Nuclear translocation of Gasdermin D (GSDMD) sensitizes chemotherapy through inhibiting PARP-1 in colorectal cancer

Abstract

Gasdermin D (GSDMD) has recently been identified to play a central role in pyroptosis of monocytes, through its N-terminal effector domain cleaved by activated inflammatory caspases to form pores on cell membrane. However, as the universally expressed protein in cancer cells, its function beyond pyroptosis, the pyroptotic function and pyroptosis-independent function of GSDMD in cancer cells have been poorly characterized. In my dissertation, I found GSDMD nuclear translocation and the apoptotic role of GSDMD for the first time. My first objective was to explore the correlation between the nuclear localization of GSDMD and overall survival rate of colorectal cancer patients. To address my first objective, I demonstrated upregulation of GSDMD mRNA in adenocarcinoma compared with GSDMD expression in adenoma and normal mucosal epithelia. Further studies using anti-GSDMD antibody in clinical specimens validated the upregulation of GSDMD protein in adenocarcinoma tissue. However, statistical analysis revealed no significant correlation of GSDMD expression levels with the overall survival rate of colorectal cancer patients. Very interestingly, I discovered the existence of GSDMD nuclear location in colorectal cancer cells of patients, and such nuclear translocation of GSDMD induces apoptosis and has significantly positive correlation with a better overall survival of patients with colorectal cancer. For the first time, I reported that non-pyroptotic GSDMD plays a beneficial role in promoting chemotherapy-induced apoptosis of tumor cells via its nuclear localization, and insufficient or absent GSDMD nuclear translocation predicts worse prognosis of colorectal cancer patients. My second objective was to define the non-pyroptotic function of GSDMD in colorectal cancer cells with chemotherapy and investigate how full-length GSDMD translocates into nucleus to induce apoptosis. My findings indicated that non-pyroptotic full-length GSDMD promotes chemotherapy-induced apoptosis of tumor cells via its nuclear translocation and subsequent regulation of poly (ADP-ribose) polymerase-1 (PARP-1) mediated DNA repair. After chemotherapeutic drug treatment induces full-length GSDMD nuclear translocation, GSDMD interacts with nuclear protein PARP-1 to dramatically inhibit its DNA Damage Repair-related function, to act similarly like the PARP inhibitor Olaparib. My findings confirmed that GSDMD related apoptosis is nuclear translocation dependent. My final objective was to dissect the potential clinical value of GSDMD nuclear translocation for chemosensitization by manipulating GSDMD expression and nuclear localization signal (NLS) and ataxia-telangiectasia mutated (ATM) signaling activation. My results indicated that NLS and ATM dependent mechanism of GSDMD nuclear translocation sensitize DNA damage related chemotherapeutic drug treatment. Together, these studies redefined the pyroptosis-independent function of GSDMD in its full-length form. My studies demonstrate that full-length GSDMD nuclear translocation facilitates chemotherapy-induced apoptosis through inhibiting PARP-1-mediated DNA Repair. Overall, our data strongly suggest that the combination of platinum-based chemotherapy and Olaparib in GSDMD low-expressed clinical cases, and subcellular localization of GSDMD may serve as a promising target for therapy of colorectal cancer and a good indicator for clinical outcomes of colorectal cancer patients. These studies are innovative in the context of GSDMD nuclear localization, cleavage-independent non-pyroptotic action, chemosensitization and better prognosis.