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HIRA protects ALT telomeres from aberrant R-loop accumulation

Lynskey, Michelle, Lee (2024) HIRA protects ALT telomeres from aberrant R-loop accumulation. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

The Alternative Lengthening of Telomeres (ALT) pathway is a homology-directed repair (HDR) mechanism of telomere extension that enables the proliferation of several aggressive cancers. Recurrent mutations inactivate the ATRX (alpha-thalassemia/mental retardation, X-linked)–DAXX (death domain associated protein) chromatin remodeling complex, which coordinates histone H3.3 deposition at telomeres. Recently, we showed that HIRA, the remaining H3.3 chaperone, adapts to compensate for ATRX loss by maintaining telomeric chromatin to sustain ALT cancer cell survival. However, the specific mechanisms underlying HIRA's ability to rescue ALT telomeres from the repercussions of ATRX loss remain unclear.
Here, we demonstrate that the HIRA-UBN1/2 complex mediates new H3.3 deposition, limiting the accumulation of unchromatinized, single-stranded DNA (ssDNA) presence at telomeres following ALT-associated HDR. HIRA adapts to maintain H3.3 levels but cannot entirely prevent R-loops, leading to collisions between RNA polymerase II and the ALT replisome and unwarranted replication stress. Furthermore, we show that Chk1-mediated H3.3 S31 phosphorylation is crucial for suppressing aberrant ssDNA accumulation and maintaining the integrity of telomeric chromatin. These studies suggest that the deposition of new histone H3.3 maintains productive ALT, and HIRA ensures ALT cell survival by limiting TERRA R-loop formation. This implicates HIRA in TERRA R-loop homeostasis, which we propose is essential in ensuring ALT cancer cell survival following ATRX loss.

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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Lynskey, Michelle, Leemichellellynskey@gmail.comMLL820000-0002-0343-3413
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairBakkenist, Christophercjb38@pitt.edu
Committee MemberBrodsky, Jeffreyjbrodsky@pitt.edu
Committee MemberLee, Adrianleeav@upmc.edu
Committee MemberSchopfer, FranciscoSchopfer, Francisco Jose <fjschopfer@katz.pitt.edu>
Thesis AdvisorO'Sullivan, Roderickosullivanr@upmc.edu
Date: 27 September 2024
Date Type: Publication
Defense Date: 19 April 2024
Approval Date: 27 September 2024
Submission Date: 30 April 2024
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 159
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Pharmacology and Chemical Biology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: Cancer, chromatin, telomere, HIRA, ALT, histone, R-loop, DNA damage
Date Deposited: 27 Sep 2024 15:33
Last Modified: 27 Sep 2024 15:33
URI: http://d-scholarship.pitt.edu/id/eprint/46362

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