RBM4–Nova1–SRSF6 splicing cascade modulates the development of brown adipocytes

• Rbm4a−/− brown adipocytes exhibit differential splicing profiles.
• RBM4a enhances the skipping of Nova 1 exon 4 in a CU-element dependent manner.
• Nova 1 reduces SRSF6 expression in pre-brown adipocyte via AS-coupled NMD pathway.
• Nova 1 and SRSF6 exert opposite effect on the development of brown adipocytes.

Alternative splicing (AS) is a pivotal mechanism for the expansion of gene diversity, which determines the cellular fate or specification. However, the effect of AS networks on brown adipogenesis has not been comprehensively investigated. In this study, we identified the discriminative splicing profiles of RNA-binding motif protein 4a-knockout (RBM4a−/−) brown adipocytes (BAs) and compared them with those of their wild-type counterparts through deep RNA-sequencing. Among these candidates, RBM4a ablation enhanced the relative level of exon 4-excluded neuro-oncological ventral antigen 1 (Nova1− 4) transcripts, which were predominantly generated in embryonic BAs. By contrast, most of the Nova1 transcripts were exon 4-included (Nova1+ 4) in mature BAs. The Nova1 isoforms exhibited differential effects on repressing the development of BAs. Moreover, overexpression of Nova1 proteins reduced the serine/arginine splicing factor 6 (SRSF6) level by enhancing the generation of intron 2-included (SRSF6+ intron 2) transcripts, which are a putative candidate of the AS-coupled nonsense-mediated decay mechanism. Furthermore, we observed the positive effect of SRSF6 on BA development. These results highlight the hierarchical role of RBM4a in an AS cascade that manipulates brown adipogenesis.