title: The polyadenylation complex of Trypanosoma brucei: Characterization of the functional poly(A) polymerase.
abstract: The generation of mature mRNA in the protozoan parasite Trypanosoma brucei requires coupled polyadenylation and trans splicing. In contrast to other eukaryotes, we still know very little on components, mechanisms, and dynamics of the 3' end-processing machinery in trypanosomes. To characterize the catalytic core of the polyadenylation complex in T. brucei, we first identified the poly(A) polymerase [Tb927.7.3780] as the major functional, nuclear-localized enzyme in trypanosomes. In contrast, another poly(A) polymerase, encoded by an intron-containing gene [Tb927.3.3160], localizes mainly in the cytoplasm and appears not to be functional in general 3' end processing of mRNAs. Based on tandem-affinity purification with tagged CPSF160 and mass spectrometry, we identified ten associated components of the trypanosome polyadenylation complex, including homologues to all four CPSF subunits, Fip1, CstF50/64, and Symplekin, as well as two hypothetical proteins. RNAi-mediated knockdown revealed that most of these factors are essential for growth and required for both in vivo polyadenylation and trans splicing, arguing for a general coupling of these two mRNA-processing reactions.
authors.affiliation: a Institute of Biochemistry, Justus Liebig University of Giessen , D-35392 Giessen , Germany.
authors.forename: Henrik
authors.initials: H
authors.lastname: Koch
authors.affiliation: b Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry , D-37077 Göttingen , Germany.
authors.forename: Monika
authors.initials: M
authors.lastname: Raabe
authors.affiliation: b Bioanalytical Mass Spectrometry Group, Max Planck Institute for Biophysical Chemistry , D-37077 Göttingen , Germany.|c Bioanalytics Group, Institute for Clinical Chemistry, University Medical Center Göttingen , D-37075 Göttingen , Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Urlaub
authors.affiliation: a Institute of Biochemistry, Justus Liebig University of Giessen , D-35392 Giessen , Germany.
authors.forename: Albrecht
authors.initials: A
authors.lastname: Bindereif
authors.affiliation: a Institute of Biochemistry, Justus Liebig University of Giessen , D-35392 Giessen , Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Preußer
mesh_terms: D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D007438:Introns; D011061:Poly A / Q000235:genetics; D026723:Polyadenylation / Q000235:genetics*; D012333:RNA, Messenger; D020040:Trans-Splicing / Q000235:genetics*; D014346:Trypanosoma brucei brucei / Q000235:genetics*
chemical_list: D012333:RNA, Messenger; D011061:Poly A; D012321:DNA-Directed RNA Polymerases
keywords: CPSF; Trypanosoma brucei; mRNA processing; poly(A) polymerase; polyadenylation; trans splicing