Antagonistic effects of the SRp30c protein and cryptic 5′ splice sites on the alternative splicing of the apoptotic regulator Bcl-x

Abstract

Alternative 5′ splice site selection allows Bcl-x to produce two isoforms with opposite effects on apoptosis. The pro-apoptotic Bcl-xS variant is up-regulated by ceramide and down-regulated by protein kinase C through specific cis-acting exonic elements, one of which is bound by SAP155. Splicing to the Bcl-xS 5′ splice site is also enforced by heterogeneous nuclear ribonucleoprotein (hnRNP) F/H proteins and by Sam68 in cooperation with hnRNP A1. Here, we have characterized exon elements that influence splicing to the 5′ splice site of the anti-apoptotic Bcl-x L isoform. Within a 86-nucleotide region (B3) located immediately upstream of the Bcl-xL donor site we have identified two elements (ML2 and AM2) that stimulate splicing to the Bcl-xL 5′ splice site. SRp30c binds to these elements and can shift splicing to the 5′ splice site of Bcl-xL in an ML2/AM2-dependent manner in vitro and in vivo. The B3 region also contains an element that represses the use of Bcl-xL. This element is bound by U1 small nuclear ribonucleoprotein and contains two 5′ splice sites that can be used when the Bcl-x L 5′ splice site is mutated or the ML2/AM2 elements are deleted. Conversely, mutating the cryptic 5′ splice sites stimulates splicing to the Bcl-xL site. Thus, SRp30c stimulates splicing to the downstream 5′ splice site of Bcl-xL, thereby attenuating the repressive effect of upstream U1 snRNP binding sites. © 2008 by The American Society for Biochemistry and Molecular Biology, Inc.