Archaeal Tuc1/Ncs6 Homolog Required for Wobble Uridine tRNA Thiolation Is Associated with Ubiquitin-Proteasome,Translation, and RNA Processing System Homologs |
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| Authors: | Nikita E Chavarria Sungmin Hwang Shiyun Cao Xian Fu Mary Holman Dina Elbanna Suzanne Rodriguez Deanna Arrington Markus Englert Sivakumar Uthandi Dieter S?ll Julie A Maupin-Furlow |
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| Institution: | 1Department of Microbiology and Cell Science, University of Florida, Gainesville, Florida, United States of America;2Genetics Institute, University of Florida, Gainesville, Florida, United States of America;3Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, United States of America;4Department of Chemistry, Yale University, New Haven, Connecticut, United States of America;Keio University, Japan |
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| Abstract: | While cytoplasmic tRNA 2-thiolation protein 1 (Tuc1/Ncs6) and ubiquitin-related modifier-1 (Urm1) are important in the 2-thiolation of 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) at wobble uridines of tRNAs in eukaryotes, the biocatalytic roles and properties of Ncs6/Tuc1 and its homologs are poorly understood. Here we present the first report of an Ncs6 homolog of archaea (NcsA of Haloferax volcanii) that is essential for maintaining cellular pools of thiolated tRNALysUUU and for growth at high temperature. When purified from Hfx. volcanii, NcsA was found to be modified at Lys204 by isopeptide linkage to polymeric chains of the ubiquitin-fold protein SAMP2. The ubiquitin-activating E1 enzyme homolog of archaea (UbaA) was required for this covalent modification. Non-covalent protein partners that specifically associated with NcsA were also identified including UbaA, SAMP2, proteasome activating nucleotidase (PAN)-A/1, translation elongation factor aEF-1α and a β-CASP ribonuclease homolog of the archaeal cleavage and polyadenylation specificity factor 1 family (aCPSF1). Together, our study reveals that NcsA is essential for growth at high temperature, required for formation of thiolated tRNALysUUU and intimately linked to homologs of ubiquitin-proteasome, translation and RNA processing systems. |
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