807 Alternative mRNA splicing regulates epidermal differentiation

Alternative splicing (AS) of messenger RNAs diversifies gene function by generating RNA and protein isoforms with different biological properties. Over 95% of multi-exon human genes are alternatively spliced, but the functional consequences of most AS events are still not known. We identified 5,826 AS events, most involving alternatively spliced cassette exons. RNA binding motif analysis nominated 35 candidate RNA binding proteins (RBPs) that control epidermal AS. RNAi depletion and functional analysis of the previously unreported RBPs identified CNOT4, DAZAP1, RBM4, RBM42, and TIA1 as novel factors controlling keratinocyte proliferation and differentiation. Within the splicing dataset, 428 AS events occurred in genes whose steady-state expression was generally stable between progenitor vs differentiated keratinocytes, indicating the potential for widespread AS-mediated regulation in the absence of overall gene expression differences. We functionally characterized one of these AS events in mitogen-activated protein kinase kinase kinase (MAP3K7), which displayed a highly reproducible cassette exon AS event (p<0.0001, FDR<0.001) in which expression of a shorter MAP3K7 isoform in basal keratinocytes switched to a longer isoform in the suprabasal epidermis. Isoform-specific perturbation demonstrated that the short MAP3K7 isoform is necessary to maintain epidermal progenitor features, while expression of the long isoform is required to induce epidermal differentiation. In summary, our study defines many alternative mRNA splicing events associated with epidermal differentiation and demonstrates a critical role for alternative mRNA splicing and its regulators in skin homeostasis.