Technische Universität Darmstadt
Anugwom (geb. Biehs), Ronja.
Resection-dependent canonical non-homologous end-joining induces genomic rearrangements.
Degree: PhD, 10 Department of BiologyRadiation Biology and DNA Repair, 2017, Technische Universität Darmstadt
DNA double-strand breaks (DSBs) are the main threat to genomic integrity. The majority of DSBs are repaired by canonical non-homologous end-joining (c-NHEJ), where the two DSB ends are rejoined with minimal processing. Some studies suggest that the rejoining of DSBs by c-NHEJ can be error-prone by causing sequence alterations and/or the misrejoining of two separate DSBs. Such genomic rearrangements are a driving force in carcinogenesis. However, it remains an ongoing discussion as to how such rearrangements arise, as other studies associate genomic rearrangements with alternative end-joining (alt-EJ) and factors favoring resection. During alt-EJ the involvement of resection results in the loss of genetic information. However, there is evidence that alt-EJ mechanisms only play a role in human cells, which are deficient for certain repair proteins. This is the case in combination with genetic defects or in tumor cells. Thus, it remains unclear how mutagenic end-joining, which results in genomic rearrangements, operates and is regulated in wild-type human cells.
To answer this question, mutagenic end-joining repair was investigated in this study by combining a reporter assay with other molecular biological assays. This reporter assay monitors the misrejoining of two 3.2 kilobase distant DSB ends under the loss of the intervening fragment. In addition, the sequence alterations at the misrejoined break sites were analyzed and overall repair was investigated. Furthermore, two approaches were taken to understand the circumstances under which error-prone end-joining is employed in wild-type human cells: the misrejoining of DSBs was compared between different reporter assays and the interactions of proteins involved in this pathway were analyzed.
The results of this study show that distant DSB ends are misrejoined by a hitherto undescribed slow repair mechanism, which is specific for the G1 phase of the cell cycle. These misrejoined break sites are frequently associated with sequence alterations, especially small deletions (less than 50 nucleotides). These deletions are the result of limited resection. Surprisingly, the DSB ends remain protected by the key c-NHEJ factor KU during this undescribed repair mechanism. This feature distinguishes this end-joining process from all previously described pathways involving resection. Indeed, the absence of factors such as KU or the anti-resection factor 53BP1 results in increased genomic rearrangements by alt-EJ. Hence, in this novel repair pathway, c-NHEJ factors are pivotal in ensuring resection remains limited.
Resection-dependent c-NHEJ is initiated by the activation of the resection factor CtIP. This is achieved through damage-inducible phosphorylation by the kinase PLK3 at Ser327, Thr847, and probably additional sites. Subsequently, the phosphorylation of CtIP at Ser327 results in its interaction with the pro-resection factor BRCA1. Similar to its role in S/G2 phase, the BRCA1-CtIP interaction seems to be important to overcome the resection barrier posed by 53BP1. Resection…
Advisors/Committee Members: Löbrich, Markus (advisor), Hamacher, Kay (advisor).
to Zotero / EndNote / Reference
APA (6th Edition):
Anugwom (geb. Biehs), R. (2017). Resection-dependent canonical non-homologous end-joining induces genomic rearrangements. (Doctoral Dissertation). Technische Universität Darmstadt. Retrieved from http://tuprints.ulb.tu-darmstadt.de/7044/
Chicago Manual of Style (16th Edition):
Anugwom (geb. Biehs), Ronja. “Resection-dependent canonical non-homologous end-joining induces genomic rearrangements.” 2017. Doctoral Dissertation, Technische Universität Darmstadt. Accessed April 21, 2018.
MLA Handbook (7th Edition):
Anugwom (geb. Biehs), Ronja. “Resection-dependent canonical non-homologous end-joining induces genomic rearrangements.” 2017. Web. 21 Apr 2018.
Anugwom (geb. Biehs) R. Resection-dependent canonical non-homologous end-joining induces genomic rearrangements. [Internet] [Doctoral dissertation]. Technische Universität Darmstadt; 2017. [cited 2018 Apr 21].
Available from: http://tuprints.ulb.tu-darmstadt.de/7044/.
Council of Science Editors:
Anugwom (geb. Biehs) R. Resection-dependent canonical non-homologous end-joining induces genomic rearrangements. [Doctoral Dissertation]. Technische Universität Darmstadt; 2017. Available from: http://tuprints.ulb.tu-darmstadt.de/7044/