SUMO CONJUGATION TO SPLICEOSOMAL PROTEINS

POZZI, BERTA; BRAGADO, LAUREANO; MAMMI, PABLO; RISSO, GUILLERMO; SREBROW, ANABELLA

Mar del Plata

Congreso; LI Reunión de la Sociedad Argentina de Investigación Bioquímica y Biología Molecular; 2015

Since previous work from our laboratory has revealed the splicing factor SRSF1 as a regulator of the SUMOconjugation pathway, we started to explore a possible link between SUMO and the splicing process, focusing on thespliceosome, the multimegadalton ribonucleoprotein machine responsible for it. We found that the addition ofrecombinant SUMO-conjugating enzyme to an in vitro splicing reaction accelerates the appearance of mature mRNAwhile a SUMO protease retards it. By Mass Spec analysis of anti-SUMO immunoprecipitated proteins obtained frompre-mRNA-bound complexes at different steps of the splicing reaction, we identify several spliceosomal SUMOsubstrates, such as Prp3, Prp28 and Snu114, which we have validated in cultured cells. After identifying SUMOattachment sites in Prp3, we obtained a SUMOylation mutant (Prp3 K289,559R) that fails to increase splicingefficiency when overexpressed, and is unable to co-precipitate U2 and U5 snRNA as well as the spliceosomalproteins SF3a and Snu114, compared to the wt version. We are currently validating the hypothesis that the Prp3SUMOylation mutant is unable to achieve similar splicing efficiency levels to the wt protein due to its diminishedrecruitment to active spliceosomes. We propose that SUMO conjugation to spliceosomal proteins could play a role insplicing dynamics by modulating protein-protein and/or protein-RNA interactions.