HOW TO TRAIN OUR CELLS TO BECOME YOUNGER – QUANTITATIVE BIOPHYSICS OF HUMAN TELOMERASE AND ITS GUARD SHELTERIN

Publikace nespadá pod Přírodovědeckou fakultu, ale pod Středoevropský technologický institut. Oficiální stránka publikace je na webu muni.cz.

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HOFR Ctirad NEČASOVÁ Ivona JANOUŠKOVÁ Eliška JANOVIČ Tomáš STOJASPAL Martin VEVERKA Pavel

Druh Článek ve sborníku
Konference 15 th International Interdisciplinary Meeting on Bioanalysis - Conference Proceedings
Fakulta / Pracoviště MU

Středoevropský technologický institut

Citace
WWW http://www.ce-ce.org
Klíčová slova Telomere; SHELTERIN
Popis Telomere maintenance is a highly coordinated process that controls cell aging. Misregulation of telomere maintenance is linked to cancer and telomere-shortening syndromes. Recent studies have shown that the TEL-patch is a cluster of amino acids onthe surface of the shelterin component TPP1 that is essential for the recruitment of telomerase to the telomere in human cells. The Cech laboratory (Colorado University Boulder) and our laboratory (Masaryk University) optimized an in vitro assay to quantitatively measure binding of the TEL-patch to telomerase and extension of the first telomeric repeat. We quantified how the TEL-patch contributes to the translocation and stabilizes the association between telomerase and telomeric DNA substrates, providing a molecular explanation for its contributions to telomerase recruitment and action. Additionally, we quantitatively described interactions of TRF2 - central shelterin subunit that folds human telomeres into loops to prevent unwanted DNA repair and chromosome end joining. We found that the basic B-domain of TRF2 stabilizes the displacement loop (D-loop) and thus reduces unwinding by RPA and BLM helicase, whereas the formation of the RAP1–TRF2 complex restores DNA unwinding. To understand how the B-domain of TRF2 affects DNA binding and D-loop processing, we analyzed DNA binding of full-length TRF2 and a truncated TRF2 construct lacking the B-domain. We found that the Bdomain improves TRF2’s binding to DNA via enhanced long-range electrostatic interactions. We determined a structural envelope model revealing that the B-domain is flexible in solution but becomes rigid upon binding to telomeric DNA. We propose a mechanism for how the Bdomain stabilizes the D-loop and contributes to improved DNA affinity of TRF2 in general. Additionally, we suggest how human RAP1 regulates TRF2 attraction and specificity to DNA and thus degree of telomere protection by shelterin.
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