Box H/ACA snoRNAs are preferred substrates for the trimethylguanosine synthase in the divergent unicellular eukaryote Trichomonas vaginalis Journal Article
| Authors: | Simoes-Barbosa, A.; Chakrabarti, K.; Pearson, M.; Benarroch, D.; Shuman, S.; Johnson, P. J. |
| Article Title: | Box H/ACA snoRNAs are preferred substrates for the trimethylguanosine synthase in the divergent unicellular eukaryote Trichomonas vaginalis |
| Abstract: | The 2,2,7-trimethylguanosine caps of eukaryal snRNAs and snoRNA are formed by the enzyme Tgs1, which catalyzes sequential guanine-N2 methylations of m 7G caps. Atypically, in the divergent unicellular eukaryote Trichomonas vaginalis, spliceosomal snRNAs lack a guanosine cap and the recombinant T. vaginalis trimethylguanosine synthase (TvTgs) produces only m 2,7G in vitro. Here, we show by direct metabolic labeling that endogenous T. vaginalis RNAs contain m 7G, m 2,7G, and m 2,2,7G caps. Immunodepletion of TvTgs from cell extracts and TvTgs add-back experiments demonstrate that TvTgs produces m 2,7G and m 2,2,7G caps. Expression of TvTgs in yeast tgs1Δ cells leads to the formation of m 2,7G and m 2,2,7G caps and complementation of the lethality of a tgs1Δ mud2Δ strain. Whereas TvTgs is present in the nucleus and cytosol of T. vaginalis cells, TMG-containing RNAs are localized primarily in the nucleolus. Molecular cloning of anti-TMG affinity-purified T. vaginalis RNAs identified 16 box H/ACA snoRNAs, which are implicated in guiding RNA pseudouridylation. The ensemble of new T. vaginalis H/ACA snoRNAs allowed us to predict and partially validate an extensive map of pseudouridines in T. vaginalis rRNA. Published by Cold Spring Harbor Laboratory Press. Copyright © 2012 RNA Society. |
| Keywords: | protein expression; unclassified drug; methylation; nonhuman; enzyme activity; molecular cloning; methyltransferases; molecular sequence data; saccharomyces cerevisiae; guanosine; rna caps; sequence alignment; mimivirus; substrate specificity; immunoprecipitation; cellular distribution; base sequence; cell nucleus; nucleic acid conformation; eukaryotic cell; ribosome rna; small nucleolar rna; rna, small nucleolar; biological transport; yeast cell; cytosol; lethality; enzyme synthesis; biogenesis; nucleolus; trimethylguanosine synthase; rna capping; synthetase; rna methylation; cell extract; small nuclear rna; polyclonal antibody; rna purification; pseudouridine; capping; snorna; trichomonas vaginalis |
| Journal Title: | RNA |
| Volume: | 18 |
| Issue: | 9 |
| ISSN: | 1355-8382 |
| Publisher: | Cold Spring Harbor Laboratory Press |
| Date Published: | 2012-09-01 |
| Start Page: | 1656 |
| End Page: | 1665 |
| Language: | English |
| DOI: | 10.1261/rna.034249.112 |
| PROVIDER: | scopus |
| PMCID: | PMC3425780 |
| PUBMED: | 22847815 |
| DOI/URL: | |
| Notes: | --- - "Export Date: 4 September 2012" - "CODEN: RNARF" - "Source: Scopus" |
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