Alternative splicing of BCL-x is controlled by RBM25 binding to a G-quadruplex in BCL-x pre-mRNA

Abstract

BCL-x is a master regulator of apoptosis whose pre-mRNA is alternatively spliced into either a long (canonical) anti-apoptotic Bcl-xL isoform, or a short (alternative) pro-apoptotic Bcl-xS isoform. The balance between these two antagonistic isoforms is tightly regulated and overexpression of Bcl-xL has been linked to resistance to chemotherapy in several cancers, whereas overexpression of Bcl-xS is associated to some forms of diabetes and cardiac disorders. The splicing factor RBM25 controls alternative splicing of BCL-x: its overexpression favours the production of Bcl-xS, whereas its downregulation has the opposite effect. Here we show that RBM25 directly and specifically binds to GQ-2, an RNA G-quadruplex (rG4) of BCL-x pre-mRNA that forms at the vicinity of the alternative 5′ splice site leading to the alternative Bcl-xS isoform. This RBM25/rG4 interaction is crucial for the production of Bcl-xS and depends on the RE (arginine-glutamate-rich) motif of RBM25, thus defining a new type of rG4-interacting domain. PhenDC3, a benchmark G4 ligand, enhances the binding of RBM25 to the GQ-2 rG4 of BCL-x pre-mRNA, thereby promoting the alternative pro-apoptotic Bcl-xS isoform and triggering apoptosis. Furthermore, the screening of a combinatorial library of 90 putative G4 ligands led to the identification of two original compounds, PhenDH8 and PhenDH9, superior to PhenDC3 in promoting the Bcl-xS isoform and apoptosis. Thus, favouring the interaction between RBM25 and the GQ-2 rG4 of BCL-x pre-mRNA represents a relevant intervention point to re-sensitize cancer cells to chemotherapy.

G-quadruplex forming regions inGCKandTM6SF2are targets for differential DNA methylation in metabolic disease and hepatocellular carcinoma patients

“…Splicing factors like U2AF1, which recognizes the 3ʹ splice site, are among the top candidates associated with G4s 37 . G4 formation regulates alternative splicing of BCL-x, since stabilization of the G4 structure promotes the expression of the pro-apoptotic BCL-xS isoform and triggering apoptosis 106 . However, this requires critical discussion since the G4 motif is found on the template strand, meaning the pre-mRNA cannot not form the G4.…”

Neoantigens in cancer immunotherapy: focusing on alternative splicing

“…The utilization of alternative 5′ splice sites in Bcl-x pre-mRNA results in the formation of the anti-apoptotic Bcl-x(L) and pro-apoptotic Bcl-x(S) protein isoforms ( 32 , 33 ). Bcl-x(L) is transcriptionally upregulated in numerous cancers and is linked to chemoresistance, as well as to the RAS-induced expression of stemness regulators and the preservation of a cancer-initiating cell phenotype ( 34 ). Indeed, the simultaneous expression of mutant versions of SF3B1 and SRSF2 in hematopoietic progenitors has been demonstrated to induce cell death ( 35 , 36 ).…”