In vivo architecture of the telomerase RNA catalytic core in Trypanosoma brucei.

Abstract

Telomerase is a unique ribonucleoprotein (RNP) reverse transcriptase that utilizes its cognate RNA molecule as a template for telomere DNA repeat synthesis. Telomerase contains the reverse transcriptase protein, TERT and the template RNA, TR, as its core components. The 5'-half of TR forms a highly conserved catalytic core comprising of the template region and adjacent domains necessary for telomere synthesis. However, how telomerase RNA folding takes place in vivo has not been fully understood due to low abundance of the native RNP. Here, using unicellular pathogen Trypanosoma brucei as a model, we reveal important regional folding information of the native telomerase RNA core domains, i.e. TR template, template boundary element, template proximal helix and Helix IV (eCR4-CR5) domain. For this purpose, we uniquely combined in-cell probing with targeted high-throughput RNA sequencing and mutational mapping under three conditions: in vivo (in WT and TERT-/- cells), in an immunopurified catalytically active telomerase RNP complex and ex vivo (deproteinized). We discover that TR forms at least two different conformers with distinct folding topologies in the insect and mammalian developmental stages of T. brucei. Also, TERT does not significantly affect the RNA folding in vivo, suggesting that the telomerase RNA in T. brucei exists in a conformationally preorganized stable structure. Our observed differences in RNA (TR) folding at two distinct developmental stages of T. brucei suggest that important conformational changes are a key component of T. brucei development.

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Published Version (Please cite this version)

10.1093/nar/gkab1042

Publication Info

Dey, Abhishek, Anais Monroy-Eklund, Kaitlin Klotz, Arpita Saha, Justin Davis, Bibo Li, Alain Laederach, Kausik Chakrabarti, et al. (2021). In vivo architecture of the telomerase RNA catalytic core in Trypanosoma brucei. Nucleic acids research, 49(21). pp. 12445–12466. 10.1093/nar/gkab1042 Retrieved from https://hdl.handle.net/10161/31419.

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Davis

Justin Davis

Postdoctoral Associate

I am a postdoctoral associate working in the Derbyshire lab in the Department of Chemistry. I graduated with my Ph. D. in Molecular Biology from the University of North Carolina at Charlotte in 2024. I am fascinated by parasite biology and the mechanisms these organisms use to adapt to their host. In my free time, I enjoy playing the acoustic guitar and studying languages. 



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