Faculty Opinions recommendation of Yeast telomerase RNA: a flexible scaffold for protein subunits.

Telomerase RNA contains a template for synthesizing telomeric DNA and has been proposed to act as a flexible scaffold for holoenzyme protein subunits in the RNP. In Saccharomyces cerevisiae, the telomerase RNA, TLC1, is bound by the Sm7 protein complex, which is required for stabilization of the predominant, non-polyadenylated (poly(A)–) TLC1 isoform. However, it remains unclear (1) whether Sm7 retains this function when its binding site is repositioned within TLC1, as has been shown for other TLC1-binding telomerase subunits, and (2) how Sm7 stabilizes poly(A)– TLC1. Here, we first show that Sm7 can stabilize poly(A)– TLC1 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 show that when an Sm site is inserted 5′ of its native position and the native site is mutated, Sm7 stabilizes shorter forms of poly(A)– TLC1 in a manner corresponding to how far upstream the new site was inserted, providing strong evidence that Sm7 binding to TLC1 controls where the mature poly(A)– 3′ is formed by directing a 3′-to-5′ processing mechanism. 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.

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Yeast Telomerase RNA Flexibly Scaffolds Protein Subunits: Results and Repercussions

Molecules ◽

10.3390/molecules25122750 ◽

2020 ◽

Vol 25 (12) ◽

pp. 2750 ◽

Cited By ~ 1

Author(s):

David C. Zappulla

Keyword(s):

Saccharomyces Cerevisiae ◽

Noncoding Rna ◽

Noncoding Rnas ◽

Telomerase Rna ◽

Protein Subunits ◽

Rna Molecules ◽

Yeast Telomerase ◽

Opportune Time ◽

Rnp Complex ◽

Functional Features

It is said that “hindsight is 20-20,” so, given the current year, it is an opportune time to review and learn from experiences studying long noncoding RNAs. Investigation of the Saccharomyces cerevisiae telomerase RNA, TLC1, has unveiled striking flexibility in terms of both structural and functional features. Results support the “flexible scaffold” hypothesis for this 1157-nt telomerase RNA. This model describes TLC1 acting as a tether for holoenzyme protein subunits, and it also may apply to a plethora of RNAs beyond telomerase, such as types of lncRNAs. In this short perspective review, I summarize findings from studying the large yeast telomerase ribonucleoprotein (RNP) complex in the hope that this hindsight will sharpen foresight as so many of us seek to mechanistically understand noncoding RNA molecules from vast transcriptomes.

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Repositioning the Sm-binding site in S. cerevisiae telomerase RNA reveals RNP organizational flexibility and Sm-directed 3′-end formation

10.1101/167361 ◽

2017 ◽

Cited By ~ 2

Author(s):

Evan P. Hass ◽

David C. Zappulla

Keyword(s):

Saccharomyces Cerevisiae ◽

Binding Site ◽

Strong Evidence ◽

Binding Sites ◽

Telomerase Rna ◽

Telomeric Dna ◽

Protein Subunits ◽

Yeast Telomerase ◽

Rna Biogenesis

ABSTRACTTelomerase 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.

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Maturation and shuttling of the yeast telomerase RNP: assembling something new using recycled parts

Current Genetics ◽

10.1007/s00294-021-01210-2 ◽

2021 ◽

Author(s):

Louise Bartle ◽

Yulia Vasianovich ◽

Raymund J. Wellinger

Keyword(s):

Budding Yeast ◽

New Combination ◽

Telomerase Rna ◽

Complete Understanding ◽

Protein Subunits ◽

Yeast Telomerase ◽

Quality Checks ◽

Human Telomerase ◽

Rna Maturation ◽

Transport Factors

AbstractAs the limiting component of the budding yeast telomerase, the Tlc1 RNA must undergo multiple consecutive modifications and rigorous quality checks throughout its lifecycle. These steps will ensure that only correctly processed and matured molecules are assembled into telomerase complexes that subsequently act at telomeres. The complex pathway of Tlc1 RNA maturation, involving 5'- and 3'-end processing, stabilisation and assembly with the protein subunits, requires at least one nucleo-cytoplasmic passage. Furthermore, it appears that the pathway is tightly coordinated with the association of various and changing proteins, including the export factor Xpo1, the Mex67/Mtr2 complex, the Kap122 importin, the Sm7 ring and possibly the CBC and TREX-1 complexes. Although many of these maturation processes also affect other RNA species, the Tlc1 RNA exploits them in a new combination and, therefore, ultimately follows its own and unique pathway. In this review, we highlight recent new insights in maturation and subcellular shuttling of the budding yeast telomerase RNA and discuss how these events may be fine-tuned by the biochemical characteristics of the varying processing and transport factors as well as the final telomerase components. Finally, we indicate outstanding questions that we feel are important to be addressed for a complete understanding of the telomerase RNA lifecycle and that could have implications for the human telomerase as well.

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Essential Regions of Saccharomyces cerevisiae Telomerase RNA: Separate Elements for Est1p and Est2p Interaction

Molecular and Cellular Biology ◽

10.1128/mcb.22.7.2366-2374.2002 ◽

2002 ◽

Vol 22 (7) ◽

pp. 2366-2374 ◽

Cited By ~ 70

Author(s):

April J. Livengood ◽

Arthur J. Zaug ◽

Thomas R. Cech

Keyword(s):

Saccharomyces Cerevisiae ◽

Nucleotide Sequence ◽

Dna Synthesis ◽

Active Site ◽

Telomerase Rna ◽

Telomeric Dna ◽

Protein Subunits ◽

Essential Region ◽

Selection For

ABSTRACT The Saccharomyces cerevisiae telomerase RNA subunit is encoded by the TLC1 gene. A selection for viable alleles of TLC1 RNA from a large library of random deletion alleles revealed that less than half (∼0.5 kb of the ∼1.3-kb RNA) is required for telomerase function in vivo. The main essential region (430 nucleotides), which contains the template for telomeric DNA synthesis, was required for coimmunoprecipitation with Est1p and Est2p. Furthermore, the subregion required for interaction with Est1p, the telomerase recruitment subunit, differed from those required for interaction with Est2p, the reverse transcriptase subunit. Two regions of the RNA distant from the template in the nucleotide sequence were required for Est2p binding, but the template itself was not. Having the RNA secured to the protein away from the template is proposed to facilitate the translocation of the RNA template through the active site. More generally, our results support a role for the telomerase RNA serving as a scaffold for binding key protein subunits.

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