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Identification of Novel Factors and Mechanisms regulating the Mitotic DNA Damage Response

De Marco Zompit, Mara. Identification of Novel Factors and Mechanisms regulating the Mitotic DNA Damage Response. 2022, University of Zurich, Faculty of Science.

Abstract

DNA double strand breaks (DSBs) are amongst the most toxic and dangerous lesions and can therefore severely threaten the integrity of our genome. The DNA damage response (DDR) pathway evolved to promptly signal and repair DSBs by initiating a series of chromatin remodeling events and recruiting many DNA repair factors to the site of lesion. Depending on the cell cycle phase, cells can take advantage of two main pathways for repair of the broken chromosome ends. The non-homologous end joining (NHEJ) is mainly active in G1 and early-S phases and represents an error-prone mechanism. On the contrary, the homologous recombination (HR) pathway is a more faithful process that requires the presence of the sister chromatids as template for accurate repair and thus, can only take place from S phase onwards. The DDR is fully active throughout interphase while it is partially inhibited during mitosis. DSBs arising during cell division generate acentric chromatin fragments that can lag during chromosome segregation as they lack any connection to the mitotic spindle. At the end of mitosis, the acentric fragments are often incorporated within micronuclei in one of the daughter cells. Micronuclei formation depicts a dangerous event for cycling cells as they are responsible for massive chromosomal rearrangements, including chromothripsis, which is often observed in tumor cells. Nevertheless, it is hypothesized that mitotic cells mark and stabilize DSBs via a tethering mechanism in favor of the completion of cell division and a more accurate repair during the next cell cycle.
In this thesis, I contributed to the discovery and characterization of a new genome maintenance complex consisting of the two proteins CIP2A and TOPBP1, which is implicated in the stabilization of DSBs specifically during mitosis. In particular, CIP2A and TOPBP1 are recruited downstream of MDC1 in response to DSBs. We discovered a constitutive interaction between MDC1 and TOPBP1 that is mediated by acidophilic casein kinase 2 (CK2). The latter phosphorylates newly characterized and highly conserved serine residues (S168 and S196) on MDC1 leading to its association with TOPBP1 via two of its nine BRCT domains. In contrast, we found that the CIP2A-TOPBP1 interaction is cell cycle regulated and is controlled by CRM1-dependent nuclear export of CIP2A during interphase. Hence, CIP2A can only associate with TOPBP1 upon breakdown of the nuclear envelope, which happens at the onset of mitosis. Within the context of mitotic chromatin, CIP2A is essential for the formation of TOPBP1 foci at sites of DNA damage.
Interestingly, the CIP2A-TOPBP1 mitotic complex not only marks DSBs but also recognizes and localizes to replication-associated DNA lesions, probably at under- replicated DNA, to which it is recruited in a MDC1-independent manner. Loss of CIP2A is lethal in cells deficient of the tumor suppressors BRCA1/2, which may offer a new therapeutic opportunity for the treatment of BRCA1/2 mutated breast and ovarian cancers. We showed that CIP2A-TOPBP1 mitotic interaction contributes to the survival and proliferation of BRCA1/2 mutated cancer cells, which are characterized by an increased load of replication-associated DNA lesions due to the lack of functional HR pathway. We hypothesize that the CIP2A-TOPBP1 interaction helps BRCA1/2 deficient cancer cells to cope with the high amount of DSBs during mitosis, thus evading cell death.
During cell division, CIP2A-TOPBP1 may tether broken acentric fragments, generated upon DSBs, to their cognate centric chromosome preventing micronucleation. Indeed, disruption of the CIP2A-TOPBP1 interaction is accompanied by an increased number of micronuclei and chromosomal aberrations, particularly in BRCA1/2 depleted cells. Thus, the identification of drugs targeting the CIP2A-TOPBP1 complex might be beneficial for the development of new treatments for BRCA1/2 mutated tumors, which urgently calls for an alternative therapeutic approach due to emerging resistance mechanisms to poly-(ADP)-ribose polymerase inhibitors (PARPi).

Additional indexing

Item Type:Dissertation (monographical)
Referees:Lopes Massimo, Stucki Manuel, Janscak Pavel
Communities & Collections:04 Faculty of Medicine > University Hospital Zurich > Clinic for Gynecology
UZH Dissertations
Dewey Decimal Classification:610 Medicine & health
Language:English
Date:May 2022
Deposited On:15 Feb 2023 08:35
Last Modified:15 Feb 2023 08:36
OA Status:Closed
Related URLs:https://uzb.swisscovery.slsp.ch/permalink/41SLSP_UZB/1d8t6qj/alma99117251223405508 (Library Catalogue)
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