Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress

Meifang Suo; Zhichen Lin; Dongfang Guo; Airong Zhang

doi:10.1371/journal.pone.0266374

Peer Review History

Original SubmissionJuly 27, 2021
24 Aug 2021 Decision Letter - Hiroyasu Nakano, Editor PONE-D-21-23535 Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress PLOS ONE Dear Dr. Zhang Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Although the reviewer feels that the study is potentially interesting, the reviewer requests the authors to address several concerns before publication. In addition, I also have several concerns. Specifically, the authors need to show the percentages of cells in each quadrant of the flow cytometry in Fig. 1d, 3d, 5b, 7a. Ideally, the authors should show the mean +/- SD of triplicate samples and determine the statistical significance of percentages of apoptotic cells in each condition. It is unclear why LSMCs spontaneously develop ER stress and undergo apoptosis. Moreover, it is also unclear why the conditioned medium of CAFs attenuates the expression of GPR78, CHOP, and ATF6 in LSMCs. To investigate the mechanisms underlying these issues may beyond the scope of the present study, however, the authors should discuss these points in more detail. Please submit your revised manuscript by December 24, 2021. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Hiroyasu Nakano, M.D., Ph.D. Academic Editor PLOS ONE Journal Requirements: PONE-D-21-23535 1. When submitting your revision, we need you to address these additional requirements. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match. When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section. 3. Thank you for stating the following financial disclosure: "NO Conceptualization: Meifang, Airong Zhang. Data curation: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Formal analysis: Meifang, Airong Zhang. Funding acquisition: Meifang, Airong Zhang. Methodology: Meifang, Airong Zhang. Investigation: Zhichen Lin, Dongfang Guo, Airong Zhang. Project administration: Meifang, Airong Zhang. Resources: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Supervision: Airong Zhang. Writing - original draft: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Writing - original draft: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang." Please state what role the funders took in the study. If the funders had no role, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript." If this statement is not correct you must amend it as needed. Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf. 4. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions. In your revised cover letter, please address the following prompts: a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent. b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories. We will update your Data Availability statement on your behalf to reflect the information you provide. 5. PLOS ONE now requires that authors provide the original uncropped and unadjusted images underlying all blot or gel results reported in a submission’s figures or Supporting Information files. This policy and the journal’s other requirements for blot/gel reporting and figure preparation are described in detail at https://journals.plos.org/plosone/s/figures#loc-blot-and-gel-reporting-requirements and https://journals.plos.org/plosone/s/figures#loc-preparing-figures-from-image-files. When you submit your revised manuscript, please ensure that your figures adhere fully to these guidelines and provide the original underlying images for all blot or gel data reported in your submission. See the following link for instructions on providing the original image data: https://journals.plos.org/plosone/s/figures#loc-original-images-for-blots-and-gels. In your cover letter, please note whether your blot/gel image data are in Supporting Information or posted at a public data repository, provide the repository URL if relevant, and provide specific details as to which raw blot/gel images, if any, are not available. Email us at plosone@plos.org if you have any questions. 6. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: The authors demonstrated that hsa_circ_0056686 was up-regulated in CAFs around uterine leiomyoma, and influenced the CAFs-mediated proliferation, migration and apoptosis of ULM cells. They also showed that hsa_circ_0056686 directly bound with miR-515-5p and this bounding contributed to the influence on the CAFs-mediated proliferation, migration, and apoptosis. This study is potentially interesting, however, some issues need to be addressed. Listed below are my specific concerns and suggestions 1. The effect of hsa_circ_0056686 on the CAFs-mediated proliferation, migration and apoptosis of ULM cells seems to be totally dependent on miR-515-5p. The data examining effects of the miR-515-5p antagomir on the CAFs-mediated proliferation, migration and apoptosis would strengthen the author’s claim. 2. The authors proposed AKAP13 as a possible target of miR-515-5p. Are there any previous reports or data that demonstrated the contribution of AKAP13 to proliferation, migration and apoptosis? 3. In figure 4B, the authors should demonstrate the western blotting data using anti-‘cleaved’ caspase 3 antibody. 4. Add a molecular weight marker to all western blot images in this manuscript. 5. More thorough explanations and important details on the experimental procedures is missing throughout the paper. In particular, effort should be performed for clarifying method to detect the protein amounts of collagens and the expression level of hsa_circ_0056686. ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0266374.r001
Revision 1
18 Jan 2022 Author Response Dear Editor, Thanks for your letter. We highly appreciate the valuable comments raised by the reviewers on our manuscript entitled " Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress " (ID: PONE-D-21-23535). We have improved our manuscript according to the comments, and the amendments are highlighted in red in the revised manuscript. Point to point responses are listed below. We would like to resubmit our manuscript for your kind consideration. Thanks again. Yours sincerely, Airong Zhang Department of Medical Laboratory, Zhumadian Central Hospital, Zhumadian, China. Email: airongz@21cn.com Although the reviewer feels that the study is potentially interesting, the reviewer requests the authors to address several concerns before publication. In addition, I also have several concerns. Specifically, the authors need to show the percentages of cells in each quadrant of the flow cytometry in Fig. 1d, 3d, 5b, 7a. Ideally, the authors should show the mean +/- SD of triplicate samples and determine the statistical significance of percentages of apoptotic cells in each condition. It is unclear why LSMCs spontaneously develop ER stress and undergo apoptosis. Moreover, it is also unclear why the conditioned medium of CAFs attenuates the expression of GPR78, CHOP, and ATF6 in LSMCs. To investigate the mechanisms underlying these issues may beyond the scope of the present study, however, the authors should discuss these points in more detail. Response: Good comments. We added the percentage of cells in each quadrant of flow cytometry in figs 1D, 3D, 5B, 7A. In this study，the data from at least three replicate experiments were presented as mean ± SD. We modified the poorly described sentences and added more details of statistical analysis in the materials and methods section (Statistical Analysis). As we know, ER stress signaling in the tumor microenvironment and cancer associated fibroblasts (CAFs) play important roles in tumor progression, and common drivers of ER stress are often low oxygen, nutrients, low pH, and reactive oxygen species accumulation. We fully understand the reviewer′s concerns. In fact, there is some degree of ER stress and apoptosis in cultured cells without any stress (maybe culture in vitro itself is a “stress factor”), and similar results have been reported in previous studies (including but not just Ref.1 & Ref.2). Ref.1: Tumour suppressor candidate 3 inhibits biological function and increases endoplasmic reticulum stress of melanoma cells WM451 by regulating AKT/GSK3-β/β-catenin pathway. doi: 10.1002/cbf.3515. Ref.2: MSRB3 promotes the progression of clear cell renal cell carcinoma via regulating endoplasmic reticulum stress. doi: 10.1016/j.prp.2019.152780. In recent years, several studies have found that CAFs are tightly linked to ER stress in tumor progression. Zeng et al. found that curcumin induced apoptosis and cell cycle arrest of CAFs, which were mainly caused by ROS mediated ER stress pathway, and mechanistically, curcumin induced upregulation of ROS via PERK-eIF2α-ATF4 axis triggers ER stress in CAFs (Ref.3). We fully consider the reviewers' concerns. Indeed, in a variety of tumor progressions, CAFs function as tumor promoting cellular activities, and inhibition of the ER stress pathway is an important mechanism of immune evasion in cancer cells. Therefore, we believe that CAFs could inhibit ER stress in tumor cells to some extent, and the two synergistically promote tumor progression. However, the specific regulatory mechanisms between tumor cells, CAFs and ER stress exceed the scope of this study, and still need further in-depth investigation. We added new literature in the discussion section and discussed the limitations of this study. Ref.3: Curcumin promotes cancer-associated fibroblasts apoptosis via ROS-mediated endoplasmic reticulum stress. doi: 10.1016/j.abb.2020.108613. 2. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match. When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section. Response: Thanks. We collated and revised funding information for this study. Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data-e.g., participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: The authors demonstrated that hsa_circ_0056686 was up-regulated in CAFs around uterine leiomyoma, and influenced the CAFs-mediated proliferation, migration and apoptosis of ULM cells. They also showed that hsa_circ_0056686 directly bound with miR-515-5p and this bounding contributed to the influence on the CAFs-mediated proliferation, migration, and apoptosis. This study is potentially interesting; however, some issues need to be addressed. Listed below are my specific concerns and suggestions: 1. The effect of hsa_circ_0056686 on the CAFs-mediated proliferation, migration and apoptosis of ULM cells seems to be totally dependent on miR-515-5p. The data examining effects of the miR-515-5p antagomir on the CAFs-mediated proliferation, migration and apoptosis would strengthen the author’s claim. Response: Good comments. The miR-515-5p mimic and miR-515-5p inhibitor were transfected into ULM cells mediated by CAFs, respectively. The results showed that overexpression of miR-515-5p inhibited cell migration and promoted cell apoptosis compared with CAFs cells transfected with sh-hsa_circ_0056686. In addition, overexpression of miR-515-5p promoted cleaved caspase3, GRP78, CHOP and ATF6 protein expression, inhibited Bcl-2, COL1A1, COL1A2 and COL1A3 protein expression, and transfection of miR-515-5p inhibitor to cells had the opposite effect as transfection of miR-515-5p mimic. The specific results were presented in Fig. 7 and results 7. 2. The authors proposed AKAP13 as a possible target of miR-515-5p. Are there any previous reports or data that demonstrated the contribution of AKAP13 to proliferation, migration and apoptosis? Response: Good comments. In this study, we predicted that AKAP13 was a potential target of miR-515-5p through online bioinformatics databases, and although it has not been clarified in this study whether miR-515-5p plays a regulatory role on uterine fibroid progression via AKAP13, it will be an important target for our subsequent study. In a previous study, Ng et al. found that AKAP13 was overexpressed in uterine fibroid tissue (Ref 1), which set the stage for our subsequent studies. In addition, another study claimed that AKAP13 expression knockdown suppressed the proliferation and invasion of acute myeloid leukaemia cells (Ref 2). We added the corresponding literature in the discussion section. Ref 1: A-Kinase Anchoring Protein 13 (AKAP13) Augments Progesterone Signaling in Uterine Fibroid Cells. J Clin Endocrinol Metab. doi: 10.1210/jc.2018-01216. Ref 2: Long non-coding RNA ZNF667-AS1 knockdown curbs liver metastasis in acute myeloid leukemia by regulating the microRNA-206/AKAP13 Axis. doi: 10.2147/CMAR.S269258. 3. In figure 4B, the authors should demonstrate the western blotting data using anti-‘cleaved’ caspase 3 antibody. Response: Thanks. In fact, the antibody used in the experiments was a cleaved caspase 3 antibody (in China, we verbally called it caspase 3 as brief), and we corrected its name in Figure 4B. 4. Add a molecular weight marker to all western blot images in this manuscript. Response: Thanks. We labeled the molecular weights of the proteins in all Western blot images. 5. More thorough explanations and important details on the experimental procedures is missing throughout the paper. In particular, effort should be performed for clarifying method to detect the protein amounts of collagens and the expression level of hsa_circ_0056686. Response: Thanks. We carefully checked and revised the manuscript and added more details of experimental procedures in the materials and methods section. 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No We appreciate for Editors/Reviewers’ warm work earnestly, and hope that the correction will meet with approval. Once again, thank you very much for your comments and suggestions. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0266374.r002
2 Mar 2022 Decision Letter - Hiroyasu Nakano, Editor PONE-D-21-23535R1Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stressPLOS ONE Dear Dr. Zhang Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. The reviewer has been satisfied with your revision, however, the reviewer still has minor concerns. Specifically, although the authors added the molecular weight markers in Figures, there is no molecular weight markers in Supplementary Figures. Moreover, the order of Figures are not correct; Figure 1 appears to be placed after Figure 7. Thus, I would like to recommend that the authors respond to these points raised by the reviewers. Please submit your revised manuscript by Apr 16 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Hiroyasu Nakano, M.D., Ph.D. Academic Editor PLOS ONE Journal Requirements: Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: The authors did not answer my comment #4 'Add a molecular weight marker to all western blot images.' ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0266374.r003
Revision 2
17 Mar 2022 Author Response Dear Editor, Thanks for your letter. We highly appreciate the valuable comments raised by the reviewers on our manuscript entitled " Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress " (ID: PONE-D-21-23535R1). We have improved our manuscript according to the comments, and the amendments are highlighted in red in the revised manuscript. Point to point responses are listed below. We would like to resubmit our manuscript for your kind consideration. Thanks again. Yours sincerely, Airong Zhang Department of Medical Laboratory, Zhumadian Central Hospital, Zhumadian, China. Email: airongz@21cn.com The reviewer has been satisfied with your revision, however, the reviewer still has minor concerns. Specifically, although the authors added the molecular weight markers in Figures, there is no molecular weight markers in Supplementary Figures. Moreover, the order of Figures are not correct; Figure 1 appears to be placed after Figure 7. Thus, I would like to recommend that the authors respond to these points raised by the reviewers. Response: Sorry for our carelessness. Fist, we added the molecular weight markers in Supplementary Figures. In addition, we have adapted the order of figures 1 and 7 when uploading our latest revised manuscript. 2. Thank you for stating the following financial disclosure: "No Conceptualization: Meifang, Airong Zhang. Data curation: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Formal analysis: Meifang, Airong Zhang. Funding acquisition: Meifang, Airong Zhang. Methodology: Meifang, Airong Zhang. Investigation: Zhichen Lin, Dongfang Guo, Airong Zhang. Project administration: Meifang, Airong Zhang. Resources: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Supervision: Airong Zhang. Writing - original draft: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Writing - original draft: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang." At this time, please address the following queries: a) Please clarify the sources of funding (financial or material support) for your study. List the grants or organizations that supported your study, including funding received from your institution. b) State what role the funders took in the study. If the funders had no role in your study, please state: “The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.” c) If any authors received a salary from any of your funders, please state which authors and which funders. d) If you did not receive any funding for this study, please state: “The authors received no specific funding for this work.” Response: Thanks. We collated and revised financial disclosures for this study. Details are as follows, “Authors' contributions Conceptualization: Meifang, Airong Zhang. Data curation: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Formal analysis: Meifang, Airong Zhang. Methodology: Meifang, Airong Zhang. Investigation: Zhichen Lin, Dongfang Guo, Airong Zhang. Project administration: Meifang, Airong Zhang. Resources: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Supervision: Airong Zhang. Writing - original draft: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Writing - original review & editing: Meifang, Zhichen Lin, Dongfang Guo, Airong Zhang. Funding The author(s) received no specific funding for this work.” Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: The authors did not answer my comment #4 'Add a molecular weight marker to all western blot images.' Response: Sorry for our carelessness. We added the molecular weight markers in Supplementary Figures of latest revised manuscript. 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No We appreciate for Editors/Reviewers’ warm work earnestly, and hope that the correction will meet with approval. Once again, thank you very much for your comments and suggestions. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0266374.r004
21 Mar 2022 Decision Letter - Hiroyasu Nakano, Editor Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress PONE-D-21-23535R2 Dear Dr. Zhang We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Hiroyasu Nakano, M.D., Ph.D. Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: (No Response) ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: (No Response) ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: (No Response) ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: (No Response) ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: (No Response) ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No https://doi.org/10.1371/journal.pone.0266374.r005
Formally Accepted
29 Mar 2022 Acceptance Letter - Hiroyasu Nakano, Editor PONE-D-21-23535R2 Hsa_circ_0056686, derived from cancer-associated fibroblasts, promotes cell proliferation and suppresses apoptosis in uterine leiomyoma through inhibiting endoplasmic reticulum stress Dear Dr. Zhang: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Professor Hiroyasu Nakano Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0266374.r006

Open letter on the publication of peer review reports

PLOS recognizes the benefits of transparency in the peer review process. Therefore, we enable the publication of all of the content of peer review and author responses alongside final, published articles. Reviewers remain anonymous, unless they choose to reveal their names.

We encourage other journals to join us in this initiative. We hope that our action inspires the community, including researchers, research funders, and research institutions, to recognize the benefits of published peer review reports for all parts of the research system.

Learn more at ASAPbio .