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In silico identification of microRNAs predicted to regulate N-myristoyltransferase and Methionine Aminopeptidase 2 functions in cancer and infectious diseases

  • Ranjit Chauhan,

    Roles Conceptualization, Data curation, Formal analysis, Methodology, Writing – original draft, Writing – review & editing

    Current address: Molecular Virology and Hepatology Research Group, Division of BioMedical Sciences, Health Sciences Centre, Memorial University, St John's, Newfoundland and Labrador, St John’s, NL, Canada.

    Affiliation Department of Biology, University of Winnipeg, Winnipeg, Manitoba, Canada

  • David Datzkiw,

    Roles Data curation, Formal analysis, Methodology, Writing – original draft, Writing – review & editing

    Affiliation Department of Biology, University of Winnipeg, Winnipeg, Manitoba, Canada

  • Shailly Varma Shrivastav ,

    Roles Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Resources, Supervision, Writing – original draft, Writing – review & editing

    ‡ These authors are joint senior authors on this work.

    Affiliation Department of Biology, University of Winnipeg, Winnipeg, Manitoba, Canada

  • Anuraag Shrivastav

    Roles Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Writing – original draft, Writing – review & editing

    a.shrivastav@uwinnipeg.ca

    ‡ These authors are joint senior authors on this work.

    Affiliations Department of Biology, University of Winnipeg, Winnipeg, Manitoba, Canada, Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada, Research Institute of Oncology and Hematology, CancerCare Manitoba, Winnipeg, Manitoba, Canada

In silico identification of microRNAs predicted to regulate N-myristoyltransferase and Methionine Aminopeptidase 2 functions in cancer and infectious diseases

  • Ranjit Chauhan, 
  • David Datzkiw, 
  • Shailly Varma Shrivastav, 
  • Anuraag Shrivastav
PLOS
x

Abstract

Protein myristoylation is a key protein modification carried out by N-Myristoyltransferase (NMT) after Methionine aminopeptidase 2 (MetAP2) removes methionine from the amino-terminus of the target protein. Protein myristoylation by NMT augments several signaling pathways involved in a myriad of cellular processes, including developmental pathways and pathways that when dysregulated lead to cancer or immune dysfunction. The emerging evidence pointing to NMT-mediated myristoylation as a major cellular regulator underscores the importance of understanding the framework of this type of signaling event. Various studies have investigated the role that myristoylation plays in signaling dysfunction by examining differential gene or protein expression between normal and diseased states, such as cancers or following HIV-1 infection, however no study exists that addresses the role of microRNAs (miRNAs) in the regulation of myristoylation. By performing a large scale bioinformatics and functional analysis of the miRNAs that target key genes involved in myristoylation (NMT1, NMT2, MetAP2), we have narrowed down a list of promising candidates for further analysis. Our condensed panel of miRNAs identifies 35 miRNAs linked to cancer, 21 miRNAs linked to developmental and immune signaling pathways, and 14 miRNAs linked to infectious disease (primarily HIV). The miRNAs panel that was analyzed revealed several NMT-targeting mRNAs (messenger RNA) that are implicated in diseases associated with NMT signaling alteration, providing a link between the realms of miRNA and myristoylation signaling. These findings verify miRNA as an additional facet of myristoylation signaling that must be considered to gain a full perspective. This study provides the groundwork for future studies concerning NMT-transcript-binding miRNAs, and will potentially lead to the development of new diagnostic/prognostic biomarkers and therapeutic targets for several important diseases.

Introduction

The onset of carcinogenesis is initiated by mutations to begin with in a normal cell that results in the loss of growth control (hyperplasia). Hyperplasia proceeds with the loss of senescence control, replicative immortality, apoptosis resistance and the ability to evade the immune system, which are the hallmark features of cancer and are intrinsic properties in nature [1]. Later on extrinsic factors get involved where the abnormal cancerous cell begins the process of angiogenesis (acquiring blood supply) in order to thrive, evade the surrounding tissue and colonize at distal sites (metastases) through the epithelial to mesenchyme transition. The culprit for driving such traits in the journey of a normal cell to cancer is primarily dysregulation of the signaling in the abnormal microenvironment in which these cancerous cells exists, which plays an important role in escaping the immune cells responsible for policing them. A healthy microenvironment (extracellular matrix) has the ability to suppress the cancerous growth.

In order to create effective and personalized strategies for cancer treatment, it is imperative to understand the many dimensions of signaling dysregulation that characterize different types of cancer and compromised immune cells. Of the signaling molecules implicated in either immune dysfunction or cancer, N-myristoyltransferase (NMT), the enzyme responsible for the covalent attachment of a 14C myristic fatty acid to the N-terminus of target proteins, has been shown to be implicated in both the development of cancer and impaired immune cell function [26]. Myristoylation is preceded by the removal of the N-terminal methionine of the target protein by Methionine Aminopeptidase 2 (MetAP2) (and aids in protein trafficking directed to cellular membrane systems (Fig 1) [6]. Prokaryotes lack NMT, whereas lower eukaryotes like protozoans and fungus have a single copy of NMT, and mammals and other vertebrates, NMT has been shown to be present in two major isoforms, NMT1 and NMT2, which catalyze the same reaction, and are coded respectively by different genes [7, 8]. The biological role of NMT serves as a promising candidate to study with regards to cancer progression and immune function as its dysregulation has been shown to contribute to defective embryo and monocyte development, cell growth, T-cell signaling, and HIV infection. Little is known about the regulation of its expression, signaling, and localization [4, 5, 810].

The methionine residue of a nascent polypeptide is removed co-translationally by MetAP2, which is followed by addition of a myristoyl group to the exposed N-terminal glycine residue by NMT.

We are increasingly aware of the multitude of complex and intertwined signaling events occurring within a cell under various stimuli and stresses, and the many forms of regulation that govern and balance them. These events include functions prone to driving cancer, such as cell proliferation, mitosis and migration, and the respective checks in place to limit growth. At the heart of cell signaling and regulation is the myriad of genes that make up the genome, and their respective protein end products that serve to exert specific cellular functions. However, the human genome, once estimated to contain over 100, 000 protein coding genes, has been re-evaluated over the years to be comprised of merely ~20, 000 distinct genes, which in turn code for an estimated 293, 000 non-redundant peptides [11, 12]. This revelation has led researchers to seek and shed light on the various forms of genomic regulation that influence gene expression, fine-tune spatio-temporal aspects of cell signaling, and account for the massive repertoire of distinct peptides. To date, several regulatory systems have been identified and well established, including transcriptional regulation through various pleiotropic transcription-factor family proteins, and epigenetic methylation of DNA, as well as post-translational mechanisms of gene regulation, such as increased proteome diversity through mRNA splicing mediated by the spliceosome, and gene downregulation via mRNA silencing by various RNA products [1316]. Of the various agents responsible for RNA silencing, there has been emerging evidence pointing to the importance of microRNAs (miRNAs) in several facets of cellular functional regulation, including those involved in cancer progression and immune function [1719].

MicroRNAs (miRNAs) are a family of endogenously expressed small non-coding single stranded RNAs that are generally 21–23 nucleotides in length [20]. The maturation of miRNA in humans is facilitated by two consecutive cleavages mediated by RNAIII enzymes DROSHA and DICER. miRNAs are the final product of larger pri-mRNAs, the latter transcribed by either RNA polymerase II or III. DROSHA complexes along with its partner DiGeorge syndrome critical region 8 (DGCR8) in a stoichiometry of 1:2 respectively form the microprocessor complex. The microprocessor complex reduces the pri-mRNA to an ~85 nucleotide long pre-miRNA containing a hairpin loop [21, 22]. Pre-miRNA is transported into the cytoplasm, recruited to the spliceosome, and subsequently cleaved by the DICER enzyme to yield a mature miRNA duplex [23]. One of the strands of the cytoplasmic miRNA, known as guide is used to target miRNA onto a protein named Argonaute to form the RNA-induced silencing complex (RISC). RISC is a multi-protein complex that is guided by the sequence of miRNA transcript to a target (through complementary base pairs) and the protein Argonaute cleaves mRNA (messenger RNA) [24]. Thus, miRNA is capable of causing degradation of the target mRNA if perfect nucleotide complementation is achieved, otherwise translational repression of the target mRNA occurs in the case of imperfect complementarity [25]. miRNA have been shown to have a certain degree of genomic organization, adding an additional layer of complexity to miRNA systems that can be manipulated to drive evolution and specialization [26]. Some miRNAs have been shown to form polycistronic clusters that in some cases co-express several miRNAs that target different mRNAs responsible for proteins within the same protein complex [27, 28]. These findings demonstrate the ability of miRNA to influence protein-protein interactions.

The repertoire of miRNA that can be expressed by a cell constitutes an essential layer of post-transcriptional gene control for many cellular processes. miRNAs are relatively young on the evolutionary timescale, being expressed only in animals, plants, and some viruses [2931]. Around 30% of the Homo sapiens protein-coding genes are regulated by miRNAs, which control the genes at the post-transcriptional level [32]. There are two modes by which miRNAs regulates the expression of genes; first, miRNA-mediated transcript degradation and second, inhibition of protein translation [25, 33]. For target degradation model, miRNA binds predominantly to the target sequence found within the 3’ untranslated region (UTR) of the target mRNA with perfect complementarity, leading the mRNA to be cleaved [34, 35]. Similarly to inhibit the translation of target genes, miRNA binds with imperfect complementarity with the target. However, recent studies suggest that even with the imperfect complementarity between miRNA and target mRNA sequences, miRNAs are capable of carrying out target recognition and subsequent translational inhibition and/or transcriptional decay [36]. In addition to its functions in post-transcriptional gene regulation, miRNAs are also known for regulating protein complexes and acting as a key-determining molecule in protein-protein interaction [27, 28].

The role of miRNA has been suggested to be more of a fine-tuning mechanism of gene regulation, rather than as a master regulator; however, increasing evidence has shown that miRNAs are heavily dysregulated in many diseases, including cancer [3740]. There are several studies linking microRNA as a driving factor in the progression of some cancers, such as the promotion of colorectal cancer proliferation and invasion by miR-320b [41]. In some cases miRNA can act as a tumor suppressor, such as miR-29c, which is correlated with breast cancer survival and downregulates B7-H3 protein which is associated with metastasis and poor prognosis in breast cancer patients [42]. Beyond the potential of miRNAs to play a positive or negative role in disease, they may serve to act as novel biomarkers [43]. Increasing evidence is revealing that specific circulating miRNAs may be used as non-invasive biomarkers for neoplastic diseases, such as breast cancer (miR-29c, 199a, 424) [44], colorectal cancer (miR-24, 320a, 423-5p) [45], and liver cancer (miR-200 family) as well as its regression (miR-199a-3p) [46, 47]. In cancer biology, miRNAs may be playing a critical role by modulating key signaling pathways, as they have been shown to affect the sensitivity of a cell to signal transduction by signaling molecules such as epidermal growth factor, Notch, TGF- β, and WNT [48, 49].

We suspect that the role of critical signaling modulation by miRNAs can be extended to their interactions with NMT translation and influence over protein-protein interaction related to NMT signaling. Dysregulation of NMT1 activity is implicated in cancer and stem cell differentiation [8, 50]. The plasticity of transition from normal to cancerous cells as well as stem cell differentiation and proliferation to T lymphocytes depend on the miRNA target gene regulation [4, 51]. In this study we predicted the miRNAs that target NMT1/NMT2/MetAP2 transcripts, and the possibility of these interactions to inhibit NMT1/2 expression, and their function in relation to cancer, stem cell, T-cell/B-cell signaling and infectious diseases, using the bioinformatics techniques TargetScan 7 and DIANA.

Materials and methods

Sequence selection for computational analysis

For NMT1 and NMT2 reference sequences were retrieved from the GenBank. NMT1 and NMT2 reference numbers were NM_021079.4 and NM_004808, respectively. For MetAP2, the reference sequence used was NM_001317182.1.

Prediction of miRNA targets

The putative miRNA targets were predicted using two annotation programs as described previously [52]. Prediction of miRNAs was done in the subtractive library. The sequences were submitted for in silico annotation of ncRNAs. During the annotation process, we searched for the RNA structures by using Infernal (INFERence of RNA ALignment) software as described in other study [53]. The BLAST program was used to search similar miRNA sequences in the National Center for Biotechnology Information (NCBI) database as described elsewhere [54]. Finally, miRNAs with the best p-value (≤0.05) were selected for further analysis and their details are presented in tabulated form (S1 Table).

Identification and functional annotation of miRNAs regulating NMT and MetAP2 genes using TargetScan and DIANA Tools mirPath

Sequences for human NMT1/2 and MetAP2 genes (18 and 11 transcripts respectively) were downloaded from the ENSEMBL genome browser (NMT: ENSG00000136448, MetAP2: ENSG00000111142). Sequences of human mature miRNAs (2,588) were downloaded from miRBase version 21 (http://mirdb.org) [55]. Normalized mRNA and miRNA expression values were downloaded from Gene Expression Omnibus (GEO) repository (accession ID: GSE62030). miRNAs targeting NMT1/2 and MetAP2 genes were predicted by TargetScan7 (v7.0; targetscan.org) [56]. To validate the predicted miRNA: target interactions, Pearson correlation coefficient (PCC) was calculated using the normalized expression values of miRNA and target genes. All the miRNA:target interactions having significantly high inverse/negative PCC (r < -0.5, p ≤ 0.05) were considered as true miRNA:target interactions. The p-value of PCC was calculated by Student’s t-test using “R” software. Biological pathways influenced by miRNAs targeting NMT and METAP2 genes were identified using DIANA Tools mirPath version 3 (v3.0) (http://www.microrna.gr/miRPathv3) [57].

Functional enrichment analysis of miRNA targets

To determine or predict the function(s) of miRNAs which targets these genes, pathway enrichment analysis was performed using DIANA Tools mirPath (v3.0) as described previously [57]. This tool provides information on experimentally-supported miRNA functional annotation using Gene Ontology (GO) or GOSlim terms [58], combined with statistically-enriched pathways, mainly Kyoto Encyclopedia of Genes and Genomes (KEGG) molecular pathways, and based on target genes which query miRNAs targets [57].

Literature search for miRNAs associated with cancer and signaling pathways

The chosen set of miRNAs from bioinformatics analyses were thoroughly researched in the literature for their association, functions and expression changes in cancer and signaling pathways. PubMed, PubMed Central and Google Scholar search engines were utilized to perform the literature survey. MiRNAs that showed at least one publication record were considered for further analysis in the study.

MiRNA clustering analysis

MiRNA expression values were extracted from “GSE62037” GEO dataset [59]. The miRNAs that were predicted to target NMT1/2 transcripts were filtered out and their expression values were specifically extracted from the above dataset. In order to recognize the normalized expression patterns of these miRNAs, an unsupervised hierarchical cluster analysis were carried out using Cluster v3.0 as described elsewhere [60]. TreeView software was used to generate and visualize the heatmaps. Green color shows positive PCC values (0.5 ≤ r ≤ 1) and red color shows inverse/negative PCC values (-0.5 ≤ r ≤ -1). Thereafter, it was determined whether the miRNA data fit into known post-transcriptional ‘RNA regulon (operon) model’ which describes how RNA molecules are organized at a higher-organization level and how their functional dynamics are connected to post-transcriptional regulatory events such as stability and translation [61, 62].

Results

Identification of miRNAs targeting NMT and METAP2 genes

Mature miRNAs targeting 18 transcripts and 11 transcripts of NMT1/2 and MetAP2 respectively, were identified using TargetScan7 [63]. Using the stringent cutoff, a total of 13,798 miRNA-target interactions were predicted by TargetScan for the NMT1/2 genes. In contrast, 7,708 interactions were predicted for the MetAP2 gene.

In order to filter out false positive miRNA: target interactions, Pearson correlation coefficient (PCC) was calculated between targeting miRNA and target gene expression. In general, miRNAs down-regulates the expression of a target gene. Based on previous reports, the threshold PCC for determining the true positive miRNA: target interaction was set at 0.5 [64, 65]. Thus, a true interaction will be indicated by significantly high inverse PCC (r ≤ -0.5, p ≤ -1). Out of 13,798 putative miRNAs: target interactions predicted for the NMT1/2 genes, only 221 miRNAs: target interactions showed r ≤ -0.5 (S1 Table). The top five miRNAs targeting the NMT1/2 genes with the highest inverse PCC were miR-421, miR-4317, miR-606, miR-140-5p and miR-941. Interestingly, these miRNAs were also found to be regulating multiple NMT1/2 transcripts (S1 Table). Similarly, for the METAP2 gene, out of 7,708 interactions predicted, 165 miRNAs: target interactions showed PCC values above our cutoff threshold (r ≤ -0.5) (S2 Table). Based on the PCC values the top five miRNAs targeting the MetAP2 gene turned out to be miR-330-3p, miR-421, miR-409-3p, miR-139-3p, and miR-1246 (S2 Table).

Next, to determine whether same miRNA targets NMT1/2 and MetAP2 genes, we compared the targeting miRNAs lists (S1 and S2 Tables). The results revealed that 7 miRNAs are common which targets NMT1/2 and MetAP2 genes while 20 miRNAs were found to be exclusively targeting either NMT1/2 or MetAP2 genes (Table 1; Fig 2). From the list of commonly targeting miRNAs, miR-421 has the highest inverse co-expression with the target genes and appeared in the top five miRNAs. Making this miRNA a top candidate for further evaluation.

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Fig 2. The distribution of microRNAs (miRNAs) that target N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

The miRNAs that target either NMT or MetAP2, or both genes are illustrated in the Venn diagram. Seven miRNAs target both NMT1/2 and MetAP2 genes.

https://doi.org/10.1371/journal.pone.0194612.g002

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Table 1. MicroRNAs (miRNAs) that are specific and common to N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

https://doi.org/10.1371/journal.pone.0194612.t001

Functional enrichment analysis of miRNA targets

To identify the biological pathways under regulation of the miRNAs identified in the previous section, functional enrichment was done using DIANA Tools mirPath version 3 [57]. The statistically significant KEGG pathways enriched from the analyses are summarized in Table 2. The analysis revealed 88 pathways that belong under the KEGG pathways. It was found that the most significantly enriched pathway regulated by miRNAs that target both NMT1/2 genes and the MetAP2 gene was proteoglycans in cancer. Interestingly, one pathway regulated by the selected miRNAs is the ErbB signaling pathway. This pathway plays an important role in regulating cancer [66]. Furthermore, Wnt, mTOR, and VEGF signaling pathways were found to be regulated by miRNAs that target NMT gene. Additionally, we also predicted KEGG pathways that are associated with the miRNAs that exclusively targets NMT1/2 or MetAP2 genes. From these pathways, the most significant ones were filtered for further exploration in terms of their relevance in cancer.

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Table 2. KEGG pathways enrichment annotation of the microRNAs (miRNAs) that target N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

https://doi.org/10.1371/journal.pone.0194612.t002

The DIANA Tools mirPath analysis predicted that most significantly enriched KEGG pathway regulated by miRNAs targeting both NMT and MetAP2 genes was ‘ErbB signaling pathway’ (p = 3.60E-29) which involved 48 genes and 17 miRNAs (Table 2; Fig 3). ErbB receptor molecules regulate cell proliferation, differentiation, cell motility, and cell survival. Therefore, ErbB receptor mutations or overexpression have been associated with cancer cell migration, development, invasion and progression of cancers such as non-small cell lung cancer [67], breast cancer [68], ovarian cancer and bladder cancer [69]. This is mainly due to the role of this pathway in phosphorylation of many important kinases involved in cancer pathology.

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Fig 3. ErbB signaling pathway that is enriched with target genes of microRNAs (miRNAs) which negatively regulate N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

The figure illustrates ErbB signaling pathway that contain genes that are targeted by miRNAs which regulate NMT gene. (EGF, epidermal growth factor; TGF, transforming growth factor; BTC, betacellulin; HB-EGF, heparin-binding epidermal growth factor (EGF)-like growth factor; EREG, epiregulin; NRG1, neuregulin-1; NRG2, neuregulin-2; NRG3, neuregulin-3; NRG4, neuregulin-4; PLCγ, phospholipase C type gamma; CAMK2B, calcium/calmodulin dependent protein kinase; PRKCB, Protein kinase C-beta; STAT5, Signal transducer and activator of transcription 5; src, Rous sarcoma virus gene; CRK, C T10 regulator of a tyrosine kinase; NCL, NCK Adaptor Protein 2; PTK2, PTK2 protein tyrosine kinase 2; ABL2, V-Abl Abelson Murine Leukemia Viral Oncogene Homolog 2; PAK2, P21 (RAC1) Activated Kinase 2; MAP2K4, Mitogen-Activated Protein Kinase Kinase 4; MAPK10, Mitogen-Activated Protein Kinase 10; SOS1, SOS Ras/Rac Guanine Nucleotide Exchange Factor 1; Grb2, Growth Factor Receptor Bound Protein 2; SHC4, Src Homology 2 Domain-Containing-Transforming Protein C4; PIK3C4, Phosphatidylinositol-4,5-Bisphosphate 3-Kinase Catalytic Subunit; AKT3, KT Serine/Threonine Kinase 3; mTOR, Mechanistic Target Of Rapamycin Kinase; BCL2, BCL2 Associated Agonist Of Cell Death; GSK3B, Glycogen Synthase Kinase 3 Beta; CDKN1A, Cyclin Dependent Kinase Inhibitor 1A; EIF4EBP1, Eukaryotic Translation Initiation Factor 4E Binding Protein 1; BRAF, B-Raf Proto-Oncogene, Serine/Threonine Kinase; RPS6KB1, Ribosomal Protein S6 Kinase B1; KRAS, KRAS Proto-Oncogene, GTPase; JUN, Jun Proto-Oncogene, AP-1 Transcription Factor Subunit; ELK, ETS Transcription Factor; Myc, MYC Proto-Oncogene, BHLH Transcription Factor; ER, endoplasmic reticulum. DNA, deoxyribonucleic acid).

https://doi.org/10.1371/journal.pone.0194612.g003

When analyzing pathways associated with colorectal (Fig 4A) and prostate cancers (Fig 4B), we observed that miRNAs targeting both NMT1/2 and MetAP2 genes also regulate the expression of key genes involved in these pathways, including AKT1, GSK3B, BRAF, MAPK and many others (Fig 4).

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Fig 4. Colorectal cancer and prostate cancer pathways that are enriched with target genes of microRNAs (miRNAs) which negatively regulate N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

The figure illustrates A) colorectal cancer pathway and B) prostate cancer pathway that contain genes that are targeted by miRNAs which regulate NMT gene. (Rac1, Rac family small GTPase 1; Wnt, Wingless-related integration site; FOS, Fos Proto-Oncogene, AP-1 Transcription Factor Subunit; APC, Adenomatosis Polyposis Coli Tumor Suppressor; AXIN2, Axis Inhibition Protein 2; CTNNB1, Catenin Beta 1; CASP3, Caspase 3; APPL, Adaptor Protein, Phosphotyrosine Interacting With PH Domain And Leucine Zipper 1; CCND1, Cyclin D1; RALGDS, Ral Guanine nucleotide dissociation stimulator; MSH6, MutS homolog 6; BCL2, B-Cell CLL/Lymphoma 2; TGFBR2, Transforming Growth Factor Beta Receptor 2; SMAD, SMAD Family Member 3 (mothers against decapentaplegic); Myc, MYC Proto-Oncogene; BAX, BCL2 Associated X, Apoptosis Regulator; CYCS, Cytochrome C, DCC, Deleted In Colorectal Carcinoma; LEF, lymphoid enhancer binding factor 1; P53, Phosphoprotein-53. AR, androgen receptor; FOXO1, Forkhead Box O1; PTEN, Phosphatase And Tensin Homolog; GSTP1, Glutathione S-Transferase Pi 1; CDK2, Cyclin Dependent Kinase 2; Rb1, Retinoblastoma 1; E2F3, E2F Transcription Factor 3; EGF, Epidermal Growth Factor; SRD5A2, Steroid 5 Alpha-Reductase 2; PSA, Kallikrein 3; MDM2, RING-Type E3 Ubiquitin Transferase Mdm2; PDK1, Pyruvate Dehydrogenase Kinase 1; CDKN1A, Cyclin Dependent Kinase Inhibitor 1A; CREBBP, CREB Binding Protein; CREB3L1, AMP Responsive Element Binding Protein 3 Like 1).

https://doi.org/10.1371/journal.pone.0194612.g004

Apart from cancer pathways, miRNAs were also shown to regulate genes in immune cells. Therefore, we further analyzed KEGG pathways that are associated with immune responses. For this, we performed pathway enrichment analysis using miRNAs that target only NMT transcripts. Interestingly, T cell receptor signaling pathway (p = 6.87E-09) involving 43 genes and 18 miRNAs (Table 2; Fig 5A); and B cell receptor signaling pathway (p = 9.73E-15) involving 36 genes and 17 miRNAs were predicted by the analysis (Table 2; Fig 5B). A total of 11 and 8 miRNAs that target NMT1 and NMT2, respectively were identified to be associated with T cell and B cell receptor pathways (Table 3). Among these miRNAs, miR-654 displayed an interesting relationship with NMT1/2 and MetAP2 genes where miR-654-5p targeted NMT1/2 while miR-654-3p targeted MetAP2. Similarly, miRNA-199b-5p was found to be targeting NMT1/2 while miRNA-199b-3p targeted MetAP2. This 5p and 3p pattern was not limited to these two miRNAs. Same pattern was observed with miR-628, and miRNA-139. These observations suggest that the miRNA species deriving from the 5' arm (5p) and 3' arm (3p) of the same pre-miRNA can regulate both NMT1/2 and MetAP2 genes depending upon which mature miRNA is loaded on to Argonaute protein. Additionally, miR-1246 was also common between NMT1/2 and MetAP2 genes.

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Fig 5. T-cell and B-cell receptor pathways that are enriched with target genes of microRNAs (miRNAs) which negatively regulate N-myristoyltransferase (NMT1/2) and methionine aminopeptidase 2 (MetAP2) genes.

The figure illustrates two cellular pathways A) T cell receptor signaling and B) B cell receptor signaling that contains genes that are targeted by miRNAs, which regulate NMT1/2 gene. (PD1, programmed cell death-1; ZAP70, Zeta Chain Of T-Cell Receptor Associated Protein Kinase 70; LAT, Linker for Activation Of T-Cells; ICOS, Inducible T-Cell Co-stimulator; DLG1, Discs Large MAGUK Scaffold Protein 1; NCK2, NCK Adaptor Protein 2; LCP2, Lymphocyte Cytosolic Protein 2; GM-CSF, Granulocyte-macrophage colony-stimulating factor; IFN-γ, Interferon gamma; TNFα, Tumor Necrosis Factor alpha; IL, Interleukin; NFKB1, Nuclear Factor Kappa B Subunit 1; CDK4, Cyclin Dependent Kinase 4; PPP3R2, Protein Phosphatase 3 Regulatory Subunit B, Beta; CTLA4, Cytotoxic T-lymphocyte Associated Protein; PKC8, protein kinase C-8; FYN, FYN Proto-Oncogene, Src Family Tyrosine Kinase; Raf-1, Raf-1 Proto-Oncogene, Serine/Threonine Kinase. CD, cluster of differentiation; BCR, break-point cluster region; BTK, Bruton Tyrosine Kinase; DAPP1, Dual Adaptor Of Phosphotyrosine And 3-Phosphoinositides 1; VAV3, Vav Guanine Nucleotide Exchange Factor 3; SHIP, SH2 Domain-Containing Inositol 5-Phosphatase; Rac1, Rac Family Small GTPase 1; SYK, Spleen Associated Tyrosine Kinase; NFATC1, Nuclear Factor of activated T-cells 1; PPP3R2, Protein Phosphatase 3 Regulatory Subunit B, Beta; RASGRP3, RAS Guanyl Releasing Protein 3; MALT1, Mucosa Associated Lymphoid Tissue Lymphoma Translocation; Lyn, LYN Proto-oncogene src family tyrosine kinase).

https://doi.org/10.1371/journal.pone.0194612.g005

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Table 3. MicroRNAs (miRNAs) that target T cell and B cell signaling pathways and stem cell signaling.

https://doi.org/10.1371/journal.pone.0194612.t003

Further analysis of the KEGG pathways enriched with miRNAs that targets only NMT1/2 gene transcripts revealed signaling pathways that regulate the pluripotency of stem cells, Hippo signaling pathway, estrogen signaling pathway and glioma. On the other hand, a similar analysis for miRNAs that target MetAP2 gene revealed regulation of fatty acid biosynthesis, signaling pathways regulating pluripotency of stem cells, ErbB signaling pathway and prion diseases as the top KEGG pathways involved. Interestingly, all these KEGG pathways are related to the genes/proteins that regulate cellular growth, survival, migration, proliferation and development, which are basically essential cellular processes involved in cancer progression [62]. Taken together, the predicted data strongly support the association of miRNAs targeting NMT1/2 and MetAP2 genes to signaling pathways implicated in cancer and immune response.

Further analysis of signaling networks

Cancer signaling.

We mined data to identify the roles of 221 miRNAs that regulate NMT in cancer. We found that 35 (15.8%) miRNAs were found to have a clear role in cancer. Of these filtered out 35 miRNAs, 15 miRNAs (42.8%) target NMT1 transcript, while 20 miRNAs (57%) target the NMT2 transcripts. These miRNAs are listed in Table 4. Furthermore, out of 35 miRNAs 14 (40%) miRNAs were found to be associated with prostate cancer whereas 4 (11.4%) miRNAs showed association with colorectal, breast and liver cancers. Additionally, 3/35 (8.5%) miRNAs were found to have roles in esophageal, renal and squamous cell carcinomas.

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Table 4. The association of microRNAs (miRNAs) that target N-myristoyltransferase (NMT) transcript 1 and 2 (NMT1 and NMT2) with different cancer types and their expression changes.

https://doi.org/10.1371/journal.pone.0194612.t004

Stem cell, T cell and B cell receptor signaling.

A thorough search of the identified 221 miRNAs revealed a defined role for 21 (9.5%) miRNAs in TCR, BCR and stem cell signaling. Of the 21 miRNAs, 13 and 8 miRNAs contained complementarity with NMT1 and NMT2 transcripts’ 3’UTR, respectively. The differential regulation of miRNAs that are associated with signaling pathways are summarized in Table 3 and may provide insights on how differential expression of miRNAs may impact NMT levels.

Infectious diseases

We found 14 miRNAs that were associated with functions in infectious diseases, 8 of which target NMT1/2 genes and 6 miRNAs target MetAP2 gene. Of the total 14 miRNAs targeting either NMT1/2 genes or MetAP gene, 13 miRNAs (miR-29a-5p, miR-132-5p, miR-134-5p, miR-137, miR-139-3p, miR-140-5p, miR-199a-3p, mir-520d-5p, miR-548p, miR-943, miR-4317 and miR-628-3p) have been found to have roles in viral infections and 4 miRNAs (miR-127-3p, miR-140-5p and miR-199a-3p and miR-4317) have been implicated in bacterial infections. Importantly, two miRNAs: miR-199a-3p and miR-4317 play a role in both viral and bacterial infections. Furthermore, two miRNAs (miR-132-5p and miR-140-5p), have roles in both viral and fungal infections.

HIV infection.

The NMT1/2 targeting miR-137 and MetAP2 targeting miR-199a-3p were found to be upregulated in HIV infection, while another miRNA-324-5p, which targets NMT1/2 genes, also modulates HIV infection by targeting viral infectivity factor gene.

Hepatitis virus infection.

Four miRNAs were found to be associated with hepatitis viral infections namely miR-29a-5p, miR-548p, miR-199a-3p and miR-520d-5p. Of these four miRNAs, three miRNAs (miR-29a-5p, miR-199a-3p, miR-520d-5p) targeted MetAP2 whereas, the fourth miR-548p targeted NMT1/2 genes.

Human Papilloma Virus (HPV) infection.

The two miRNAs (miR-324-5p and miR-139-3p) that were found to be associated with HPV infection also targeted NMT1/2 and MetAP2 genes. Human Papillomavirus subtype-16 E5 protein downregulates miR-324-5p and contrary to this miR-139-3p is downregulated in HPV-16-induced carcinomas.

Fungal infection.

The NMT1/2 and MetAP2 genes’ targeting miR-132-5p was found to be associated with fungal infection of human dendritic cells with Candida albicans and Aspergillus fumigatus and regulates immune responses through the interactions with FKBP1B, KLF4, and SPN genes. Furthermore, miR-140, miR-628-3p and miR-943 miRNAs that target NMT1/2 and MetAP2 genes were found to have roles in infectious diseases. Of which miR-943 is upregulated in Herpes Zoster viral infection and miR-628-3p serves as a biomarker for the enterovirus 71 infection. The variable expression of miRNAs associated with infectious diseases is summarized in Table 5 with their corresponding references.

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Table 5. The association of microRNAs (miRNAs) that target N-myristoyltransferase (NMT) transcript 1 and 2 (NMT1 and NMT2) and MetAP transcripts with different infectious diseases and their expression changes.

https://doi.org/10.1371/journal.pone.0194612.t005

Targeting miRNAs of NMT follows RNA Regulon model

The miRNAs targeting NMT1/2 genes were clustered together based on their expression, which is supported by a previous report proposing that miRNAs performing similar functions usually co-express with each other [61]. This phenomenon of miRNA regulon was previously observed between miRNAs regulated by same RNA Binding Proteins [144]. We deemed it worth to identify whether NMT1/2 and MetAP2 gene expression followed a pattern indicative of a miRNA regulon. To find an active regulon model, PCC between miRNAs were calculated based on the expression values and miRNAs were clustered based on the correlation value. It was observed that miR-4306 share highest co-expression with miR-1244 followed by miR-3179 (Fig 6). This regulon model suggests that some common biological pathway is regulated by these miRNAs apart from targeting of NMT1/2 genes. Our data supports this notion as many of the miRNAs detected in this study were found to be involved in targeting genes involved in same pathways. Most of these miRNAs co-express with each other indicating miRNA regulon model is similar to the RNA regulon model, which needs further evaluation.

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Fig 6. The clustering of microRNAs (miRNAs) targeting N-myristoyltransferase (NMT) genes.

The figure illustrates clustering of the miRNAs that target NMT1/2 genes based on normalized expression values obtained from “GSE62037”. Data suggest miRNA regulon model is similar to the RNA regulon model. Green color shows positive Pearson correlation coefficient (PCC) values (0.5 ≤ r ≤ 1) and red color shows inverse/negative PCC values (-0.5 ≤ r ≤ 1). Heatmap shows that most of miRNAs have high PCC values, and these miRNAs co-express with each other to regulate common biological processes.

https://doi.org/10.1371/journal.pone.0194612.g006

Discussion

Understanding miRNA-mRNA interactions that play role in tumor development, cancer progression and associated cellular processes such as immune responses is a critical step necessary to use miRNAs as therapeutic or diagnostic targets for cancer and other diseases. miRNAs are emerging as a crucial component in the regulation of cellular signaling pathways, including several processes involved in diseases such as cancer and immune dysfunction. Investigating miRNA function and expression patterns is necessary for fully understanding how miRNAs fit to interact with more well studied mechanisms of cell regulation, giving us a bigger and clearer picture of the signaling framework of cells under various conditions. Specifically, studying miRNA interactions with the NMT1/2 and MetAP2 genes will shed light on the poorly understood regulation of myristoylation, a key cellular function critical to oncogenesis, adaptive immune function, and infectious disease onset. Our findings have verified that miRNAs are likely an important driving force in the regulation of myristoylation, and identified several NMT-transcript-binding miRNAs that have been linked to processes mediated by myristoylation.

The present study predicted (1) miRNAs that significantly bind to the NMT1 and NMT2 transcripts, (2) miRNAs in which NMT1/2 and MetAP2 transcripts are common binding targets, (3) a possible role for the involvement of NMT1/2 transcript-binding miRNAs in Cancer, T/B cell receptor signaling and infectious diseases. By using TargetScan software, we initially found a total of 13,798 miRNA-target hits for the NMT1/2 genes, whereas the MetAP2 gene had 7,708 interactions detected. Statistical analysis using “R” software package further delineated significant binding of these miRNAs onto their target; based on this, 221 NMT miRNA-NMT-target interactions, and 165 miRNA-METAP2-target interactions were verified. A thorough literature review was performed to find if these miRNAs have any role in Cancer, T/B cell receptor signaling, and infectious diseases. Of the 35 miRNAs, 15 (42.8%) target the NMT1 transcript and 20 (57%) target the NMT2 transcripts. Interestingly, 14 miRNAs (40%) have a defined role in prostate cancer, and 4 (11.4%) miRNAs each that play a role in colorectal, breast and liver cancers. Furthermore 3 (8.5%) miRNAs are involved in esophageal cancer, and one miRNA each that has been detected to play a role in renal cell carcinoma, brain cancer, and role in squamous cell carcinomas. Secondly, with regards to T/B cell receptor signaling, 21 (9.5%) NMT1/2 targeting miRNAs were detected. Of the 21 miRNAs, 13 had NMT1 as their target site and 8 had NMT2 as their target site. Lastly, 2 of each of the miRNAs targeting NMT1/2 were involved in infectious diseases, including HIV.

Numerous recent reports clearly point to a role of miRNA in cancer establishment, evasion, differentiation, and metastasis, which when targeted, have shown the promising results as therapeutic targets in preclinical as well as in clinical trials [145, 146]. Our findings indicate that several miRNAs that target myristoylation related genes (NMT1/2 and MetAP2) are associated with cancer. Interestingly, of these identified miRNAs, some have been found to be associated with colorectal or breast cancers wherein role of NMT has already been established [2, 50]. Furthermore, miR-409-3p and miR-106b-3p, and miR-127 that all bind to the NMT1 transcript, have been reportedly involved in colorectal cancer and breast cancer respectively. miR-409-3p was previously found to be a colorectal tumor suppressor, with reduced expression of miR-409p present in colorectal cancer tissue and correlated with metastasis [94]. In contrast, miR-106b-3p upregulation was correlated with colorectal tumor growth [147]. Wang et al demonstrated that miR-127 is downregulated in breast cancer tissues, and reduced miR-127 expression is correlated with late stage lymph node metastasis. Furthermore, miR-127 downregulation was an independent prognostic factor that predicted lower overall survival in breast cancer patients and upregulation of miR-127 inhibited breast cancer cells growth, survival, and migration [148].

We also identified NMT2-targeting miRNAs that have been previously demonstrated to play a role in regulating or advancing colorectal and breast cancers. Our analysis identified miR-133a and miR-301a as NMT2-binding miRNAs that act to suppress and promote colorectal cancer respectively. Wan et al established through gene analysis that miR-133a expression was reduced in 83.2% of colorectal cancer patient tumors compared to healthy mucosal tissue, and Dong et al demonstrated that ectopic overexpression of miR-133a suppresses colorectal tumor growth in-vitro and in-vivo [112, 149]. A study into the colorectal cancer promoting mechanism of miR-301a, which is upregulated in colorectal cancers, revealed that miR-301a expression confers growth and invasion by downregulating SOCS6 expression [150]. Our investigation of NMT2 transcript-binding miRNAs revealed miR-411-5p and miR-187, which respectively function as a tumor suppressor and an oncogenic agent of breast cancer. miR-411-5p was observed to suppress breast cancer by downregulating GRB2 and Ras expression [114]. Finally, a report by Mulrane et al identified miR-187 expression as an independent factor that drives the in vitro development of increased aggressiveness and invasiveness of breast cancer [113].

There are several lines of evidence that demonstrate the critical role miRNAs play in stem cell, T-cell, and B-cell signaling [151154]. This is of relevance to this study as besides cancer, studies on myristoylation signaling have focused on NMT’s role with respect to embryogenesis, innate immune cell differentiation, and adaptive immune development and signaling, and HIV infection [4, 5, 8, 10]. During our analysis we identified several NMT1/2-binding-miRNAs involved in the previously mentioned functions and have chosen to discuss a selection of the most relevant and best-annotated examples. With regards to certain developmental pathways, miR-200, 199b and 26a have been shown to play a decisive role, with the former binding to the NMT1 transcript, and latter two binding to the NMT2 transcript. miR-200 family microRNAs were found to be expressed exclusively in epithelial type tissues and when downregulated could stimulate an epithelial-mesenchymal transition, which may implicate their reduced activity with cancer metastasis [155]. miR-199b has been shown to block stem cell differentiation through inhibition of the Notch pathway [81]. miR-26a was identified as a required factor for the differentiation of skeletal muscle during mouse development, with inhibition of miR-26a resulting in delayed muscle regeneration [156].

In terms of immune function, the NMT2-transcript binding miR-628 was found to be upregulated following TLR mediated LPS detection in monocytes, and targeted MyD88 as part of a negative feedback loop for TLR signaling [157]. Two NMT1-transcript-binding miRNAs, miR-29b and 326, have evident roles in T-cell signaling. miR-29b overexpression in CD8+ T cells of renal carcinoma patients has been found to induce immune dysfunction through down-regulation of JAK3 and MCL-1 [158]. Furthermore, heightened expression of miR-326, which is associated with multiple sclerosis (MS), was shown to drive TH-17 T helper cell differentiation and MS pathogenesis in mouse models [78]. Two mRNA hits during our analysis were functionally implicated in the regulation of B-cell signaling, specifically with regards to the autoimmune disease lupus. The B-cells of lupus patients were determined to have increased expression of miR-148, an NMT1-transcript-binding miRNA, and decreased expression of miR-1246, an NMT2-transcript-binding miRNA. miR-148a is thought to affect B cell signaling by impairing immune tolerance pathways through suppression of PTEN, thus contributing to autoimmune symptoms [159]. In contrast, expression of miR-1246 normally regulated B cell activation through suppression of EBF1, however miR-1246 expression is reduced in the activated B cells of lupus patients through Akt-p53 signaling [160]. Lastly, miR-132 (NMT1-transcript-binding miRNA) and miR-29 (NMT2-transcript-binding miRNA) are involved in HIV infection, with the former promoting infection and the latter acting as an antiviral agent. miR-132 is highly upregulated during CD4+ T cell activation and are thought to enhance HIV-1 replication [161]; miR-29 family miRNAs are expressed by increased STAT3 transcription following IL-21 stimulation and lead to an anti-viral environment in CD4+ T cells during initial control of HIV-1 in vivo [162].

The overlap between the effects of NMT-transcript-binding miRNAs and previously reported NMT dysfunction are encouraging for the future development of therapeutics or diagnostic/prognostic biomarkers that exploit myristoylation-mediated cell signaling. The high stability of miRNAs have made them a promising biomarkers as they can be easily obtained from the buffy coat associated immune cells of blood samples [163, 164]. Additionally, methods of in vivo miRNA modulation are being developed for future clinical settings [165].

A recent report by Chen et al. documented dynamic interaction of miR-99 and miR-100 and the ability of miR-100 to downregulate NMT1 transcript levels [166]. Several other recent studies demonstrated that miR-99 modulates many molecular signaling pathways that are not limited to AKT, mTOR, MMP1 and IGFR1 signaling and contribute to the tumorigenesis of cancer conditions such as head and neck squamous cell carcinoma [167], oral squamous cell carcinoma [168, 169] and esophageal squamous cell carcinoma [169]. Interestingly, one functional study indeed proved that miR-99a inhibits cell proliferation, colony formation ability, migration and invasion by targeting fibroblast growth factor receptor 3 (FGFR3) in prostate cancer [170] and bladder cancer [171]. A study by Androulidaki et al. demonstrated that AKT1 controls macrophage responses to the LPS by regulation miRNA [172]. All these studies provide supporting evidence that miR-99 is a critical player in cancer pathogenesis, which is possibly derived from post-transcriptional regulation of NMT. Study by Schramedei et al. demonstrated that MetAP2 is a putative target of miR-21 in B-cell lymphomas [173].

In addition to prediction of the nature of miRNA-mRNA relationships, our study also revealed another layer of complexity and interconnectedness of miRNAs, long noncoding RNAs (lncRNA) and target mRNAs. The NMT1 targeting miR-675-5p, which is down regulated in metastatic prostate cancer, derives from the lncRNA H19 [91]. In prostate cancer cells, H19 is upregulated that aids in expression of miR-675. The expression of lncRNA H19 and miR-675 were associated with repression of extracellular matrix protein, TGFβ1, that regulate cellular migration and cancer metastasis. It appears that the H19-miR-675 lncRNA-miRNA interactions function in different cancer cell types by targeting different mRNAs. For instance, in gastric cancer, H19-miR-675 regulates cell proliferation by repressing RUNX1 [174].

One of the bottlenecks in HIV infection is the neurodegeneration, which occurs through modulation of host axon guidance and associated neurotrophin signaling pathways. Zhou et al., comprehensively profiled miRNAs in patients with dementia who were infected with HIV. In this study, they uncovered three important miRNAs that were dysregulated particularly in HIV infected dementia patients compared to only dementia patients. The miR-137 that targets NMT1/2 and MetAP2 genes was on top of the list. Further enrichment of the miRNA pathways revealed that miR-137 is involved in more than 5 neurodegeneration pathways that included WNT and MAPK pathways, which happens to be dysregulated in cancers as well [125]. Since miR-137 targets NMT1/2 genes and particularly in dentate gyrus and hippocampus, it would be interesting to further investigate the dynamics of miR-137 and NMT in dementia patients with HIV. Hepatocellular carcinoma is a dreaded cancer of the liver whose etiology is linked to chronic liver inflammation most commonly due to infection with hepatitis B and hepatitis C viruses. Our analysis revealed four NMT1/2 and MetAP2 targeting miRNAs (miR-29a-5p, miR-548p, miR-199a-3p and miR-520d-5p) that have roles in hepatitis viral infections. miR-29a-5p is demonstrated to be upregulated in Hepatitis B Virus infection related to hepatocellular carcinoma and may function through inhibition of PTEN (131). Mounting evidences in the literature suggest that miR-199 plays dominant and multi-faceted role not only in cancer but also in infectious diseases including HBV, HCV and Schistosoma mansoni. It not only inhibits the HBV viral replication but also regresses the hepatocellular carcinoma [47, 133]. Moreover, miR-199a-3p can also be used as early biomarker in HCV infection [138]. As revealed in our analysis, since miR-199a-3p also inhibits the MetAP2 gene, which plays key role in angiogenesis, further studies are warranted to underpin the role of miR-199a-3p in oncogenesis. Apart from these miRNAs, we also found that miR-520d-5p is associated with immune clearance by transitioning the immune tolerant to immune activation state in chronic hepatitis B and miR-548p suppresses hepatitis B virus associated HCC by downregulating expression of hepatitis B x-interacting protein [128, 175].

Our in-depth analyses of miRNAs that regulate NMT1/2 and MetAP2 genes have cemented the significance of miRNAs in disease etiology and progression. The emergence of miRNAs’ abilities in regulating various important facets of cellular functions dictates the significance of miRNAs in designing new therapies around cell signaling molecules.

Several miRNAs in this study were predicted to interact with mRNA transcripts responsible for translating the key proteins involved in myristoylation (NMT1/NMT2/METAP2). This preliminary in-silico analysis requires future validation within in-vitro cell models to demonstrate expression of the identified miRNAs and their interaction with the target transcripts.

Initial validation should include quantitative PCR (qPCR) to confirm co-expression of each miRNA and their respective transcript target using miRNA and transcript specific primers. Since, in-silico study may reveal a large number of potential miRNA binding sites on a target mRNA, it is important that a reporter assay should be performed to validate specific binding sites.

The interaction between a miRNA of interest and its target(s) needs to be demonstrated, which can be done using in-situ hybridization with locked nucleic acid (LNA) modified oligonucleotide probes labelled with digoxigenin that can be visualized appropriately using digoxigenin antibody conjugated to alkaline phosphatase to act upon a chromogenic substrate. A major challenge to these initial approaches includes a potential tissue specific expression of the analyzed miRNAs, emphasizing the need to test a spectrum of tissue types and cell lines. Tissue specific expression of NMT1, NMT2 and MetAP2 is well documented and therefore, initial studies would require validation of miRNAs in cell lines and tissues that express the target genes.

Furthermore, miRNAs of interest may be overexpressed (gain-in-function) in cell lines through a lentiviral infection or plasmid transfection to determine their effect on NMT1/NMT2/METAP2 mRNA transcript levels using qPCR and the resulting protein expression by Western blot analysis. Narrowed down miRNAs from the aforementioned validation techniques will be used for further studies in order to determine their effect(s) on cell signaling pathways that are involved in regulating cell proliferation, apoptosis, T/B cell signaling and viral infection.

Conclusion

Our original data set encompassed over 13, 000 miRNAs that potentially target and regulate NMT. Together, the functional data available for NMT1/2-transcript-targeting miRNAs helped create a condensed data set of miRNAs that serve as strong candidates for further investigation into the regulation of myristoylation with respect to various types of signaling dysregulation. In total, 35 NMT-transcript-binding miRNAs were linked to cancer, 21 linked to development or immune dysfunction, and 14 linked to infectious disease, including HIV and HBV infection. Future analysis of this miRNA panel is needed to verify a direct link between these miRNAs and myristoylation dysfunction associated diseases. This study provides a platform for studying an additional facet of NMT regulation, and may lead to the development of new therapeutic targets or biomarkers for several types of cancer, development or immune dysfunction, HIV and hepatitis viral infections.

Supporting information

S1 Table. Putative miRNAs that target N-myristoyltransferase (NMT) gene.

https://doi.org/10.1371/journal.pone.0194612.s001

(DOCX)

S2 Table. Putative microRNAs (miRNAs) that target methionine aminopeptidase 2 (METAP2) gene.

https://doi.org/10.1371/journal.pone.0194612.s002

(DOC)

Acknowledgments

Authors would like to thank Dr. Ashwani Jha (Department of Biochemistry and Molecular Biology, Centre for High-Throughput Biology, University of British Columbia) for helping with target scan. AS acknowledges the funding from The University of Winnipeg and the Prostate Cancer Fight Foundation. DD acknowledges financial support from CIHR and QE2. SV acknowledges funding support from Manitoba Medical Service Foundation. RC’s position as an RA at the University of Winnipeg was supported by CFI.

References

  1. 1. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011 Mar 4;144(5):646–74. pmid:21376230. Epub 2011/03/08. Eng.
  2. 2. Shrivastav A, Varma S, Saxena A, DeCoteau J, Sharma RK. N-myristoyltransferase: a potential novel diagnostic marker for colon cancer. Journal of translational medicine. 2007;5:58. pmid:18021392. Pubmed Central PMCID: Pmc2203986. Epub 2007/11/21. eng.
  3. 3. Selvakumar P, Sharma RK. Role of calpain and caspase system in the regulation of N-myristoyltransferase in human colon cancer (Review). International journal of molecular medicine. 2007 May;19(5):823–7. PubMed pmid:17390089. Epub 2007/03/29. eng.
  4. 4. Rampoldi F, Bonrouhi M, Boehm ME, Lehmann WD, Popovic ZV, Kaden S, et al. Immunosuppression and Aberrant T Cell Development in the Absence of N-Myristoylation. Journal of immunology (Baltimore, Md: 1950). 2015 Nov 1;195(9):4228–43. pmid:26423150. Epub 2015/10/02. eng.
  5. 5. Shrivastav A, Varma S, Lawman Z, Yang SH, Ritchie SA, Bonham K, et al. Requirement of N-myristoyltransferase 1 in the development of monocytic lineage. Journal of immunology (Baltimore, Md: 1950). 2008 Jan 15;180(2):1019–28. PubMed pmid:18178842. Epub 2008/01/08. eng.
  6. 6. Towler DA, Eubanks SR, Towery DS, Adams SP, Glaser L. Amino-terminal processing of proteins by N-myristoylation. Substrate specificity of N-myristoyl transferase. The Journal of biological chemistry. 1987 Jan 25;262(3):1030–6. PubMed pmid:3100524. Epub 1987/01/25. eng.
  7. 7. Giang DK, Cravatt BF. A second mammalian N-myristoyltransferase. The Journal of biological chemistry. 1998 Mar 20;273(12):6595–8. PubMed pmid:9506952. Epub 1998/04/18. eng.
  8. 8. Yang SH, Shrivastav A, Kosinski C, Sharma RK, Chen MH, Berthiaume LG, et al. N-myristoyltransferase 1 is essential in early mouse development. The Journal of biological chemistry. 2005 May 13;280(19):18990–5. pmid:15753093. Epub 2005/03/09. eng.
  9. 9. Ducker CE, Upson JJ, French KJ, Smith CD. Two N-myristoyltransferase isozymes play unique roles in protein myristoylation, proliferation, and apoptosis. Molecular cancer research: MCR. 2005 Aug;3(8):463–76. pmid:16123142. Pubmed Central PMCID: Pmc2908404. Epub 2005/08/27. eng.
  10. 10. Li H, Dou J, Ding L, Spearman P. Myristoylation is required for human immunodeficiency virus type 1 Gag-Gag multimerization in mammalian cells. Journal of virology. 2007 Dec;81(23):12899–910. pmid:17881447. Pubmed Central PMCID: Pmc2169113. Epub 2007/09/21. eng.
  11. 11. Pertea M, Salzberg SL. Between a chicken and a grape: estimating the number of human genes. Genome Biology. 2010 05/05;11(5):206–. PubMed PMID: PMC2898077. pmid:20441615
  12. 12. Kim M-S, Pinto SM, Getnet D, Nirujogi RS, Manda SS, Chaerkady R, et al. A draft map of the human proteome. Nature. 2014;509(7502):575–81. PubMed PMID: PMC4403737. pmid:24870542
  13. 13. Iwafuchi-Doi M, Zaret KS. Pioneer transcription factors in cell reprogramming. Genes & development. 2014;28(24):2679–92. PubMed PMID: PMC4265672.
  14. 14. Han Y, Garcia BA. Combining genomic and proteomic approaches for epigenetics research. Epigenomics. 2013;5(4):439–52. PubMed PMID: PMC4055025. pmid:23895656
  15. 15. Witten JT, Ule J. Understanding splicing regulation through RNA splicing maps. Trends in Genetics. 2011;27(3–2):89–97. PubMed PMID: PMC3165201.
  16. 16. Kasai M, Kanazawa A. RNA silencing as a tool to uncover gene function and engineer novel traits in soybean. Breeding Science. 2012 02/04 07/04/received 09/14/accepted;61(5):468–79. PubMed PMID: PMC3406797. pmid:23136487
  17. 17. Chen K, Rajewsky N. The evolution of gene regulation by transcription factors and microRNAs. Nature reviews Genetics. 2007 Feb;8(2):93–103. pmid:17230196. Epub 2007/01/19. Eng.
  18. 18. Jebbawi F, Fayyad-Kazan H, Merimi M, Lewalle P, Verougstraete JC, Leo O, et al. A microRNA profile of human CD8(+) regulatory T cells and characterization of the effects of microRNAs on Treg cell-associated genes. Journal of translational medicine. 2014 Aug 06;12:218. pmid:25090912. Pubmed Central PMCID: Pmc4440568. Epub 2014/08/06. Eng.
  19. 19. O'Donnell KA, Wentzel EA, Zeller KI, Dang CV, Mendell JT. c-Myc-regulated microRNAs modulate E2F1 expression. Nature. 2005 Jun 9;435(7043):839–43. pmid:15944709. Epub 2005/06/10. Eng.
  20. 20. Zhou X, Yang PC. MicroRNA: a small molecule with a big biological impact. MicroRNA (Shariqah, United Arab Emirates). 2012;1(1):1. PubMed pmid:25048083. Epub 2012/01/01. Eng.
  21. 21. Lee Y, Kim M, Han J, Yeom KH, Lee S, Baek SH, et al. MicroRNA genes are transcribed by RNA polymerase II. The EMBO journal. 2004 Oct 13;23(20):4051–60. pmid:15372072. Pubmed Central PMCID: Pmc524334. Epub 2004/09/17. Eng.
  22. 22. Borchert GM, Lanier W, Davidson BL. RNA polymerase III transcribes human microRNAs. Nature structural & molecular biology. 2006 Dec;13(12):1097–101. pmid:17099701. Epub 2006/11/14. Eng.
  23. 23. Kurzynska-Kokorniak A, Koralewska N, Pokornowska M, Urbanowicz A, Tworak A, Mickiewicz A, et al. The many faces of Dicer: the complexity of the mechanisms regulating Dicer gene expression and enzyme activities. Nucleic acids research. 2015;43(9):4365–80. PubMed PMID: PMC4482082. pmid:25883138
  24. 24. Flores O, Kennedy EM, Skalsky RL, Cullen BR. Differential RISC association of endogenous human microRNAs predicts their inhibitory potential. Nucleic acids research. 2014;42(7):4629–39. PubMed PMID: PMC3985621. pmid:24464996
  25. 25. Valencia-Sanchez MA, Liu J, Hannon GJ, Parker R. Control of translation and mRNA degradation by miRNAs and siRNAs. Genes & development. 2006 Mar 1;20(5):515–24. pmid:16510870. Epub 2006/03/03. Eng.
  26. 26. Turner M, Yu O, Subramanian S. Genome organization and characteristics of soybean microRNAs. BMC genomics. 2012 May 04;13:169. pmid:22559273. Pubmed Central PMCID: Pmc3481472. Epub 2012/05/09. Eng.
  27. 27. Sass S, Dietmann S, Burk UC, Brabletz S, Lutter D, Kowarsch A, et al. MicroRNAs coordinately regulate protein complexes. BMC systems biology. 2011 Aug 25;5:136. pmid:21867514. Pubmed Central PMCID: Pmc3170341. Epub 2011/08/27. Eng.
  28. 28. Liang H, Li WH. MicroRNA regulation of human protein protein interaction network. RNA (New York, NY). 2007 Sep;13(9):1402–8. pmid:17652130. Pubmed Central PMCID: Pmc1950750. Epub 2007/07/27. Eng.
  29. 29. Guerra-Assuncao JA, Enright AJ. Large-scale analysis of microRNA evolution. BMC genomics. 2012 Jun 06;13:218. pmid:22672736. Pubmed Central PMCID: Pmc3497579. Epub 2012/06/08. Eng.
  30. 30. Pashkovskiy PP, Ryazansky SS. Biogenesis, evolution, and functions of plant microRNAs. Biochemistry Biokhimiia. 2013 Jun;78(6):627–37. pmid:23980889. Epub 2013/08/29. Eng.
  31. 31. Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004 Jan 23;116(2):281–97. PubMed pmid:14744438. Epub 2004/01/28. Eng.
  32. 32. Lewis BP, Burge CB, Bartel DP. Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets. Cell. 2005 Jan 14;120(1):15–20. pmid:15652477. Epub 2005/01/18. eng.
  33. 33. Pillai RS, Bhattacharyya SN, Filipowicz W. Repression of protein synthesis by miRNAs: how many mechanisms? Trends in cell biology. 2007 Mar;17(3):118–26. pmid:17197185. Epub 2007/01/02. eng.
  34. 34. Catalanotto C, Cogoni C, Zardo G. MicroRNA in Control of Gene Expression: An Overview of Nuclear Functions. Int J Mol Sci. 2016 Oct 13;17(10). pmid:27754357. Pubmed Central PMCID: Pmc5085744. Epub 2016/10/19. eng.
  35. 35. Fabian MR, Sonenberg N, Filipowicz W. Regulation of mRNA translation and stability by microRNAs. Annual review of biochemistry. 2010;79:351–79. pmid:20533884. Epub 2010/06/11. eng.
  36. 36. Martin HC, Wani S, Steptoe AL, Krishnan K, Nones K, Nourbakhsh E, et al. Imperfect centered miRNA binding sites are common and can mediate repression of target mRNAs. Genome Biol. 2014 Mar 14;15(3):R51. pmid:24629056. Pubmed Central PMCID: Pmc4053950. Epub 2014/03/19. eng.
  37. 37. Greenberg DS, Soreq H. MicroRNA therapeutics in neurological disease. Current pharmaceutical design. 2014;20(38):6022–7. PubMed pmid:24641229. Epub 2014/03/20. Eng.
  38. 38. Fisher K, Lin J. MicroRNA in inflammatory bowel disease: Translational research and clinical implication. World J Gastroenterol. 2015 Nov 21;21(43):12274–82. pmid:26604636. Pubmed Central PMCID: Pmc4649112. Epub 2015/11/26. Eng.
  39. 39. Pers YM, Jorgensen C. MicroRNA in 2012: Biotherapeutic potential of microRNAs in rheumatic diseases. Nature reviews Rheumatology. 2013 Feb;9(2):76–8. pmid:23321608. Epub 2013/01/17. Eng.
  40. 40. Shrestha S, Hsu SD, Huang WY, Huang HY, Chen W, Weng SL, et al. A systematic review of microRNA expression profiling studies in human gastric cancer. Cancer medicine. 2014 Aug;3(4):878–88. pmid:24902858. Pubmed Central PMCID: Pmc4303155. Epub 2014/06/07. Eng.
  41. 41. Zhou J, Zhang M, Huang Y, Feng L, Chen H, Hu Y, et al. MicroRNA-320b promotes colorectal cancer proliferation and invasion by competing with its homologous microRNA-320a. Cancer letters. 2015 Jan 28;356(2 Pt B):669–75. pmid:25458952. Pubmed Central PMCID: Pmc4397650. Epub 2014/12/03. Eng.
  42. 42. Nygren MK, Tekle C, Ingebrigtsen VA, Makela R, Krohn M, Aure MR, et al. Identifying microRNAs regulating B7-H3 in breast cancer: the clinical impact of microRNA-29c. British journal of cancer. 2014 Apr 15;110(8):2072–80. pmid:24577056. Pubmed Central PMCID: Pmc3992492. Epub 2014/03/01. Eng.
  43. 43. Chauhan R, Lahiri N. Tissue- and Serum-Associated Biomarkers of Hepatocellular Carcinoma. Biomarkers in cancer. 2016;8(Suppl 1):37–55. pmid:27398029. Pubmed Central PMCID: 4933537.
  44. 44. Zhang L, Xu Y, Jin X, Wang Z, Wu Y, Zhao D, et al. A circulating miRNA signature as a diagnostic biomarker for non-invasive early detection of breast cancer. Breast cancer research and treatment. 2015 Nov;154(2):423–34. pmid:26476723. Epub 2015/10/20. Eng.
  45. 45. Fang Z, Tang J, Bai Y, Lin H, You H, Jin H, et al. Plasma levels of microRNA-24, microRNA-320a, and microRNA-423-5p are potential biomarkers for colorectal carcinoma. Journal of experimental & clinical cancer research: CR. 2015 Aug 22;34:86. pmid:26297223. Pubmed Central PMCID: Pmc4546358. Epub 2015/08/25. Eng.
  46. 46. Dhayat SA, Husing A, Senninger N, Schmidt HH, Haier J, Wolters H, et al. Circulating microRNA-200 Family as Diagnostic Marker in Hepatocellular Carcinoma. PloS one. 2015;10(10):e0140066. pmid:26447841. Pubmed Central PMCID: Pmc4598187. Epub 2015/10/09. Eng.
  47. 47. Varshney A, Panda JJ, Singh AK, Yadav N, Bihari C, Biswas S, et al. Targeted delivery of miR-199a-3p using self-assembled dipeptide nanoparticles efficiently reduces hepatocellular carcinoma. Hepatology. 2017 Nov 6. pmid:29108133.
  48. 48. Avraham R, Yarden Y. Regulation of signalling by microRNAs. Biochemical Society transactions. 2012 Feb;40(1):26–30. pmid:22260661. Pubmed Central PMCID: Pmc3621035. Epub 2012/01/21. Eng.
  49. 49. Inui M, Martello G, Piccolo S. MicroRNA control of signal transduction. Nature reviews Molecular cell biology. 2010 Apr;11(4):252–63. pmid:20216554. Epub 2010/03/11. Eng.
  50. 50. Shrivastav A, Varma S, Senger A, Khandelwal RL, Carlsen S, Sharma RK. Overexpression of Akt/PKB modulates N-myristoyltransferase activity in cancer cells. The Journal of pathology. 2009 Jul;218(3):391–8. pmid:19360752. Epub 2009/04/11. eng.
  51. 51. Thinon E, Morales-Sanfrutos J, Mann DJ, Tate EW. N-Myristoyltransferase Inhibition Induces ER-Stress, Cell Cycle Arrest, and Apoptosis in Cancer Cells. ACS chemical biology. 2016 Aug 19;11(8):2165–76. pmid:27267252. Pubmed Central PMCID: Pmc5077176. Epub 2016/06/09. Eng.
  52. 52. Baumgart M, Barth E. A miRNA catalogue and ncRNA annotation of the short-living fish Nothobranchius furzeri. 2017 Sep 5;18(1):693. pmid:28874118.
  53. 53. Nawrocki EP, Eddy SR. Infernal 1.1: 100-fold faster RNA homology searches. Bioinformatics (Oxford, England). 2013 Nov 15;29(22):2933–5. pmid:24008419. Pubmed Central PMCID: Pmc3810854. Epub 2013/09/07. eng.
  54. 54. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, et al. BLAST+: architecture and applications. BMC bioinformatics. 2009 Dec 15;10:421. pmid:20003500. Pubmed Central PMCID: Pmc2803857. Epub 2009/12/17. eng.
  55. 55. Wong N, Wang X. miRDB: an online resource for microRNA target prediction and functional annotations. Nucleic acids research. 2015 Jan;43(Database issue):D146–52. pmid:25378301. Pubmed Central PMCID: Pmc4383922. Epub 2014/11/08. eng.
  56. 56. Agarwal V, Bell GW, Nam JW, Bartel DP. Predicting effective microRNA target sites in mammalian mRNAs. eLife. 2015 Aug 12;4. pmid:26267216. Pubmed Central PMCID: Pmc4532895. Epub 2015/08/13. eng.
  57. 57. Vlachos IS, Zagganas K, Paraskevopoulou MD, Georgakilas G, Karagkouni D, Vergoulis T, et al. DIANA-miRPath v3.0: deciphering microRNA function with experimental support. Nucleic acids research. 2015 Jul 1;43(W1):W460–6. pmid:25977294. Pubmed Central PMCID: Pmc4489228. Epub 2015/05/16. eng.
  58. 58. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, et al. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nature genetics. 2000 May;25(1):25–9. pmid:10802651. Pubmed Central PMCID: Pmc3037419. Epub 2000/05/10. eng.
  59. 59. Diaz G, Zamboni F, Tice A, Farci P. Integrated ordination of miRNA and mRNA expression profiles. BMC genomics. 2015 Oct 12;16:767. pmid:26459852. Pubmed Central PMCID: Pmc4603994. Epub 2015/10/16. eng.
  60. 60. Nadal E, Truini A, Nakata A, Lin J, Reddy RM, Chang AC, et al. A Novel Serum 4-microRNA Signature for Lung Cancer Detection. Scientific reports. 2015 Jul 23;5:12464. pmid:26202143. Pubmed Central PMCID: Pmc5378885. Epub 2015/07/24. eng.
  61. 61. Keene JD. RNA regulons: coordination of post-transcriptional events. Nature reviews Genetics. 2007 Jul;8(7):533–43. pmid:17572691. Epub 2007/06/19. Eng.
  62. 62. Jha A, Panzade G, Pandey R, Shankar R. A legion of potential regulatory sRNAs exists beyond the typical microRNAs microcosm. Nucleic acids research. 2015 Oct 15;43(18):8713–24. pmid:26354861. Pubmed Central PMCID: Pmc4605316. Epub 2015/09/12. eng.
  63. 63. Riffo-Campos ÁL, Riquelme I, Brebi-Mieville P. Tools for Sequence-Based miRNA Target Prediction: What to Choose? International Journal of Molecular Sciences. 2016;17(12):1987. PubMed PMID: PMC5187787.
  64. 64. Maccani A, Hackl M, Leitner C, Steinfellner W, Graf AB, Tatto NE, et al. Identification of microRNAs specific for high producer CHO cell lines using steady-state cultivation. Applied microbiology and biotechnology. 2014 Sep;98(17):7535–48. pmid:25052466. Pubmed Central PMCID: Pmc4139590. Epub 2014/07/24. eng.
  65. 65. Xiao Y, Xu C, Guan J, Ping Y, Fan H, Li Y, et al. Discovering dysfunction of multiple microRNAs cooperation in disease by a conserved microRNA co-expression network. PloS one. 2012;7(2):e32201. pmid:22384175. Pubmed Central PMCID: Pmc3285207. Epub 2012/03/03. eng.
  66. 66. Hoque MO, Brait M, Rosenbaum E, Poeta ML, Pal P, Begum S, et al. Genetic and epigenetic analysis of erbB signaling pathway genes in lung cancer. Journal of thoracic oncology: official publication of the International Association for the Study of Lung Cancer. 2010 Dec;5(12):1887–93. pmid:21102258. Pubmed Central PMCID: Pmc3057519. Epub 2010/11/26. Eng.
  67. 67. Hynes NE, MacDonald G. ErbB receptors and signaling pathways in cancer. Current opinion in cell biology. 2009 Apr;21(2):177–84. pmid:19208461. Epub 2009/02/12. eng.
  68. 68. Stern DF. Tyrosine kinase signalling in breast cancer: ErbB family receptor tyrosine kinases. Breast Cancer Research. 2000;2(3):176–83. PubMed PMID: PMC138772. pmid:11250707
  69. 69. Appert-Collin A, Hubert P, Crémel G, Bennasroune A. Role of ErbB Receptors in Cancer Cell Migration and Invasion. Frontiers in Pharmacology. 2015;6:283. PubMed PMID: PMC4657385. pmid:26635612
  70. 70. Roderburg C, Mollnow T, Bongaerts B, Elfimova N, Vargas Cardenas D, Berger K, et al. Micro-RNA profiling in human serum reveals compartment-specific roles of miR-571 and miR-652 in liver cirrhosis. PloS one. 2012;7(3):e32999. pmid:22412969. Pubmed Central PMCID: Pmc3296762. Epub 2012/03/14. eng.
  71. 71. Min S, Li L, Zhang M, Zhang Y, Liang X, Xie Y, et al. TGF-beta-associated miR-27a inhibits dendritic cell-mediated differentiation of Th1 and Th17 cells by TAB3, p38 MAPK, MAP2K4 and MAP2K7. Genes and immunity. 2012 Dec;13(8):621–31. pmid:23034448. Epub 2012/10/05. eng.
  72. 72. Wei JQ, Chen H, Zheng XF, Zhang BX, Wang Y, Tang PF, et al. [Hsa-miR-654-5p regulates osteogenic differentiation of human bone marrow mesenchymal stem cells by repressing bone morphogenetic protein 2]. Nan fang yi ke da xue xue bao = Journal of Southern Medical University. 2012 Mar;32(3):291–5. PubMed pmid:22445969. Epub 2012/03/27. chi.
  73. 73. Guglielmelli P, Bisognin A, Saccoman C, Mannarelli C, Coppe A, Vannucchi AM, et al. Small RNA Sequencing Uncovers New miRNAs and moRNAs Differentially Expressed in Normal and Primary Myelofibrosis CD34+ Cells. PloS one. 2015;10(10):e0140445. pmid:26468945. Pubmed Central PMCID: Pmc4607157. Epub 2015/10/16. eng.
  74. 74. Liu C, Iqbal J, Teruya-Feldstein J, Shen Y, Dabrowska MJ, Dybkaer K, et al. MicroRNA expression profiling identifies molecular signatures associated with anaplastic large cell lymphoma. Blood. 2013 Sep 19;122(12):2083–92. pmid:23801630. Pubmed Central PMCID: Pmc3778551. Epub 2013/06/27. eng.
  75. 75. Yu T, Zuo Q-F, Gong L, Wang L-N, Zou Q-M, Xiao B. MicroRNA-491 regulates the proliferation and apoptosis of CD8+ T cells. Scientific reports. 2016 08/03/online;6:30923. pmid:27484289
  76. 76. Xie H, Sun L, Zhang L, Liu T, Chen L, Zhao A, et al. Mesenchymal Stem Cell-Derived Microvesicles Support Ex Vivo Expansion of Cord Blood-Derived CD34(+) Cells. Stem cells international. 2016;2016:6493241. pmid:27042183. Pubmed Central PMCID: 4799819.
  77. 77. Yu X, Wang Q-L, Li Y-F, Wang X-D, Xu A, Li Y. A novel miR-200b-3p/p38IP pair regulates monocyte/macrophage differentiation. Cell discovery. 2016 01/26/online;2:15043. pmid:27462440
  78. 78. Du C, Liu C, Kang J, Zhao G, Ye Z, Huang S, et al. MicroRNA miR-326 regulates TH-17 differentiation and is associated with the pathogenesis of multiple sclerosis. Nature immunology. 2009 Dec;10(12):1252–9. pmid:19838199. Epub 2009/10/20. eng.
  79. 79. Schmitt MJ, Philippidou D, Reinsbach SE, Margue C, Wienecke-Baldacchino A, Nashan D, et al. Interferon-gamma-induced activation of Signal Transducer and Activator of Transcription 1 (STAT1) up-regulates the tumor suppressing microRNA-29 family in melanoma cells. Cell communication and signaling: CCS. 2012 Dec 17;10(1):41. pmid:23245396. Pubmed Central PMCID: Pmc3541122. Epub 2012/12/19. eng.
  80. 80. Liu X, Zhan Z, Xu L, Ma F, Li D, Guo Z, et al. MicroRNA-148/152 impair innate response and antigen presentation of TLR-triggered dendritic cells by targeting CaMKIIalpha. Journal of immunology (Baltimore, Md: 1950). 2010 Dec 15;185(12):7244–51. pmid:21068402. Epub 2010/11/12. eng.
  81. 81. Garzia L, Andolfo I, Cusanelli E, Marino N, Petrosino G, De Martino D, et al. MicroRNA-199b-5p impairs cancer stem cells through negative regulation of HES1 in medulloblastoma. PloS one. 2009;4(3):e4998. pmid:19308264. Pubmed Central PMCID: Pmc2656623. Epub 2009/03/25. eng.
  82. 82. Favreau AJ, Sathyanarayana P. miR-590-5p, miR-219-5p, miR-15b and miR-628-5p are commonly regulated by IL-3, GM-CSF and G-CSF in acute myeloid leukemia. Leukemia research. 2012 Mar;36(3):334–41. pmid:22015065. Pubmed Central PMCID: Pmc3264807. Epub 2011/10/22. eng.
  83. 83. Smigielska-Czepiel K, van den Berg A, Jellema P, van der Lei RJ, Bijzet J, Kluiver J, et al. Comprehensive analysis of miRNA expression in T-cell subsets of rheumatoid arthritis patients reveals defined signatures of naive and memory Tregs. Genes and immunity. 2014;15(2):115–25. PubMed PMID: PMC3959220. pmid:24401767
  84. 84. Otaegui D, Baranzini SE, Armananzas R, Calvo B, Munoz-Culla M, Khankhanian P, et al. Differential micro RNA expression in PBMC from multiple sclerosis patients. PloS one. 2009 Jul 20;4(7):e6309. pmid:19617918. Pubmed Central PMCID: Pmc2708922. Epub 2009/07/21. eng.
  85. 85. Floyd DH, Zhang Y, Dey BK, Kefas B, Breit H, Marks K, et al. Novel anti-apoptotic microRNAs 582-5p and 363 promote human glioblastoma stem cell survival via direct inhibition of caspase 3, caspase 9, and Bim. PloS one. 2014;9(5):e96239. pmid:24805821. Pubmed Central PMCID: Pmc4013090. Epub 2014/05/09. eng.
  86. 86. Wang-Renault SF, Boudaoud S, Nocturne G, Roche E, Sigrist N, Daviaud C, et al. Deregulation of microRNA expression in purified T and B lymphocytes from patients with primary Sjogren's syndrome. Annals of the rheumatic diseases. 2018 Jan;77(1):133–40. pmid:28916716. Epub 2017/09/17. eng.
  87. 87. Emmrich S, Engeland F, El-Khatib M, Henke K, Obulkasim A, Schoning J, et al. miR-139-5p controls translation in myeloid leukemia through EIF4G2. Oncogene. 2016 Apr 7;35(14):1822–31. pmid:26165837. Epub 2015/07/15. eng.
  88. 88. Chen QY, Jiao DM, Zhu Y, Hu H, Wang J, Tang X, et al. Identification of carcinogenic potential-associated molecular mechanisms in CD133(+) A549 cells based on microRNA profiles. Tumour biology: the journal of the International Society for Oncodevelopmental Biology and Medicine. 2016 Jan;37(1):521–30. pmid:26227219. Epub 2015/08/01. eng.
  89. 89. Meng D, Yang S, Wan X, Zhang Y, Huang W, Zhao P, et al. A transcriptional target of androgen receptor, miR-421 regulates proliferation and metabolism of prostate cancer cells. The international journal of biochemistry & cell biology. 2016 Apr;73:30–40. pmid:26827675. Epub 2016/02/02. eng.
  90. 90. Jones DZ, Schmidt ML, Hobbing KR, Clark G, Kidd LR. miR-186 suppresses cell proliferation and anchorage-independence in a metastatic prostate cancer cell line. The FASEB Journal. 2016 April 1, 2016;30(1 Supplement):936.6.
  91. 91. Zhu M, Chen Q, Liu X, Sun Q, Zhao X, Deng R, et al. lncRNA H19/miR-675 axis represses prostate cancer metastasis by targeting TGFBI. The FEBS journal. 2014 Aug;281(16):3766–75. pmid:24988946. Epub 2014/07/06. eng.
  92. 92. Kong XJ, Duan LJ, Qian XQ, Xu D, Liu HL, Zhu YJ, et al. Tumor-suppressive microRNA-497 targets IKKbeta to regulate NF-kappaB signaling pathway in human prostate cancer cells. Am J Cancer Res. 2015;5(5):1795–804. PubMed pmid:26175947. Pubmed Central PMCID: Pmc4497445. Epub 2015/07/16. eng.
  93. 93. Saini S, Majid S, Shahryari V, Arora S, Yamamura S, Chang I, et al. miRNA-708 control of CD44(+) prostate cancer-initiating cells. Cancer research. 2012 Jul 15;72(14):3618–30. pmid:22552290. Epub 2012/05/04. eng.
  94. 94. Bai R, Weng C, Dong H, Li S, Chen G, Xu Z. MicroRNA-409-3p suppresses colorectal cancer invasion and metastasis partly by targeting GAB1 expression. International journal of cancer Journal international du cancer. 2015 Nov 15;137(10):2310–22. pmid:25991585. Epub 2015/05/21. Eng.
  95. 95. Luo X, Stock C, Burwinkel B, Brenner H. Identification and evaluation of plasma microRNAs for early detection of colorectal cancer. PloS one. 2013;8(5):e62880. pmid:23690963. Pubmed Central PMCID: Pmc3653912. Epub 2013/05/22. eng.
  96. 96. Li W, Jiang G, Zhou J, Wang H, Gong Z, Zhang Z, et al. Down-regulation of miR-140 induces EMT and promotes invasion by targeting Slug in esophageal cancer. Cellular physiology and biochemistry: international journal of experimental cellular physiology, biochemistry, and pharmacology. 2014;34(5):1466–76. pmid:25322669. Epub 2014/10/18. eng.
  97. 97. Bibby BA, Reynolds JV, Maher SG. MicroRNA-330-5p as a Putative Modulator of Neoadjuvant Chemoradiotherapy Sensitivity in Oesophageal Adenocarcinoma. PloS one. 2015;10(7):e0134180. pmid:26221725. Pubmed Central PMCID: Pmc4519309. Epub 2015/07/30. eng.
  98. 98. Tu L, Zhao E, Zhao W, Zhang Z, Tang D, Zhang Y, et al. hsa-miR-376c-3p Regulates Gastric Tumor Growth Both In Vitro and In Vivo. 2016;2016:9604257. pmid:27965982.
  99. 99. Xie J, Tan ZH, Tang X, Mo MS, Liu YP, Gan RL, et al. MiR-374b-5p suppresses RECK expression and promotes gastric cancer cell invasion and metastasis. World J Gastroenterol. 2014 Dec 14;20(46):17439–47. pmid:25516656. Pubmed Central PMCID: Pmc4265603. Epub 2014/12/18. eng.
  100. 100. Lu YC, Cheng AJ, Lee LY, You GR, Li YL, Chen HY, et al. MiR-520b as a novel molecular target for suppressing stemness phenotype of head-neck cancer by inhibiting CD44. Scientific reports. 2017 May 17;7(1):2042. pmid:28515423. Pubmed Central PMCID: Pmc5435724. Epub 2017/05/19. eng.
  101. 101. Chen J, Wang M, Guo M, Xie Y, Cong YS. miR-127 regulates cell proliferation and senescence by targeting BCL6. PloS one. 2013;8(11):e80266. pmid:24282530. Pubmed Central PMCID: Pmc3840165. Epub 2013/11/28. eng.
  102. 102. Liu H, Song Z, Liao D, Zhang T, Liu F, Zheng W, et al. miR-503 inhibits cell proliferation and invasion in glioma by targeting L1CAM. International journal of clinical and experimental medicine. 2015;8(10):18441–7. PubMed pmid:26770450. Pubmed Central PMCID: Pmc4694350. Epub 2016/01/16. eng.
  103. 103. Chen B, Pan W, Lin X, Hu Z, Jin Y, Chen H, et al. MicroRNA-346 functions as an oncogene in cutaneous squamous cell carcinoma. Tumour biology: the journal of the International Society for Oncodevelopmental Biology and Medicine. 2016 Feb;37(2):2765–71. pmid:26408183. Epub 2015/09/27. eng.
  104. 104. Formosa A, Markert EK, Lena AM, Italiano D, Finazzi-Agro E, Levine AJ, et al. MicroRNAs, miR-154, miR-299-5p, miR-376a, miR-376c, miR-377, miR-381, miR-487b, miR-485-3p, miR-495 and miR-654-3p, mapped to the 14q32.31 locus, regulate proliferation, apoptosis, migration and invasion in metastatic prostate cancer cells. Oncogene. 2014 Oct 30;33(44):5173–82. pmid:24166498. Epub 2013/10/30. eng.
  105. 105. Andrews K. Functional Validation of MicroRNAs Correlated with Second Biochemical Recurrence After Post-Prostatectomy Radiation in Prostate Cancer: The Ohio State University; 2017.
  106. 106. Wu J, Ji A, Wang X, Zhu Y, Yu Y, Lin Y, et al. MicroRNA-195-5p, a new regulator of Fra-1, suppresses the migration and invasion of prostate cancer cells. Journal of translational medicine. 2015 Sep 4;13:289. pmid:26337460. Pubmed Central PMCID: Pmc4558968. Epub 2015/09/05. eng.
  107. 107. Durojaiye V, Ilboudo A, Levine F, Osborne J, Park JY, Ogunwobi OO. Abstract 187: miR-1205/FRYL as a novel regulatory mechanism in androgen-independent prostate cancer. Cancer research. 2015;75(15 Supplement):187–.
  108. 108. Srivastava A, Goldberger H, Dimtchev A, Ramalinga M, Chijioke J, Marian C, et al. MicroRNA profiling in prostate cancer—the diagnostic potential of urinary miR-205 and miR-214. PloS one. 2013;8(10):e76994. pmid:24167554. Pubmed Central PMCID: Pmc3805541. Epub 2013/10/30. eng.
  109. 109. Hu J, Cai G, Xu Y, Cai S. The Plasma microRNA miR-1914* and -1915 Suppresses Chemoresistant in Colorectal Cancer Patients by Down-regulating NFIX. Current molecular medicine. 2016;16(1):70–82. PubMed pmid:26695693.
  110. 110. Ramalinga M, Roy A, Srivastava A, Bhattarai A, Harish V, Suy S, et al. MicroRNA-212 negatively regulates starvation induced autophagy in prostate cancer cells by inhibiting SIRT1 and is a modulator of angiogenesis and cellular senescence. Oncotarget. 2015 Oct 27;6(33):34446–57. pmid:26439987. Pubmed Central PMCID: Pmc4741465. Epub 2015/10/07. eng.
  111. 111. Zhang W, Zhang T, Jin R, Zhao H, Hu J, Feng B, et al. MicroRNA-301a promotes migration and invasion by targeting TGFBR2 in human colorectal cancer. Journal of Experimental & Clinical Cancer Research. 2014 December 31;33(1):113.
  112. 112. Dong Y, Zhao J, Wu CW, Zhang L, Liu X, Kang W, et al. Tumor suppressor functions of miR-133a in colorectal cancer. Molecular cancer research: MCR. 2013 Sep;11(9):1051–60. pmid:23723074. Epub 2013/06/01. Eng.
  113. 113. Mulrane L, Madden SF, Brennan DJ, Gremel G, McGee SF, McNally S, et al. miR-187 is an independent prognostic factor in breast cancer and confers increased invasive potential in vitro. Clinical cancer research: an official journal of the American Association for Cancer Research. 2012 Dec 15;18(24):6702–13. pmid:23060431. Epub 2012/10/13. eng.
  114. 114. Zhang Y, Xu G, Liu G, Ye Y, Zhang C, Fan C, et al. miR-411-5p inhibits proliferation and metastasis of breast cancer cell via targeting GRB2. Biochemical and biophysical research communications. 2016 Aug 05;476(4):607–13. pmid:27264952. Epub 2016/06/07. eng.
  115. 115. Yu G, Zhang T, Jing Y, Bao Q, Tang Q, Zhang Y. miR-519 suppresses nasopharyngeal carcinoma cell proliferation by targeting oncogene URG4/URGCP. Life sciences. 2017 Apr 15;175:47–51. pmid:28315691.
  116. 116. Zhang Y, Huang F, Wang J, Peng L, Luo H. MiR-15b mediates liver cancer cells proliferation through targeting BCL-2. International journal of clinical and experimental pathology. 2015;8(12):15677–83. PubMed PMID: PMC4730050. pmid:26884837
  117. 117. Guo W, Qiu Z, Wang Z, Wang Q, Tan N, Chen T, et al. MiR-199a-5p is negatively associated with malignancies and regulates glycolysis and lactate production by targeting hexokinase 2 in liver cancer. Hepatology (Baltimore, Md). 2015 Oct;62(4):1132–44. pmid:26054020. Epub 2015/06/09. eng.
  118. 118. Fang L, Zhang HB, Li H, Fu Y, Yang GS. miR-548c-5p inhibits proliferation and migration and promotes apoptosis in CD90(+) HepG2 cells. Radiology and oncology. 2012 Sep;46(3):233–41. pmid:23077462. Pubmed Central PMCID: Pmc3472946. Epub 2012/10/19. eng.
  119. 119. Wang X, Jiang F, Song H, Li X, Xian J, Gu X. MicroRNA-200a-3p suppresses tumor proliferation and induces apoptosis by targeting SPAG9 in renal cell carcinoma. Biochemical and biophysical research communications. 2016 Feb 12;470(3):620–6. pmid:26797273. Epub 2016/01/23. eng.
  120. 120. Zeng JS, Zhang ZD, Pei L, Bai ZZ, Yang Y, Yang H, et al. CBX4 exhibits oncogenic activities in breast cancer via Notch1 signaling. The international journal of biochemistry & cell biology. 2017 Dec 8. pmid:29229426.
  121. 121. Liu SG, Qin XG, Zhao BS, Qi B, Yao WJ, Wang TY, et al. Differential expression of miRNAs in esophageal cancer tissue. Oncology letters. 2013 May;5(5):1639–42. pmid:23761828. Pubmed Central PMCID: Pmc3678876. Epub 2013/06/14. eng.
  122. 122. Rau CS, Wu SC, Yang JC, Lu TH, Wu YC, Chen YC, et al. Profiling the circulating miRNAs in mice exposed to gram-positive and gram-negative bacteria by Illumina small RNA deep sequencing. Journal of biomedical science. 2015 Jan 7;22:1. pmid:25563241. Pubmed Central PMCID: Pmc4300083. Epub 2015/01/08. eng.
  123. 123. Dix A, Czakai K, Leonhardt I, Schaferhoff K, Bonin M, Guthke R, et al. Specific and Novel microRNAs Are Regulated as Response to Fungal Infection in Human Dendritic Cells. Frontiers in microbiology. 2017;8:270. pmid:28280489. Pubmed Central PMCID: 5322194.
  124. 124. Orr-Burks NL, Shim BS, Wu W, Bakre AA, Karpilow J, Tripp RA. MicroRNA screening identifies miR-134 as a regulator of poliovirus and enterovirus 71 infection. Scientific data. 2017 Mar 1;4:170023. pmid:28248924. Pubmed Central PMCID: 5332013.
  125. 125. Zhou L, Pupo GM, Gupta P, Liu B, Tran SL, Rahme R, et al. A parallel genome-wide mRNA and microRNA profiling of the frontal cortex of HIV patients with and without HIV-associated dementia shows the role of axon guidance and downstream pathways in HIV-mediated neurodegeneration. BMC genomics. 2012 Nov 28;13:677. pmid:23190615. Pubmed Central PMCID: Pmc3560210. Epub 2012/11/30. eng.
  126. 126. Cui L, Qi Y, Li H, Ge Y, Zhao K, Qi X, et al. Serum microRNA expression profile distinguishes enterovirus 71 and coxsackievirus 16 infections in patients with hand-foot-and-mouth disease. PloS one. 2011;6(11):e27071. pmid:22087245. Pubmed Central PMCID: Pmc3210764. Epub 2011/11/17. eng.
  127. 127. Wu CX, Cheng J, Wang YY, Wang JJ, Guo H, Sun H. Microrna Expression Profiling of Macrophage Line Raw264.7 Infected by Candida Albicans. Shock (Augusta, Ga). 2017 Apr;47(4):520–30. pmid:27749761. Epub 2016/10/18. eng.
  128. 128. Hu XM, Yan XH, Hu YW, Huang JL, Cao SW, Ren TY, et al. miRNA-548p suppresses hepatitis B virus X protein associated hepatocellular carcinoma by downregulating oncoprotein hepatitis B x-interacting protein. Hepatology research: the official journal of the Japan Society of Hepatology. 2016 Jul;46(8):804–15. pmid:26583881. Epub 2015/11/20. eng.
  129. 129. Li X, Huang Y, Zhang Y, He N. Evaluation of microRNA Expression in Patients with Herpes Zoster. Viruses. 2016 Dec 2;8(12). pmid:27918431. Pubmed Central PMCID: Pmc5192387. Epub 2016/12/06. eng.
  130. 130. Jaeger A, Hadlich F, Kemper N, Lubke-Becker A, Murani E, Wimmers K, et al. MicroRNA expression profiling of porcine mammary epithelial cells after challenge with Escherichia coli in vitro. BMC genomics. 2017 Aug 24;18(1):660. pmid:28836962. Pubmed Central PMCID: Pmc5571640. Epub 2017/08/25. eng.
  131. 131. Kong G, Zhang J, Zhang S, Shan C, Ye L, Zhang X. Upregulated microRNA-29a by hepatitis B virus X protein enhances hepatoma cell migration by targeting PTEN in cell culture model. PloS one. 2011 May 5;6(5):e19518. pmid:21573166. Pubmed Central PMCID: 3088678.
  132. 132. Sannigrahi MK, Sharma R, Singh V, Panda NK, Rattan V, Khullar M. Role of Host miRNA Hsa-miR-139-3p in HPV-16-Induced Carcinomas. Clinical cancer research: an official journal of the American Association for Cancer Research. 2017 Jul 15;23(14):3884–95. pmid:28143871. Epub 2017/02/02. eng.
  133. 133. Zhang GL, Li YX, Zheng SQ, Liu M, Li X, Tang H. Suppression of hepatitis B virus replication by microRNA-199a-3p and microRNA-210. Antiviral research. 2010 Nov;88(2):169–75. pmid:20728471.
  134. 134. El-Guendy NM, Helwa R, El-Halawany MS, Abdel Rahman Ali S, Tantawy Aly M, Hasan Alieldin N, et al. The Liver MicroRNA Expression Profiles Associated With Chronic Hepatitis C Virus (HCV) Genotype-4 Infection: A Preliminary Study. Hepatitis monthly. 2016 Apr;16(4):e33881. pmid:27275163. Pubmed Central PMCID: Pmc4893413. Epub 2016/06/09. eng.
  135. 135. Hoy AM, Lundie RJ, Ivens A, Quintana JF, Nausch N, Forster T, et al. Parasite-derived microRNAs in host serum as novel biomarkers of helminth infection. PLoS neglected tropical diseases. 2014 Feb;8(2):e2701. pmid:24587461. Pubmed Central PMCID: Pmc3930507. Epub 2014/03/04. eng.
  136. 136. Santhakumar D, Forster T, Laqtom NN, Fragkoudis R, Dickinson P, Abreu-Goodger C, et al. Combined agonist-antagonist genome-wide functional screening identifies broadly active antiviral microRNAs. Proceedings of the National Academy of Sciences of the United States of America. 2010 Aug 3;107(31):13830–5. pmid:20643939. Pubmed Central PMCID: Pmc2922253. Epub 2010/07/21. eng.
  137. 137. Squillace N, Bresciani E, Torsello A, Bandera A, Sabbatini F, Giovannetti C, et al. Changes in subcutaneous adipose tissue microRNA expression in HIV-infected patients. The Journal of antimicrobial chemotherapy. 2014 Nov;69(11):3067–75. pmid:25063777. Epub 2014/07/27. eng.
  138. 138. Zekri AN, Youssef AS, El-Desouky ED, Ahmed OS, Lotfy MM, Nassar AA, et al. Serum microRNA panels as potential biomarkers for early detection of hepatocellular carcinoma on top of HCV infection. Tumour biology: the journal of the International Society for Oncodevelopmental Biology and Medicine. 2016 Sep;37(9):12273–86. pmid:27271989. Epub 2016/10/27. eng.
  139. 139. Greco D, Kivi N, Qian K, Leivonen S-K, Auvinen P, Auvinen E. Human Papillomavirus 16 E5 Modulates the Expression of Host MicroRNAs. PloS one. 2011;6(7):e21646. PubMed PMID: PMC3128596. pmid:21747943
  140. 140. Hariharan M, Scaria V, Pillai B, Brahmachari SK. Targets for human encoded microRNAs in HIV genes. Biochemical and biophysical research communications. 2005 Dec 2;337(4):1214–8. pmid:16236258. Epub 2005/10/21. eng.
  141. 141. Hui AB, Lin A, Xu W, Waldron L, Perez-Ordonez B, Weinreb I, et al. Potentially prognostic miRNAs in HPV-associated oropharyngeal carcinoma. Clinical cancer research: an official journal of the American Association for Cancer Research. 2013 Apr 15;19(8):2154–62. pmid:23459718. Epub 2013/03/06. eng.
  142. 142. Xing TJ, Wang B, Zhang J. Screening and bioinformatic analysis of microRNA-associated immune clearance in patients with chronic hepatitis B. International journal of clinical and experimental medicine. 2014;7(12):4924–32. PubMed pmid:25663989. Pubmed Central PMCID: Pmc4307436. Epub 2015/02/11. eng.
  143. 143. Zhang X, Daucher M, Armistead D, Russell R, Kottilil S. MicroRNA expression profiling in HCV-infected human hepatoma cells identifies potential anti-viral targets induced by interferon-alpha. PloS one. 2013;8(2):e55733. pmid:23418453. Pubmed Central PMCID: Pmc3572124. Epub 2013/02/19. eng.
  144. 144. Jha A, Mehra M, Shankar R. The regulatory epicenter of miRNAs. Journal of biosciences. 2011 Sep;36(4):621–38. PubMed pmid:21857109. Epub 2011/08/23. Eng.
  145. 145. Kota J, Chivukula RR, O'Donnell KA, Wentzel EA, Montgomery CL, Hwang HW, et al. Therapeutic microRNA delivery suppresses tumorigenesis in a murine liver cancer model. Cell. 2009 Jun 12;137(6):1005–17. pmid:19524505. Pubmed Central PMCID: Pmc2722880. Epub 2009/06/16. Eng.
  146. 146. Liu C, Kelnar K, Liu B, Chen X, Calhoun-Davis T, Li H, et al. The microRNA miR-34a inhibits prostate cancer stem cells and metastasis by directly repressing CD44. Nature medicine. 2011 Feb;17(2):211–5. pmid:21240262. Pubmed Central PMCID: Pmc3076220. Epub 2011/01/18. Eng.
  147. 147. Wang YX, Lang F, Liu YX, Yang CQ, Gao HJ. In situ hybridization analysis of the expression of miR-106b in colonic cancer. International journal of clinical and experimental pathology. 2015;8(1):786–92. PubMed pmid:25755775. Pubmed Central PMCID: Pmc4348869. Epub 2015/03/11. Eng.
  148. 148. Wang S, Li H, Wang J, Wang D, Yao A, Li Q. Prognostic and biological significance of microRNA-127 expression in human breast cancer. Disease markers. 2014;2014:401986. pmid:25477702. Pubmed Central PMCID: Pmc4244930. Epub 2014/12/06. Eng.
  149. 149. Wan TM, Lam CS, Ng L, Chow AK, Wong SK, Li HS, et al. The clinicopathological significance of miR-133a in colorectal cancer. Disease markers. 2014;2014:919283. pmid:25104873. Pubmed Central PMCID: Pmc4101241. Epub 2014/08/12. Eng.
  150. 150. Fang Y, Sun B, Xiang J, Chen Z. MiR-301a promotes colorectal cancer cell growth and invasion by directly targeting SOCS6. Cellular physiology and biochemistry: international journal of experimental cellular physiology, biochemistry, and pharmacology. 2015;35(1):227–36. pmid:25591765. Epub 2015/01/17. Eng.
  151. 151. Mathieu J, Ruohola-Baker H. Regulation of stem cell populations by microRNAs. Advances in experimental medicine and biology. 2013;786:329–51. pmid:23696365. Pubmed Central PMCID: Pmc3901537. Epub 2013/05/23. eng.
  152. 152. Utikal J, Abba M, Novak D, Moniuszko M, Allgayer H. Function and significance of MicroRNAs in benign and malignant human stem cells. Seminars in cancer biology. 2015 Dec;35:200–11. pmid:26192966. Epub 2015/07/21. eng.
  153. 153. Jeker LT, Bluestone JA. MicroRNA regulation of T-cell differentiation and function. Immunological reviews. 2013 May;253(1):65–81. pmid:23550639. Pubmed Central PMCID: Pmc3621017. Epub 2013/04/05. eng.
  154. 154. Kurosaki T, Shinohara H, Baba Y. B cell signaling and fate decision. Annual review of immunology. 2010;28:21–55. pmid:19827951. Epub 2009/10/16. eng.
  155. 155. Korpal M, Kang Y. The emerging role of miR-200 family of microRNAs in epithelial-mesenchymal transition and cancer metastasis. RNA biology. 2008 Jul-Sep;5(3):115–9. PubMed PMID: PMC3532896. pmid:19182522
  156. 156. Dey BK, Gagan J, Yan Z, Dutta A. miR-26a is required for skeletal muscle differentiation and regeneration in mice. Genes & development. 2012 Oct 01;26(19):2180–91. pmid:23028144. Pubmed Central PMCID: Pmc3465739. Epub 2012/10/03. eng.
  157. 157. Jun H, Ying H, Daiwen C, Bing Y, Xiangbing M, Ping Z, et al. miR-628, a microRNA that is induced by Toll-like receptor stimulation, regulates porcine innate immune responses. Scientific reports. 2015 Jul 31;5:12226. pmid:26227240. Pubmed Central PMCID: Pmc4521185. Epub 2015/08/01. eng.
  158. 158. Gigante M, Pontrelli P, Herr W, Gigante M, D’Avenia M, Zaza G, et al. miR-29b and miR-198 overexpression in CD8(+) T cells of renal cell carcinoma patients down-modulates JAK3 and MCL-1 leading to immune dysfunction. Journal of translational medicine. 2016;14:84. PubMed PMID: PMC4827202. pmid:27063186
  159. 159. Gonzalez-Martin A, Adams BD. The microRNA miR-148a functions as a critical regulator of B cell tolerance and autoimmunity. 2016 Apr;17(4):433–40. pmid:26901150.
  160. 160. Luo S, Liu Y, Liang G, Zhao M, Wu H, Liang Y, et al. The role of microRNA-1246 in the regulation of B cell activation and the pathogenesis of systemic lupus erythematosus. Clinical epigenetics. 2015;7:24. pmid:25789080. Pubmed Central PMCID: Pmc4364674. Epub 2015/03/20. eng.
  161. 161. Chiang K, Liu H, Rice AP. miR-132 enhances HIV-1 replication. Virology. 2013 Mar 30;438(1):1–4. pmid:23357732. Pubmed Central PMCID: Pmc3594373. Epub 2013/01/30. eng.
  162. 162. Adoro S, Cubillos-Ruiz JR, Chen X, Deruaz M, Vrbanac VD, Song M, et al. IL-21 induces antiviral microRNA-29 in CD4 T cells to limit HIV-1 infection. Nature communications. 2015 Jun 25;6:7562. pmid:26108174. Pubmed Central PMCID: Pmc4481879. Epub 2015/06/26. eng.
  163. 163. Etheridge A, Lee I, Hood L, Galas D, Wang K. Extracellular microRNA: a new source of biomarkers. Mutation research. 2011 03/23;717(1–2):85–90. PubMed PMID: PMC3199035. pmid:21402084
  164. 164. Donzelli S, Blandino G, Muti P. Use of Buffy Coat miRNA Profiling for Breast Cancer Prediction in Healthy Women. Methods in molecular biology (Clifton, NJ). 2016;1379:13–9. pmid:26608286. Epub 2015/11/27. Eng.
  165. 165. Pereira DM, Rodrigues PM, Borralho PM, Rodrigues CM. Delivering the promise of miRNA cancer therapeutics. Drug discovery today. 2013 Mar;18(5–6):282–9. pmid:23064097. Epub 2012/10/16. eng.
  166. 166. Chen D, Chen Z, Jin Y, Dragas D, Zhang L, Adjei BS, et al. MicroRNA-99 family members suppress Homeobox A1 expression in epithelial cells. PloS one. 2013;8(12):e80625. pmid:24312487. Pubmed Central PMCID: Pmc3849180. Epub 2013/12/07. Eng.
  167. 167. Chen Z, Jin Y, Yu D, Wang A, Mahjabeen I, Wang C, et al. Down-regulation of the microRNA-99 family members in head and neck squamous cell carcinoma. Oral oncology. 2012 Aug;48(8):686–91. pmid:22425712. Pubmed Central PMCID: Pmc3380146. Epub 2012/03/20. eng.
  168. 168. Yen YC, Shiah SG, Chu HC, Hsu YM, Hsiao JR, Chang JY, et al. Reciprocal regulation of microRNA-99a and insulin-like growth factor I receptor signaling in oral squamous cell carcinoma cells. Molecular cancer. 2014 Jan 10;13:6. pmid:24410957. Pubmed Central PMCID: Pmc3895693. Epub 2014/01/15. eng.
  169. 169. Mei LL, Qiu YT, Huang MB, Wang WJ, Bai J, Shi ZZ. MiR-99a suppresses proliferation, migration and invasion of esophageal squamous cell carcinoma cells through inhibiting the IGF1R signaling pathway. Cancer biomarkers: section A of Disease markers. 2017 Jul 31. pmid:28800315. Epub 2017/08/12. eng.
  170. 170. Wu D, Zhou Y, Pan H, Qu P, Zhou J. microRNA99a inhibits cell proliferation, colony formation ability, migration and invasion by targeting fibroblast growth factor receptor 3 in prostate cancer. Molecular medicine reports. 2015 Feb;11(2):1469–75. pmid:25352177. Epub 2014/10/30. eng.
  171. 171. Wu D, Zhou Y, Pan H, Zhou J, Fan Y, Qu P. microRNA-99a inhibiting cell proliferation, migration and invasion by targeting fibroblast growth factor receptor 3 in bladder cancer. Oncology letters. 2014 Apr;7(4):1219–24. pmid:24944696. Pubmed Central PMCID: Pmc3961452. Epub 2014/06/20. eng.
  172. 172. Androulidaki A, Iliopoulos D, Arranz A, Doxaki C, Schworer S, Zacharioudaki V, et al. The kinase Akt1 controls macrophage response to lipopolysaccharide by regulating microRNAs. Immunity. 2009 Aug 21;31(2):220–31. pmid:19699171. Pubmed Central PMCID: Pmc2865583. Epub 2009/08/25. Eng.
  173. 173. Schramedei K, Mörbt N, Pfiefer G, Läuter J, Rosolowski M, Tomm JM et al. MicroRNA-21 targets tumor supressor genes ANP32A and SMARCA4. Oncogene. 2011;30:2975–2985. pmid:21317927.
  174. 174. Zhuang M, Gao W, Xu J, Wang P, Shu Y. The long non-coding RNA H19-derived miR-675 modulates human gastric cancer cell proliferation by targeting tumor suppressor RUNX1. Biochemical and biophysical research communications. 2014 Jun 6;448(3):315–22. pmid:24388988. Epub 2014/01/07. eng.
  175. 175. Xing TJ, Xu HT, Yu WQ, Wang B, Zhang J. MiRNA-548ah, a potential molecule associated with transition from immune tolerance to immune activation of chronic hepatitis B. International journal of molecular sciences. 2014 Aug 19;15(8):14411–26. pmid:25196343. Pubmed Central PMCID: 4159859.