The RNA-binding protein Sam68 modulates the alternative splicing of Bcl-x

• DOI: 10.1083/jcb.200701005
• Publication Date: 2007-03-20T00:20:51Z
• Authors: Maria Paola Paron...; Tilman Achsel; Autumn Massiello; Charles E. Chalfant; Claudio Sette
• Keywords: Settore BIO/16 - ANATOMIA UMANA; 0301 basic medicine; 570; Binding Sites; Serine-Arginine Splicing Factors; Heterogeneous Nuclear Ribonucleoprotein A1; bcl-X Protein; Down-Regulation; Nuclear Proteins; RNA-Binding Proteins; Cell Line; Up-Regulation; DNA-Binding Proteins; Alternative Splicing; 03 medical and health sciences; Heterogeneous-Nuclear Ribonucleoprotein Group A-B; Humans; Point Mutation; Protein Isoforms; RNA, Messenger; Phosphorylation; RNA, Small Interfering; Research Articles; Adaptor Proteins, Signal Transducing

The RNA-binding protein Sam68 is involved in apoptosis, but its cellular mRNA targets and its mechanism of action remain unknown. We demonstrate that Sam68 binds the mRNA for Bcl-x and affects its alternative splicing. Depletion of Sam68 by RNA interference caused accumulation of antiapoptotic Bcl-x(L), whereas its up-regulation increased the levels of proapoptotic Bcl-x(s). Tyrosine phosphorylation of Sam68 by Fyn inverted this effect and favored the Bcl-x(L) splice site selection. A point mutation in the RNA-binding domain of Sam68 influenced its splicing activity and subnuclear localization. Moreover, coexpression of ASF/SF2 with Sam68, or fusion with an RS domain, counteracted Sam68 splicing activity toward Bcl-x. Finally, Sam68 interacted with heterogenous nuclear RNP (hnRNP) A1, and depletion of hnRNP A1 or mutations that impair this interaction attenuated Bcl-x(s) splicing. Our results indicate that Sam68 plays a role in the regulation of Bcl-x alternative splicing and that tyrosine phosphorylation of Sam68 by Src-like kinases can switch its role from proapoptotic to antiapoptotic in live cells.