Let's sp(l)ice up pluripotency!

mRNA-splicing presents an important regulatory step that contributes to the functional repertoire of all cell types and tissues. Profound changes in alternative splicing have also been revealed during embryonic stem cell (ESC) differentiation. Based on a genome-wide screen for pluripotency regulators, Ng and colleagues now identify SON as a new splicing regulator in the maintenance of human ESC pluripotency and self-renewal (Lu et al, 2013). RNA splicing is the process of removal of introns from nascent pre-mRNAs that results in the formation of mature mRNAs, with all exons joined together. The human genome contains B40000 genes that give rise to B200000 transcripts, suggesting that each gene locus can generate several different transcripts. This phenomenon can be in part explained by alternative splicing. Alternative splicing occurs when an exon can be either included or excluded from the mature mRNA, giving rise to different transcripts called splice variants. Transcriptome analysis suggests that the vast majority of human multiexonic genes are alternatively spliced (Pan et al, 2008), and this occurs in a cell-type-specific manner. For these reasons splicing should not be considered a housekeeping cellular function, but rather a dynamically regulated process. Interestingly, little is known about the molecular mechanisms controlling it. The recent study on SON therefore provides one of the first examples of a protein that regulates alternative splicing in the context of ESC pluripotency. The Ng lab at the Genome Institute of Singapore previously performed a genome-wide RNAi screen to identify genes required for self-renewal of human embryonic stem (hES) cells (Chia et al, 2010), and found among the top candidates the spliceosome-associated factor SON. SON is a nuclear protein that has been shown to interact with the spliceosome and to regulate splicing in HeLa cells (Sharma et al, 2010; Ahn et al, 2011). First, Lu et al (2013) characterized the role of SON as a regulator of Pluripotency in hES cells and found that knockdown of SON leads to impaired self-renewal and downregulation of pluripotency markers such as Oct4, Nanog and Prdm14. The reader might find those results not surprising, because a cell with a faulty splicing machinery should not be able to perform any cellular function—let alone maintaining pluripotency—but this would only be a rather superficial perspective: Through a series of elegant experiments the authors showed that loss of SON impairs both self-renewal and cell survival, but the two processes are uncoupled. For instance, knockdown of SON in the presence of the ROCK inhibitor, a chemical commonly used to reduce cell death in hES cells, results in differentiation with negligible effects on survival. The authors then showed that SON is expressed specifically in pluripotent cells and becomes downregulated upon differentiation. Moreover, when SON is reduced only B10% of hES cellspecific transcripts show aberrant splicing and, quite remarkably, among the same gene locus only a few introns are affected. In other words, SON is a very specific regulator of splicing in hES cells. But how is such exquisite specificity achieved? To address this question, the authors deployed their powerful genomic technologies and found that the introns regulated by SON tend to be shorter, with a weaker splicing strength and more GC-rich than those not affected by SON depletion. These results indicate that the activity of SON is regulated by