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A direct link between MITF, innate immunity, and hair graying

Fig 4

MITF binds to the promoter of innate immune target genes and acts as a transcriptional repressor.

(A, B) Relative gene expression in melan-a cells 48 hours after transfection with siRNA. In (A), cells were transfected with siRNAs targeting Mitf (siMitf and siMitf-OM) or an analogous scrambled control siRNA (siMitf-scram). In (B), cells were transfected with one of two siRNAs targeting Irf4 (siIrf4_a, siIrf4_b) or siNC1. The bars represent the mean ± standard deviation, and the asterisks indicate gene expression changes with a q-value of <0.05 using the two-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli, with Q = 5%. Data presented in A and B are representative of two independent knockdown experiments performed in melan-a cells at two different cell passages. (C) Venn diagrams demonstrating the intersection of gene lists. The left diagram represents the genes exhibiting a 1.5-fold or greater difference in expression in Mitfmi-vga9/+ over wild-type McSCs (Mitfmi-vga9/+DEGs, top circle) overlapped with the list of genes associated with MITF ChIP-seq peaks reported by Webster et al. 2014 (MITF ChIP-seq genes, bottom circle) [41] and are considered potential direct targets of MITF. These 108 direct targets are provided in S4 Data. Direct target genes were further identified by comparing them with a list of known pigmentation genes in the upper right diagram and with the list of 55 innate immune-related genes up-regulated in Mitfmi-vga9/+ McSCs (as defined in Fig 3A) in the lower right diagram. (D) MITF ChIP-qPCR performed in melan-a cells and assayed for enrichment at the indicated gene loci (Dct, Ifih1, Ifit3, Stat1). A centromeric DNA sequence was amplified as a negative control (neg con). Error bars indicate mean ± standard deviation of two independent ChIP pulldowns. The raw data used to generate the graphs in (A), (B), and (D) are available in S1 Data. Actb, actin beta; ChIP, chromatin immunoprecipitation; ChIP-qPCR, chromatin immunoprecipitation quantitative polymerase chain reaction; ChIP-seq, chromatin immunoprecipitation sequencing; DEG, differentially expressed gene; IgG, immunoglobulin G; McSC, melanocyte stem cell; MITF, melanogenesis associated transcription factor; neg con, negative control; siNC1, nontargeting control siRNA; siRNA, small interfering RNA.

Fig 4