The RNA-binding protein Sam68 modulates the alternative splicing of Bcl-x.
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.
Paronetto
Maria Paola
MP
Department of Public Health and Cell Biology, Section of Anatomy, University of Rome Tor Vergata, 00133 Rome, Italy.
Achsel
Tilman
T
Massiello
Autumn
A
Chalfant
Charles E
CE
Sette
Claudio
C
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics* / Q000378:metabolism*; D017398:Alternative Splicing / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D015536:Down-Regulation / Q000502:physiology; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017354:Point Mutation / Q000235:genetics; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D000068103:Serine-Arginine Splicing Factors; D015854:Up-Regulation / Q000502:physiology; D051020:bcl-X Protein / Q000235:genetics* / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C494811:BCL2L1 protein, human; D004268:DNA-Binding Proteins; C106957:FYB1 protein, human; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; C489559:KHDRBS1 protein, human; D009687:Nuclear Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D051020:bcl-X Protein; D000068103:Serine-Arginine Splicing Factors