A previously unidentified activity of yeast and mouse RNA:pseudouridine synthases 1 (Pus1p) on tRNAs.
Mouse pseudouridine synthase 1 (mPus1p) was the first vertebrate RNA:pseudouridine synthase that was cloned and characterized biochemically. The mPus1p was previously found to catalyze Psi formation at positions 27, 28, 34, and 36 in in vitro produced yeast and human tRNAs. On the other hand, the homologous Saccharomyces cerevisiae scPus1p protein was shown to modify seven uridine residues in tRNAs (26, 27, 28, 34, 36, 65, and 67) and U44 in U2 snRNA. In this work, we expressed mPus1p in yeast cells lacking scPus1p and studied modification of U2 snRNA and several yeast tRNAs. Our data showed that, in these in vivo conditions, the mouse enzyme efficiently modifies yeast U2 snRNA at position 44 and tRNAs at positions 27, 28, 34, and 36. However, a tRNA:Psi26-synthase activity of mPus1p was not observed. Furthermore, we found that both scPus1p and mPus1p, in vivo and in vitro, have a previously unidentified activity at position 1 in cytoplasmic tRNAArg(ACG). This modification can take place in mature tRNA, as well as in pre-tRNAs with 5' and/or 3' extensions. Thus, we identified the protein carrying one of the last missing yeast tRNA:Psi synthase activities. In addition, our results reveal an additional activity of mPus1p at position 30 in tRNA that scPus1p does not possess.
Behm-Ansmant
Isabelle
I
Laboratoire de Maturation des ARN et Enzymologie Moléculaire, UMR 7567 CNRS-UHP, Nancy I, Faculté des Sciences, BP 239, 54506 Vandoeuvre-les-Nancy Cedex, France.
Massenet
Séverine
S
Immel
Françoise
F
Patton
Jeffrey R
JR
Motorin
Yuri
Y
Branlant
Christiane
C
D000818:Animals; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism*; D066298:In Vitro Techniques; D051379:Mice; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism*; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
D012322:RNA Precursors; D012331:RNA, Fungal; D012342:RNA, Small Nuclear; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C405484:U2 small nuclear RNA; D012343:RNA, Transfer; D006836:Hydro-Lyases; C022479:pseudouridylate synthetase