Protein kinase C-delta regulates chromatin remodeling and DNA repair

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PROJECT ABSTRACT Head and neck cancer (HNC) accounts for 4% of all cancers in the US, and HNC patients are mostly treated with irradiation (IR) therapy or combination of treatments. Unfortunately, besides killing cancer cells, IR treatment also kills non-tumor cells in the oral cavity and damages the salivary glands, causing side effects such as chronic oral infection, decreased saliva production, and xerostomia. IR-induced damage to the salivary glands is thought to occur in part due from loss of salivary acinar cells through apoptosis. Our lab has identified protein kinase C delta (PKCδ) as a key regulator of IR-induced apoptosis in the salivary acinar cells in vitro and in the salivary gland in vivo. We have shown that upon IR damage PKCδ is activated and imported into the nucleus. Although we know that nuclear import of PKCδ is necessary for apoptosis, the mechanism of how nuclear PKCδ regulates DNA damage-induced apoptosis is unknown. Previous studies from our lab have suggested that PKCδ does not activate the apoptotic pathway directly but may instead regulate upstream pathways that contribute to the DNA damage response and DNA repair. My preliminary studies suggest a novel mechanism of PKCδ-mediated regulation of apoptosis through chromatin remodeling and DNA repair. My preliminary data shows that depletion of PKCδ increases IR-induced DNA damage repair, and that PKCδ regulates DNA repair through both non- homologous end joining (NHEJ) and homologous recombination (HR) pathways. Furthermore, my preliminary data suggests that PKCδ may regulate chromatin structure through altering histone modifications. Thus, my overall goal is to understand mechanistically how nuclear PKCδ regulates chromatin accessibility and DNA repair in response to IR damage. In Aim 1 I will characterize the role of nuclear PKCδ in NHEJ and HR pathways and analyze whether PKCδ is biased toward NHEJ or HR using in vivo fluorescent reporter assay. I will also determine if PKCδ affects the formation of NHEJ and HR repair complexes at sites of double-stranded breaks (DSBs) and perform rapid immunoprecipitation mass spectrometry to identify chromatin interacting proteins influenced by PKCδ. In Aim 2 I will ask how PKCδ regulates histone modifications and chromatin structure, and how this relates to DNA repair regulation. I will explore the role of PKCδ in histone modifications before and after IR, and the nuclear function of PKCδ in chromatin remodeling. To further understand the mechanism of this regulation, we will analyze if PKCδ regulates upstream histone-modifying enzymes. Overall, my proposed studies should enhance our understanding of the mechanism by which PKCδ regulates DNA damage-induced apoptosis, and may lead to new safe therapeutic interventions to protect salivary gland and oral tissues of patients undergoing IR for HNC. This F32 fellowship will also provide me with specific scientific and research training, including the development of new research skills, and exposure to basic and translational cancer and oral biology. Together with a wide variety of professional development opportunities, this training will prepare me for my long-term goal to become an independent cancer researcher.