Hepatitis C viral infection leads to self DNA recognition and AIM2 activation in hepatocytes

Maynard, Arron (2023) Hepatitis C viral infection leads to self DNA recognition and AIM2 activation in hepatocytes. Masters thesis, Memorial University of Newfoundland.

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Lay Summary This thesis focuses on the cross-talk between hepatitis C (HCV) RNA viral infection and cytosolic self-DNA recognition. Cytosolic DNA is a damage associated molecular pattern that, when recognized, triggers a cascade involving DDX41 translocation and degradation, STING translocation and AIM2 inflammasome formation, ultimately leading to death of the cell. During HCV infection, and in the context of inflammation, most researchers focus on the well-studied NLRP3 inflammasome, but we believe HCV infection triggers the release of nuclear or mitochondrial DNA to the cytosol and activates the AIM2 inflammasome. This is reflected through an increase in AIM2 proteins, the visualization of macro-molecular aggregate structures tagged with anti-AIM2 antibodies and the colocalization of AIM2 and caspase-1 after HCV RNA infection. Abstract HCV is the leading cause of liver disease, cirrhosis, and hepatocellular carcinoma (HCC) in many countries. HCV, an RNA virus, may engage elements of cytosolic DNA response. Life or death of individual cells can determine disease progression in multicellular organisms, thus pyroptosis, a pro-inflammatory form of cell death, is initiated through the recognition of danger signals associated with pathological stimuli such as HCV. AIM2 inflammasome formation, triggered through the presence of cytosolic DNA, is not well characterized in the context of HCV, especially as it relates to hepatocytes in vitro. This is because there is dogma that RNA viral infection leads to NLRP3 inflammasome activation; whereas, DNA viral infection leads to AIM2 inflammasome activation, although there are some well studied exceptions. During HCV infection, AIM2 inflammasomes could potentially be activated through the loss of nuclear envelope or mitochondrial membrane integrity leading to the release of DNA to the cytosol. The proteins observed in these experiments were specifically chosen in the context of gaining a greater understanding of how JFH-1T infection in Huh-7.5 cells involves “cross talk” between RNA viral infection and self cytosolic DNA recognition. Here I demonstrated that after a three-day infection, there is a drastic decrease in the amount of DDX41 protein in the nucleus, suggesting DDX41 degradation. I also show that after infection, STING proteins were far less wide spread, and condensed into smaller pockets located just outside the nucleus, pointing to the possibility that STING proteins are being activated by DDX41 proteins. Since HCV is an RNA virus, and DDX41 is triggered by cytosolic DNA, this lays the foundation for understanding the potential for the formation of AIM2 inflammasomes within the same parameters. Next, I found that AIM2 protein colocalizes with caspase-1 after JFH-1T infection. I believe that the discovery of a new inflammasome activated by HCV may provide insight on programmed cell death and inflammation, which can have an impact on HCV pathogenesis and treatment, especially since AIM2 expression in down regulated in HCV-related cancers.

Item Type: Thesis (Masters)
URI: http://research.library.mun.ca/id/eprint/15782
Item ID: 15782
Additional Information: Includes bibliographical references (pages 97-119)
Keywords: HCV, pyroptosis, AIM2, Huh-7.5, PCD
Department(s): Medicine, Faculty of
Date: February 2023
Date Type: Submission
Digital Object Identifier (DOI): https://doi.org/10.48336/KZ70-P442
Library of Congress Subject Heading: Hepatitis C--Genetic aspects; Infection--Genetic aspects
Medical Subject Heading: Hepatitis C--genetics; RNA Viruses--genetics; Viral Diseases--genetics; Inflammasomes;

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