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Novel Pathways Linking Mammalian Septins and the DNA Damage Response

Errington, Timothy
Format
Thesis/Dissertation; Online
Author
Errington, Timothy
Advisor
Macara, Ian
Abstract
A cellular response to damaged DNA, known as the DNA damage response (DDR), has evolved to combat damage that occurs from exposure to genotoxic agents or byproducts of normal cellular metabolism. Upon recognition of DNA damage, the cell arrests the cell cycle and repairs damaged DNA to maintain genome integrity. However, if the damage is severe, cells undergo apoptosis or initiate cellular senescence. The DDR is a highly coordinated event linking many pathways involved in various cellular processes. A previous study from this lab implicated mammalian septins in the DDR, although through an unknown mechanism. These cytoskeletal proteins function as signaling platforms and diffusion barriers and associate with various proteins including the adaptor proteins SOCS7 and NCK. In response to multiple types of DNA damage, NCK relocalizes from the cytoplasm to the nucleus, using the nucleocytoplasmic shuttling protein SOCS7. The nuclear accumulation of NCK in response to UV irradiation is dependent on the kinase activity of ATR, a member of the PIKK family that is activated early in the DDR. Depletion of NCK results in elevated phosphorylation of the transcription factor p53 and an early induction of apoptosis. Depletion of SOCS7, which blocks the nuclear accumulation of NCK, also increases phosphorylation of p53 and also results in an early induction of apoptosis. This indicates the anti-apoptotic role of NCK is dependent on its nuclear translocation during the DDR. Another septin interacting protein was identified using a proteomic approach. This novel nuclease called Septin Associated Nuclease 1 (SAN1) possesses unique 5’ exonuclease activity mediated by a FEN1-related nuclease domain necessary for the proper repair of DNA interstrand crosslinks (ICL). Depletion of SAN1 results in a low rate of homologous recombination (HR) due to a decrease in end-resection of double-strand breaks generated during ICL repair. Additionally, SAN1 is mostly localized in the cytoplasm but accumulates in the nucleus following treatment with ICL-inducing agents. However, in the absence of septins, SAN1 becomes mislocalized and distributed throughout the cell. Depletion of septins also results in a decrease in HR and end-resection. This suggests septins are necessary for the proper response to ICL by regulating SAN1 localization and activation. Taken together, these data demonstrate that mammalian septins play a role in the DDR and highlight an unexpected link between cytoskeletal elements and DNA damage signaling.
Language
English
Published
University of Virginia, Department of Microbiology, Immunology, and Cancer Biology, PHD (Doctor of Philosophy), 2013
Published Date
2013-11-26
Degree
PHD (Doctor of Philosophy)
Collection
Libra ETD Repository
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