RBM4-Nova1-SRSF6 splicing cascade modulates 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.

RBM4a modulates the impact of PRDM16 on development of brown adipocytes through an alternative splicing mechanism.

Brown adipocytes (BAs) exhibit an energy-expending signature that is important in balancing metabolic homeostasis. In this study, results of transcriptome analyses revealed the reprogrammed splicing profile of the PR domain containing 16 (PRDM16) gene, a key transcription factor involved in brown adipogenesis, throughout development of wild-type brown adipose tissues (BATs). Moreover, discriminative splicing patterns of PRDM16 transcripts were noted in embryonic and postnatal RBM4a-/- BATs. Overexpression of RBM4a enhanced the relative levels of PRDM16-ex 16 transcripts by simultaneously interacting with exonic and intronic CU elements, which encoded the PRDM16S isoform containing a distinct C-terminus. The presence of the overexpressed PRDM16S isoform showed a stronger effect than the overexpressed PRDM16L isoform on enhancing transcriptional activity of the RBM4a and the PGC-1α promoter. Overexpression of the PRDM16S isoform exerted more-prominent effects on enhancing the BAT-related gene program and energy expenditure compared to those of PRDM16L-overexpressing cells. Our studies demonstrated that RBM4a-regulated alternative splicing constituted another regulatory mechanism for strengthening the influence of PRDM16 on the development of brown adipocytes.