The 3-methylcytidine (m³C) modification is ubiquitous in eukaryotic tRNA, widely found at C₃₂ in the anticodon loop of tRNA^Thr, tRNA^Ser, and some tRNA^Arg species, as well as in the variable loop (V-loop) of certain tRNA^Ser species. In the yeast Saccharomyces cerevisiae, formation of m³C₃₂ requires Trm140 for six tRNA substrates, including three tRNA^Thr species and three tRNA^Ser species, whereas in Schizosaccharomyces pombe, two Trm140 homologs are used, one for tRNA^Thr and one for tRNA^Ser The occurrence of a single Trm140 homolog is conserved broadly among Ascomycota, whereas multiple Trm140-related homologs are found in metazoans and other fungi. We investigate here how S. cerevisiae Trm140 protein recognizes its six tRNA substrates. We show that Trm140 has two modes of tRNA substrate recognition. Trm140 recognizes G₃₅-U₃₆-t⁶A₃₇ of the anticodon loop of tRNA^Thr substrates, and this sequence is an identity element because it can be used to direct m³C modification of tRNA^Phe However, Trm140 recognition of tRNA^Ser substrates is different, since their anticodons do not share G₃₅-U₃₆ and do not have any nucleotides in common. Rather, specificity of Trm140 for tRNA^Ser is achieved by seryl-tRNA synthetase and the distinctive tRNA^Ser V-loop, as well as by t⁶A₃₇ and i⁶A₃₇ We provide evidence that all of these components are important in vivo and that seryl-tRNA synthetase greatly stimulates m³C modification of tRNA^Ser(CGA) and tRNA^Ser(UGA) in vitro. In addition, our results show that Trm140 binding is a significant driving force for tRNA modification and suggest separate contributions from each recognition element for the modification.

• PMID: 28003514
• DOI full text
• PMC full text
• PubMed
• PubTator

Reference Type

Journal Article

Authors

Han L, Marcus E, D'Silva S, Phizicky EM

Primary Lit For

Additional Lit For

Review For