Characterization of subunits of the RNA polymerase I complex in Trypanosoma brucei.
The Trypanosoma brucei homologue of the RNA polymerase I (RNA Pol I) subunit Rpa12p of Saccharomyces cerevisiae was cloned and characterized. This protein did not appear to be essential for growth in either bloodstream or procyclic forms of the parasite. Trypanosomes expressing a C-terminal tagged version of TbRPA12 were generated in order to purify RNA Pol I from both developmental stages. Tandem affinity purification (TAP) revealed a number of proteins associating with TbRPA12, some of which appeared to be stage-specific. Mass spectrometry allowed the identification of four subunits in addition to TbRPA12, namely TbRPA1, TbRPA2, TbRPC40 and one isoform of TbRPB5 (Tb1RPB5), as well as an unknown 30kDa protein and histones H2A and H3. Whereas these studies demonstrated that TbRPA1 was phosphorylated, no evidence for phosphorylation of TbRPA2 was found.
Walgraffe
David
D
Laboratory of Molecular Parasitology, Department of Molecular Biology, IBMM, Free University of Brussels, 12, rue des Professeurs Jeener et Brachet, B-6041 Gosselies, Belgium.
Devaux
Sara
S
Lecordier
Laurence
L
Dierick
Jean-François
JF
Dieu
Marc
M
Van den Abbeele
Jan
J
Pays
Etienne
E
Vanhamme
Luc
L
D000595:Amino Acid Sequence; D000818:Animals; D000940:Antigenic Variation; D018507:Gene Expression Regulation, Developmental; D008018:Life Cycle Stages; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D012318:RNA Polymerase I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
D021122:Protein Subunits; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C423311:RPA12 protein, S cerevisiae; D012318:RNA Polymerase I