Actin-binding protein ABP140 is a methyltransferase for 3-methylcytidine at position 32 of tRNAs in Saccharomyces cerevisiae

Abstract

Transfer RNAs contain various modified nucleotides that are introduced enzymatically at the post-transcriptional level. In Saccharomyces cerevisiae, 3-methylcytidine (m3C) is found at position 32 of the tRNAs for Thr and Ser. We used a systematic reverse genetic approach combined with mass spectrometry (ribonucleome analysis), and identified the actin-binding protein ABP140 as the protein responsible for m3C formation in both tRNA Thr1 and tRNASer1. ABP140 consists of an N-terminal actinbinding sequence and a C-terminal S-adenosylmethionine (Ado-Met) binding motif. Deletion of the actin-binding sequence in ABP140 did not affect m 3C formation, indicating that subcellular localization of ABP140 to actin filaments is not involved in tRNA modification. m3C formation in tRNAThr1 could be reconstituted using recombinant Abp140p in the presence of Ado-Met, whereas m3C did not form in tRNASer1 in vitro, indicating the absence of a factor(s) required for tRNASer1 m3C formation. Thus, ABP140 has been designated TRM140 according to the preferred nomenclature. In addition, we observed a specific reduction of m3C formation in HeLa cells by siRNA-mediated knock down of the human ortholog of TRM140. Copyright © 2011 RNA Society.