Role of a 300-kilodalton nuclear complex in the maturation of Trypanosoma brucei initiator methionyl-tRNA.
tRNAs are transcribed as precursors containing 5' leader and 3' extensions that are removed by a series of posttranscriptional processing reactions to yield functional mature tRNAs. Here, we examined the maturation pathway of tRNA(Met) in Trypanosoma brucei, an early divergent unicellular eukaryote. We identified an approximately 300-kDa complex located in the nucleus of T. brucei that is required for trimming the 5' leader of initiator tRNA(Met) precursors. One of the subunits of the complex (T. brucei MT40 [TbMT40]) is a putative methyltransferase and a homolog of Saccharomyces cerevisiae Gcd14, which is essential for 1-methyladenosine modification in tRNAs. Down-regulation of TbMT40 by RNA interference resulted in the accumulation of precursor initiator tRNA(Met) containing 5' extensions but processed 3' ends. In addition, immunoprecipitations with anti-La antibodies revealed initiator tRNA(Met) molecules with 5' and 3' extensions in TbMT40-silenced cells, albeit at a much lower level. Interestingly, silencing of TbMT40, as well as of TbMT53, a second subunit of the complex, led to an increase in the levels of mature elongator tRNA(Met). Taken together, our data provide a glance at the maturation of tRNAs in parasitic protozoa and suggest that at least for initiator tRNA(Met), 3' trimming precedes 5' processing.
Department of Epidemiology and Public Health, Yale University Medical School, 295 Congress Avenue, New Haven, CT 06536-0812, USA.
George K
GK
Arhin
Shuiyuan
S
Shen
Henriette
H
Irmer
Elisabetta
E
Ullu
Christian
C
Tschudi
D000595:Amino Acid Sequence; D000818:Animals; D020868:Gene Silencing; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012323:RNA Processing, Post-Transcriptional; D016053:RNA, Protozoan / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014346:Trypanosoma brucei brucei / Q000235:genetics* / Q000378:metabolism; D012359:tRNA Methyltransferases
D046911:Macromolecular Substances; D021122:Protein Subunits; D015800:Protozoan Proteins; D016053:RNA, Protozoan; D012358:RNA, Transfer, Met; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012359:tRNA Methyltransferases; C119384:GCD14 protein, S cerevisiae