SOX10-Regulated Promoter Use Defines Isoform-Specific Gene Expression in Schwann Cells

Abstract

To support the development of diverse cell types, multicellular organisms have adopted mechanisms to tailor gene expression profiles to specific spatial and temporal contexts. These mechanisms—including alternative promoter use—mediate the expression of gene isoforms that are functionally required in a given cell, at a given time. Moreover, the investigation of isoform-specific gene expression profiles in a cell type of interest can provide insights into the biology of the cells and the functions of the associated isoforms. Schwann cells—the myelinating cells of the peripheral nervous system—are an example of a highly specialized cell type with a similarly specialized gene expression profile. SOX10 is a transcriptional activator that is required for the development and maintenance of these cells, and SOX10 target genes are known to be critical for Schwann cell function. Interestingly, SOX10-regulated promoter elements have been characterized and a subset of these regulatory elements dictate the expression of unique gene isoforms. These findings implicate isoform-specific expression of the associated loci in Schwann cell function. This dissertation explores the concept of SOX10-regulated promoter use as a critical factor for isoform-specific gene expression in Schwann cells. These efforts were predicted to identify gene products that contribute to Schwann cell function based on their SOX10-dependent expression in these cells. To interrogate the function of SOX10 at promoter elements, we employ multiple strategies. First, we describe a sequence conservation-based analysis that identified a SOX10-regulated promoter at a known demyelinating disease gene, MTMR2. The SOX10-regulated promoter directs expression of transcripts encoding a unique MTMR2 protein isoform, suggesting that this isoform may be particularly important for Schwann cell function. Subsequently, we took a broad, unbiased, genome-wide approach to identify SOX10-regulated promoters in Schwann cells. These studies utilize transcription start site (TSS) mapping to assess the activity of SOX10-bound promoters in Schwann cells in an intact adult peripheral nerve, in differentiating primary Schwann cells, and upon ablation of SOX10 expression in vitro. The integration of these data provides a comprehensive assessment of SOX10-dependent promoter activity as it relates to multiple models and stages of Schwann cell development. Based on these findings we explore characteristics of SOX10-dependent TSSs, which supports the characterization of these regulatory elements as bona fide SOX10-dependent Schwann cell promoters. Finally, we validate and expand upon our findings at four previously unreported SOX10 target genes in Schwann cells, each of which is a multi-TSS locus with a SOX10-regulated promoter: ARPC1A, CHN2, DDR1, and GAS7. For each gene we confirm the SOX10-mediated regulatory activity of the promoter and explore the expression of transcript and protein isoforms in Schwann cells. Importantly, these findings suggest roles for the SOX10-regulated gene products in peripheral myelination and—with further study—will likely reveal new insights into Schwann cell biology. As a whole, this dissertation contributes to our understanding of SOX10-regulated isoform expression in Schwann cells and confirms the utility of our strategy toward the identification of high-confidence candidate gene products for further study.