RRM
2009 Rustbelt RNA Meeting
RRM
|
|
|
|
|
Poster abstracts
Abstract:
Trm10 is a highly conserved SAM-dependent methyltransferase which catalyzes N1-methylation at the G9 position of tRNA substrates. While all m1G9 methylation is catalyzed by a single TRM10 in yeast, there are up to three Trm10 homologs in metazoa, including humans, for which the function has not yet been investigated. In humans, one of these homologs (RG9MTD1) is targeted to the mitochondria where it has unexpectedly been implicated in a novel proteinacious RNase P complex. Although the biochemical function of Trm10 in this complex is unknown, its presence is absolutely required for RNase P processing activity. Furthermore, the function of the remaining two, presumably cytoplasmic, homologs (RG9MTD2 and RG9MTD3) is unknown. Thus, the goal of this work is to investigate the biochemical and biological function of the multiple human Trm10 homologs.
To that end we have begun to characterize one of the human Trm10 homologs, RG9MTD2, which shares the highest sequence homology to yeast Trm10. Using site specifically-labeled yeast Gly, Val and Leu tRNAs and assay conditions optimized with respect to [Mg] and pH, we have shown that RG9MTD2, like yeast Trm10, catalyzes m1G9 formation. In addition, RG9MTD2 follows the same pattern of in vitro activity observed with yeast Trm10, modifying Gly and Val but not Leu tRNAs, which is unexpected since a different subset of tRNAs are m1G9 modified in yeast versus human cells. Previously, similar questions of substrate specificity have been raised regarding yeast Trm10, which by some unknown mechanism modifies some, but not all, G9-containing tRNAs in vivo. Therefore, the basis for Trm10 substrate specificity in both yeast and humans is unclear, and future investigations of this issue, as well as similar investigations of the remaining two human homologs (RG9MTD1 and RG9MTD3) are in progress.
Keywords: tRNA methyltransferase, yeast, enzyme catalysis
