title: Analysis of spliceosomal proteins in Trypanosomatids reveals novel functions in mRNA processing.
abstract: In trypanosomatids, all mRNAs are processed via trans-splicing, although cis-splicing also occurs. In trans-splicing, a common small exon, the spliced leader (SL), which is derived from a small SL RNA species, is added to all mRNAs. Sm and Lsm proteins are core proteins that bind to U snRNAs and are essential for both these splicing processes. In this study, SmD3- and Lsm3-associated complexes were purified to homogeneity from Leishmania tarentolae. The purified complexes were analyzed by mass spectrometry, and 54 and 39 proteins were purified from SmD3 and Lsm complexes, respectively. Interestingly, among the proteins purified from Lsm3, no mRNA degradation factors were detected, as in Lsm complexes from other eukaryotes. The U1A complex was purified and mass spectrometry analysis identified, in addition to U1 small nuclear ribonucleoprotein (snRNP) proteins, additional co-purified proteins, including the polyadenylation factor CPSF73. Defects observed in cells silenced for U1 snRNP proteins suggest that the U1 snRNP functions exclusively in cis-splicing, although U1A also participates in polyadenylation and affects trans-splicing. The study characterized several trypanosome-specific nuclear factors involved in snRNP biogenesis, whose function was elucidated in Trypanosoma brucei. Conserved factors, such as PRP19, which functions at the heart of every cis-spliceosome, also affect SL RNA modification; GEMIN2, a protein associated with SMN (survival of motor neurons) and implicated in selective association of U snRNA with core Sm proteins in trypanosomes, is a master regulator of snRNP assembly. This study demonstrates the existence of trypanosomatid-specific splicing factors but also that conserved snRNP proteins possess trypanosome-specific functions.
authors.affiliation: Mina and Everard Goodman Faculty of Life Sciences and Advanced Materials and Nanotechnology Institute, Bar-Ilan University, Ramat-Gan 52900, Israel.
authors.forename: Itai Dov
authors.initials: ID
authors.lastname: Tkacz
authors.forename: Sachin Kumar
authors.initials: SK
authors.lastname: Gupta
authors.forename: Vadim
authors.initials: V
authors.lastname: Volkov
authors.forename: Mali
authors.initials: M
authors.lastname: Romano
authors.forename: Tomer
authors.initials: T
authors.lastname: Haham
authors.forename: Pawel
authors.initials: P
authors.lastname: Tulinski
authors.forename: Ilana
authors.initials: I
authors.lastname: Lebenthal
authors.forename: Shulamit
authors.initials: S
authors.lastname: Michaeli
mesh_terms: D001692:Biological Transport; D002460:Cell Line; D007891:Leishmania / Q000166:cytology* / Q000235:genetics*; D013058:Mass Spectrometry; D026723:Polyadenylation; D015800:Protozoan Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000378:metabolism; D020038:RNA, Spliced Leader / Q000096:biosynthesis; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D013045:Species Specificity; D017381:Spliceosomes / Q000378:metabolism*
chemical_list: D015800:Protozoan Proteins; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; D020038:RNA, Spliced Leader; D017411:Ribonucleoproteins, Small Nuclear