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Single-molecule analysis of subtelomeres and telomeres in Alternative Lengthening of Telomeres (ALT) cells
Journal article   Open access   Peer reviewed

Single-molecule analysis of subtelomeres and telomeres in Alternative Lengthening of Telomeres (ALT) cells

Heba Z. Abid, Jennifer McCaffrey, Kaitlin Raseley, Eleanor Young, Katy Lassahn, Dharma Varapula, Harold Riethman and Ming Xiao
BMC genomics, v 21(1), pp 485-485
15 Jul 2020
DOI: https://doi.org/10.1186/s12864-020-06901-7
PMID: 32669102
Featured in Collection :   UN Sustainable Development Goals @ Drexel
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https://doi.org/10.1186/s12864-020-06901-7View
Published, Version of Record (VoR)CC BY V4.0,  Open

Abstract

Biotechnology & Applied Microbiology Genetics & Heredity Life Sciences & Biomedicine Science & Technology
Background Telomeric DNA is typically comprised of G-rich tandem repeat motifs and maintained by telomerase (Greider CW, Blackburn EH; Cell 51:887-898; 1987). In eukaryotes lacking telomerase, a variety of DNA repair and DNA recombination based pathways for telomere maintenance have evolved in organisms normally dependent upon telomerase for telomere elongation (Webb CJ, Wu Y, Zakian VA; Cold Spring Harb Perspect Biol 5:a012666; 2013); collectively called Alternative Lengthening of Telomeres (ALT) pathways. By measuring (TTAGGG) n tract lengths from the same large DNA molecules that were optically mapped, we simultaneously analyzed telomere length dynamics and subtelomere-linked structural changes at a large number of specific subtelomeric loci in the ALT-positive cell lines U2OS, SK-MEL-2 and Saos-2. Results Our results revealed loci-specific ALT telomere features. For example, while each subtelomere included examples of single molecules with terminal (TTAGGG) n tracts as well as examples of recombinant telomeric single molecules, the ratio of these molecules was subtelomere-specific, ranging from 33:1 (19p) to 1:25 (19q) in U2OS. The Saos-2 cell line shows a similar percentage of recombinant telomeres. The frequency of recombinant subtelomeres of SK-MEL-2 (11%) is about half that of U2OS and Saos-2 (24 and 19% respectively). Terminal (TTAGGG) n tract lengths and heterogeneity levels, the frequencies of telomere signal-free ends, and the frequency and size of retained internal telomere-like sequences (ITSs) at recombinant telomere fusion junctions all varied according to the specific subtelomere involved in a particular cell line. Very large linear extrachromosomal telomere repeat (ECTR) DNA molecules were found in all three cell lines; these are in principle capable of templating synthesis of new long telomere tracts via break-induced repair (BIR) long-tract DNA synthesis mechanisms and contributing to the very long telomere tract length and heterogeneity characteristic of ALT cells. Many of longest telomere tracts (both end-telomeres and linear ECTRs) displayed punctate CRISPR/Cas9-dependent (TTAGGG) n labeling patterns indicative of interspersion of stretches of non-canonical telomere repeats. Conclusion Identifying individual subtelomeres and characterizing linked telomere (TTAGGG) n tract lengths and structural changes using our new single-molecule methodologies reveals the structural consequences of telomere damage, repair and recombination mechanisms in human ALT cells in unprecedented molecular detail and significant differences in different ALT-positive cell lines.

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Collaboration types
Domestic collaboration
Web of Science research areas
Biotechnology & Applied Microbiology
Genetics & Heredity
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