SR family RNA binding proteins regulate splicing of nascent RNAs in vitro but their physiological role in vivo is largely unexplored, as genetic deletion of many SR protein genes results in embryonic lethality. Here we show that SRSF3HKO mice carrying a hepatocyte-specific deletion of Srsf3 (homologous to human SRSF3/SRp20) have a disrupted hepatic architecture and show pre- and postnatal growth retardation. SRSF3HKO mice exhibit impaired hepatocyte maturation with alterations in glucose and lipid homeostasis characterized by reduced glycogen storage, fasting hypoglycemia, increased insulin sensitivity and reduced cholesterol synthesis. We identify various splicing alterations in the SRSF3HKO liver that explain the in vivo phenotype. In particular, loss of SRSF3 causes aberrant splicing of Hnf1α, Ern1, Hmgcs1, Dhcr7 and Scap genes, which are critical regulators of glucose and lipid metabolism. Our study provides the first evidence for a SRSF3-driven genetic programme required for morphological and functional differentiation of hepatocytes that may have relevance for human liver disease and metabolic dysregulation.

The insulin receptor (IR) exists as two isoforms, IR-A and IR-B, which result from alternative splicing of exon 11 in the primary transcript. This alternative splicing is cell specific, and the relative proportions of exon 11 isoforms also vary during development, aging, and different disease states. We have previously demonstrated that both intron 10 and exon 11 contain regulatory sequences that affect IR splicing both positively and negatively. In this study, we sought to define the precise sequence elements within exon 11 that control exon recognition and cellular factors that recognize these elements. Using minigenes carrying linker-scanning mutations within exon 11, we detected both exonic splicing enhancer and exonic splicing silencer elements. We identified binding of SRp20 and SF2/ASF to the exonic enhancers and CUG-BP1 to the exonic silencer by RNA affinity chromatography. Overexpression and knockdown studies with hepatoma and embryonic kidney cells demonstrated that SRp20 and SF2/ASF increase exon inclusion but that CUG-BP1 causes exon skipping. We found that CUG-BP1 also binds to an additional intronic splicing silencer, located at the 3 end of intron 10, to promote exon 11 skipping. Thus, we propose that SRp20, SF2/ASF, and CUG-BP1 act antagonistically to regulate IR alternative splicing in vivo and that the relative ratios of SRp20 and SF2/ASF to CUG-BP1 in different cells determine the degree of exon inclusion.In mammals, alternative splicing is a common strategy for creating functional diversities of proteins that have cell and developmentally specific functions. Given the important role for splicing, it is not surprising that a recent estimate has proposed that 50 to 60% of mutations linked to disease affect splicing (21, 43). The majority of human genes undergo alternative pre-mRNA splicing through the use of competing 5Ј or 3Ј splice sites or through alternative inclusion/exclusion of exons in the pre-mRNA. These alternative exons often contain splice sites that diverge from the consensus site, and the presence of cis regulatory elements within the exon and/or the flanking introns determines whether these exons are recognized (18,20,31). These cis elements can have either a positive (enhancer) or a negative (silencer) effect on splicing. Both enhancers and silencers are thought to function through binding to specific trans-acting protein factors (1). Differences in the expression or activities of these trans-acting factors may modulate the recognition of the alternative exon and lead to developmental or tissue-specific differences in splicing. Proteins that bind to specific sequence elements to affect splice site selection include SR proteins, hnRNPs, and other related RNA binding proteins, such as the CELF family, TIA-1, and Raver-1 (11,12,14,25,32). Adding a further layer of regulation, local context, such as RNA secondary structure, may influence the way that binding motifs are recognized by their cognate factors (3, 10, 13).The human insulin receptor (IR) is encoded by a single INSR gene th...

Alterations in RNA splicing are associated with cancer, but it is not clear whether they result from malignant transformation or have a causative role. We show here that hepatocyte-specific deletion of splicing factor SRSF3 impairs hepatocyte maturation and metabolism in early adult life, and mice develop spontaneous hepatocellular carcinoma (HCC) with aging. Tumor development is preceded by chronic liver disease with progressive steatosis and fibrosis. SRSF3 protects mice against carbon tetrachloride-induced fibrosis and carcinogenesis, and suppresses inclusion of the profibrogenic EDA exon in fibronectin 1. Loss of SRSF3 increases expression of insulin like growth factor 2 (Igf2) and the A-isoform of the insulin receptor (Insr-A) allowing aberrant activation of mitogenic signaling, promotes aberrant splicing and expression of epithelial to mesenchymal transition (EMT) genes, and activates Wnt/β-catenin signaling leading to c-Myc induction. Finally, SRSF3 expression is either decreased or the protein miss-localized in human HCC. Conclusion Our data suggest a potential role for SRSF3 in preventing hepatic carcinogenesis by regulating splicing to suppress fibrosis, mitogenic splicing and EMT. Thus, these mice may provide an attractive model to discover the pathogenic mechanisms linking aberrant pre-mRNA splicing with liver damage, fibrosis and HCC.