Targeted RNA 2'-O-methylation for Regulating Telomerase Activity
The goal of this project is to fully develop a novel technique, namely, targeted RNA 2'-O-methylation, for regulating telomerase activity in vivo, as well as to understand how 2'-O-methylation, as a naturally occurring process, is adopted by cells to regulate telomerase activity. Our recent research on small RNA-guided RNA modifications has produced the exciting realization that such modifications are widespread and heavily utilized in eukaryotic cells, leading to the site-specific introduction of modifications (e.g., 2'-O-methylation) into telomerase RNA (TLC1 in yeast). Using box C/D RNA-guided 2'-O-methylation, we have artificially targeted TLC1 at six sites within the conserved pseudoknot region, and identified at least three sites where modification led to altered telomerase activity in vivo (two led to enhancement and one to reduction). Building on these results, we plan to extend our modification targets to each and every site throughout the TLC1 sequence.
It is quite likely that this RNA-guided 2'-O-methylation mechanism, when artificially applied to telomerase RNA, will offer us a unique opportunity to identify 2'-OH moieties important for telomerase function, and to thereby manipulate telomerase activity in vivo. Identification of the important 2'-OH moieties will also allow us to delineate how they contribute to telomerase function. In addition to artificial 2'-O-methylation, we have recently discovered that the yeast telomerase RNA TLC1 is naturally 2'-O-methylated. This exciting discovery has prompted us to further investigate the mechanism of naturally occurring 2'-O-methylation in TLC1. At the moment, we are conducting experiments directed toward identifying the enzyme responsible for the natural modification. Identification of the enzyme responsible for induced 2'-O-methylation will eventually lead to the elucidation of the mechanism of this modification, and will thus significantly advance our understanding of the regulation of telomerase activity and aging.
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