PT  - JOURNAL ARTICLE
AU  - Evan P. Hass
AU  - David C. Zappulla
TI  - Repositioning the Sm-binding site in <em>S. cerevisiae</em> telomerase RNA reveals RNP organizational flexibility and Sm-directed 3′-end formation
AID  - 10.1101/167361
DP  - 2017 Jan 01
TA  - bioRxiv
PG  - 167361
4099  - http://biorxiv.org/content/early/2017/07/22/167361.short
4100  - http://biorxiv.org/content/early/2017/07/22/167361.full
AB  - Telomerase RNA contains a template for synthesizing telomeric DNA by reverse transcription and has been proposed to act as a flexible scaffold for holoenzyme protein subunits in the RNP. In Saccharomyces cerevisiae, the telomerase subunits Est1 and Ku bind to the telomerase RNA, TLC1, and it has been shown that these proteins still function when their binding sites are repositioned within the RNA. TLC1 is also bound by the Sm7 protein complex, which is required for stabilization of the predominant, non-polyadenylated (poly(A)–) TLC1 isoform. Here, we first show that Sm7 can perform this function even when its binding site is repositioned via circular permutation to several different positions within TLC1, further supporting the conclusion that the telomerase holoenzyme is organizationally flexible. Next, we tested the hypothesis that the location of the Sm7-binding site relative to the 3′ end is contrastingly important. When we moved the Sm site to locations 5′ of its native position, we observed that this stabilized shorter forms of poly(A)– TLC1 in a manner precisely corresponding to how far upstream the Sm site was moved. This provides strong evidence that the location of Sm7 binding to TLC1 controls where the mature poly(A)– 3′ end is formed. In summary, our results show that Sm7 and the 3′ end of yeast telomerase RNA comprise an organizationally flexible module within the telomerase RNP and provide insights into the mechanistic role of Sm7 in telomerase RNA biogenesis.