Date of Award
8-4-2023
Degree Name
Doctor of Philosophy (PhD)
Degree Type
Dissertation
Abstract
DNA replication is a fundamental process and defects in replication can cause genetic mutations and instability and various human diseases including cancer. The progression of replication forks can be challenged by both intrinsic and environmental factors, such as DNA lesions, secondary structure, insufficient nucleotides, collision with the transcription apparatus, radiation, or cancer drugs. These challenges necessitate robust mechanisms to protect the fork structure in the event of replication stress. Tumor suppressors like BRCA1, BRCA2, and FANC can protect fork structure by controlling nucleases that can attack stalled replication forks, which otherwise can cause fork collapse and DNA damage. In addition to these factors, cells also activate signaling pathways for fork protection. One such pathway is the Ataxia Telangiectasia and Rad3- related/checkpoint kinase1 (ATR/Chk1)-dependent replication checkpoint. We have recently discovered a novel Ca2+-dependent signaling pathway that operates separately from the ATR/Chk1 checkpoint, although both pathways target the resection nuclease Exo1. In this Ca2+- dependent pathway, replication stress leads to the generation of DNA fragments, which upon cytoplasmic translocation, activates the sensor protein cGAS, leading to the synthesis of cGAMP from ATP and GTP. Serving as a second messenger, cGAMP binds to STING, causing its dissociation from the ion channel TRPV2 on the ER, leading to TRPV2 derepression and Ca2+ release. The resulting elevation of intracellular Ca2+ ([Ca2+]i) then activates CaMKK2 and the downstream kinase AMPK. Upon activation, AMPK phosphorylates Exo1 at S746, causing the binding of 14-3-3 proteins, which in turn prevents Exo1 fork recruitment. As a result, abnormal fork resection is avoided. Disruption of this pathway causes excessive fork processing, chromosomal instability, and reduced cell viability in the presence of replication stress. In our effort to further decipher this pathway, we identified TRP channel-associated factor 1 (TCAF1) as a novel fork protection factor that promotes TRPV2-mediated Ca2+ release from the ER after replication stress. TCAF1 does so by promoting the dissociation of STING from TRPV2. Our findings have revealed novel mechanisms of genome maintenance and have important implications for the understanding and treatment of cancer.
Language
English (en)
Chair and Committee
Zhongsheng You
Recommended Citation
Kong, Lingzhen, "Ca2+ Dynamics and Genome Protection" (2023). Arts & Sciences Electronic Theses and Dissertations. 3121.
https://openscholarship.wustl.edu/art_sci_etds/3121