Unknown,Transcriptomics,Genomics,Proteomics

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Genomic Analysis Reveals Distinct Mechanisms and Functional Classes of SOX10-Regulated Genes in Melanocytes [ChIP-Seq]

ABSTRACT: We performed ChIP-Seq analysis of SOX10, histone H3 lysine 27 acetylation (H3K27ac) and H3K27 trimethylation (H3K27me3) in melanocytes to profile the genomic binding sites of SOX10 and the chromatin landscape. In parallel, we generated Sox10 haploinsufficient cell lines using gene knockout approaches and conducted microarray gene expression analysis to identify functional gene targets of SOX10 transcriptional regulation in melanocytes. We demonstrate that SOX10 predominantly engages “open” chromatin, binds to melanocyte enhancer elements and plays a central role in transcriptional activation and repression of functionally distinct classes of genes. Furthermore, we identified cis-regulatory sequence motifs of putative co-regulatory transcription factors that define SOX10-activated and SOX10-repressed target genes. Our results uncover novel mechanisms and roles of SOX10 in global transcriptional regulation of diverse regulatory pathways in the melanocyte lineage. ChIP-seq profiling of SOX10, H3K27ac, and H3K27me3 in the mouse melanocyte cell line melan-Ink4a-Arf-1 (melan-a).

ORGANISM(S): Mus musculus

SUBMITTER: Laura Baxter

PROVIDER: E-GEOD-69949 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Gene expression profiling of tissues derived from different genetic backgrounds can provide insight to potential phenotype-related changes in gene expression. We have compared wildtype C57BL/6 (melan-Ink4a) melanocyte cell gene expression to that of, âbrownâ Tyrp1b/ b (melan-Ink4a-b) and âchocolateâ Rab38 cht/cht (melan-cht) melanocyte derived lines. We observed alterations in gene expression consistent with a physiological response of melan-Ink4a-b cells to endoplasmic reticulum protein-folding impairment. Additionally, a set of loci was identified that are expressed in a similar manner to known melanocyte genes. Importantly, the collection of 254 genes is enriched for expression in the multiple melanocyte lines relative to control cell populations. This set contains Tyr, Tyrp1, Dct, Si, Melna, Slc24a5, Slc45a2, Slc24a4, Mitf, and Sox10, in addition to genes not previously attributed to pigmentation and genes with unknown function. Further evaluation of one gene in this cohort, Gpnmb, demonstrated an expression pattern similar to Dct, Si and Mitf. This expression is vastly reduced in MitfMi mutants embryos, indicating that it is dependent on Mitf for in vivo expression. Consistent with this notion, a highly-conserved MITF binding site resides directly upstream of human GPNMB, and deletion of this sequence dramatically reduces in vitro enhancer activity. Keywords: NIH embryo fibroblast cell line, melanocyte cell line, Ink4a, Tyrp1b, Rab38 A 32k expression arrays were generated by printing Oligo nucleotides from Operon Mouse oligo set (MV3) onto epoxy slides. Total RNA from immortalized melanocyte cell lines derived from C57BL/6J Ink4a-/- (melan-Ink4a-1), C57BL/6J Ink4a-/-; Tyrp1b/b (melan-Ink4a-b) and 2 separate C57BL/6J Ink4a-/-; RaB38cht/cht lines (melan-cht4), (melan-cht5) and NIH3T3 were labeled with Cy5/3 and co-hybridized with a total RNA universal reference (Stratagene, Inc) labeled with Cy3/5. Each comparison was done in Triplicate including one dye swap.

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Genomic Analysis Reveals Distinct Mechanisms and Functional Classes of SOX10-Regulated Genes in Melanocytes [ChIP-Seq]

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