REGULATION OF THE INTEGRATOR COMPLEX BY SUMO CONJUGATION

BRAGADO, LAUREANO; MAGALNIK, MELINA; SREBROW ANABELLA

virtual

Congreso; SAIB - SAMIGE Joint meeting; 2021

Sociedad Argentina de Investigaciones en Bioquimica y Biología Molecular (SAIB)

In addition to protein-coding genes, RNA polymerase II (Pol II) transcribes numerous genes that correspond to non-coding RNAs, including small nuclear RNAs (snRNAs). snRNAs are not only a fundamental component of snRNPs, the ribonucleoprotein particles that conform the spliceosome but also some of them are necessary for Pol II transcriptional activity. Although the functions of snRNAs are well understood, the regulation of their biogenesis is still a matter of deep investigation. snRNA genes share common features with protein-coding genes, including the relative positioning of elements that control transcription and RNA processing. However, there are important differences in the set of proteins required for proper expression of these two gene types. Particularly, while CPSF complex is in charge of 3? end formation and polyadenylation of pre-mRNAs, the Integrator complex is responsible for 3?-end processing of snRNAs. Moreover, this multimeric complex composed by 14 subunits is also known to regulate the expression of other transcripts, such as PROMPTs and eRNAs. Nevertheless, how Integrator activity is regulated remains unclear. SUMOylation is a reversible post-translational modification consisting in the conjugation of SUMO (small ubiquitin-related modifier) peptides to different target proteins. It mainly modulates intra- and inter- molecular interactions and consequently the function of diverse cellular proteins. Years ago, our lab reported the influence of SUMOylation of spliceosomal proteins on spliceosome assembly and catalytic activity. We are currently studying the involvement of SUMO conjugation in snRNA biogenesis. We have observed that modifying global SUMOylation levels in mammalian cell lines alters the proportion of nascent vs. mature snRNAs. Furthermore, we have shown that several subunits of the Integrator complex are modified by SUMO. In particular, INTS11 SUMOylation is regulated by USPL1, a SUMO-isopeptidase that localizes in Cajal bodies, nuclear compartments involved in snRNA expression and snRNP maturation. Having identified the target lysine residues of this modification within INTS11 and generated a SUMOylation deficient mutant of this protein, we found that this mutant is unable to achieve proper 3? end processing of precursor snRNAs. While INTS11 SUMOylation-deficient mutant preserves its interaction with the other two subunits of the catalytic module (INTS11/INTS9/INTS4), this post-translational modification seems to be important for proper assembly of the whole Integrator complex. Moreover, INTS11 SUMOylation mutant showed decreased recruitment to chromatin and a more cytoplasmatic localization, compared to the wild-type protein. Taken together, these results represent a novel regulatory mechanism for the function of the Integrator complex.