Post-incision events induced by UV : regulation of incision and the role of post-incision factors in mammalian NER
Overmeer, R.M.
Citation
Overmeer, R. M. (2010, September 29). Post-incision events induced by UV : regulation of incision and the role of post-incision factors in mammalian NER.
Retrieved from https://hdl.handle.net/1887/15997
Version: Corrected Publisher’s Version
License: Licence agreement concerning inclusion of doctoral thesis in the Institutional Repository of the University of Leiden Downloaded from: https://hdl.handle.net/1887/15997
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Post-incision events induced by UV
Regulation of incision and the role of post-incision factors in mammalian NER
René Marcel Overmeer
Post-incision events induced by UV
Regulation of incision and the role of post-incision factors in mammalian NER
proefschrift
ter verkrijging van
de graad van Doctor aan de Universiteit Leiden,
op gezag van de Rector Magnifi cus prof. mr. P.F. van der Heijden, volgens het besluit van het College voor Promoties
te verdedigen op woensdag 29 september 2010 klokke 16.15 uur
door
René Marcel Overmeer geboren te Alphen aan den Rijn
op 23 juni 1980
Promotiecommissie:
Promotor: Prof. dr. L.H.F. Mullenders Copromotor: Dr. M.I. Fousteri
Overige leden: Prof. dr. J. Brouwer
Prof. dr. R. van Driel (UvA) Dr. W. Vermeulen (Erasmus MC)
ISBN 978-90-6464-417-7
Printed by GVO drukkers & vormgevers B.V. | Ponsen & Looijen
The printing of this thesis was fi nancially supported by the J.E. Jurriaanse Stichting and Greiner Bio-One.
Contents
Aim and outline of the thesis 7
Chapter 1: Introduction 11
Chapter 2: GG-NER complex assembly and regulation 21
Chapter 3: Perspectives 39
Chapter 4: Replication Protein A safeguards genome integrity 45 by controlling NER incision events
Chapter 5: RFC recruits DNA polymerase δ to sites of NER 69 but is not required for PCNA recruitment
Chapter 6: Three DNA polymerases, recruited by different 91 mechanisms, carry out NER repair synthesis in
human cells
Chapter 7: UV induces DNA damage signaling in non-cycling 117 human cells independent of nucleotide excision repair
Summary for the layman 141
Samenvatting voor de leek 145
Addendum 149
Curriculum Vitae 150
List of publications 151
Dankwoord 152
Aim and outline of the thesis
8
Aim and outline of the thesis
Cancer has been projected to become the most prevalent cause of death worldwide in 2010.
This is mostly due to a decreased risk of dying from cardiovascular diseases, made possible by improved understanding and treatment, in which research has had a major part. Likewise cancer treatment has greatly improved over the last century, although the main pillars behind the treatment have remained the same. Initially cancer was treated by a combination of surge- ry and DNA damage infl iction by ionizing radiation, since then many novel DNA damaging agents, also know as cytostatics, have been introduced.
It is now known that cancer is caused by an accumulation of mutations that lead to un- checked cell growth and proliferation. So how does a cell accumulate these mutations? First of all mutations are formed when cells attempt to replicate damaged DNA as the damage disrupts the base stacking and/or the backbone integrity. The sensitivity of cancer to DNA damaging agents led to the discovery that cancers are often less capable of repairing DNA damage. Moreover normal cells possess various DNA damage repair pathways which safe- guard the integrity of the genome and prevent DNA damage to induce mutations, whereas tumour cells are often defi cient in one or more of these repair pathways. Defects in repair lead to an accumulation of DNA damage, which in turn leads to a higher mutation frequency.
In addition cancer cells might replicate damaged DNA without proper functioning cell-cycle checkpoints. A consequence of such defects in cell-cycle checkpoints is that cells have less time to repair DNA damage before DNA replication and importantly might also escape from apoptosis. The combination of repair and checkpoint defects makes cancer cells highly mu- tagenic.
To summarize, the origin of cancer is therefore thought to lie in initial mutations in ei- ther DNA repair or cell-cycle checkpoint factors induced by DNA damage as well as factors that control apoptosis. In concurrence with these observations, DNA damaging agents in our environment such as components of tobacco smoke, fuel combustion, ultraviolet light (UV) from the sun and ionizing radiation have been found to lead to an increased cancer risk. However by-products from cellular metabolism can also damage DNA, indicating that complete avoidance of DNA damage is not feasible. Therefore, DNA repair and inhibition of cell-cycle progression (either to facilitate repair or to trigger controlled cell death) are critical in preventing mutations and subsequently cancer.
The fi rst chapter of this thesis gives a short overview of the major DNA damaging agents and the opposing repair pathways, subsequently nucleotide excision repair (NER), able to repair a wide range of damages, will be discussed. The second chapter will then thoroughly discuss the assembly of the NER complex, the subsequent incision and repair synthesis.
The recently elucidated regulation of NER will also be discussed in detail. In addition DNA damage induced signalling will be discussed briefl y. Perspectives in respect to NER related research will be discussed in the third chapter. The fourth chapter concerns the regulation of NER, preventing dual incision when gaps, formed by previously excised damages, cannot be
9 fi lled. Chapter fi ve and six concern the recruitment and activity of replication factors after dual incision. Finally, chapter seven describes UV damage mediated incision and signalling in NER defi cient cells.
The main focus of this thesis is the sequence of events following dual incision, as much was known about the steps and requirements leading to incision yet little was known about the handover from pre- to post-incision complexes, the recruitment of post-incision factors and how they function in NER.
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