The role of mitochondrial/ER contacts in the regulation of mtDNA release from mitochondria, innate immune signaling, and responses to viral infection

PROJECT SUMMARY Innate immunity is critical for human health, allowing cells to detect and combat invasion by pathogens. Mitochondria are essential organelles that play important roles within the regulation of innate immune pathways. Contact sites between mitochondria and the endoplasmic reticulum (mitochondria/ER contacts, or MERCs) are important for mitochondrial homeostasis (such as replication of mitochondrial DNA, or mtDNA), but also act as signaling platforms for antiviral responses to viral dsRNA. However, the role of MERCs in the regulation of innate immune responses to cytoplasmic DNA is not well understood. In addition, mtDNA activates innate immune pathways when released from mitochondria into the cytoplasm. Dr. Laura Newman has found that MERCs stimulate the release of mtDNA in response to stalled mtDNA replication caused by mtDNA damage. Though it is well-established that cytoplasmic mtDNA enhances antiviral defenses, whether MERCs regulate mtDNA release during viral infection is unknown. Certain DNA viruses (HSV-1 and EBV) damage mtDNA directly or inhibit its replication, suggesting that removal of mtDNA (and its antiviral properties) may aid viral replication. This provides an ideal model system to test whether MERCs mediate release of damaged mtDNA during infection. In addition, RNA viruses disrupt the ER and MERCs to replicate. Release of mtDNA from mitochondria occurs during infection by several RNA viruses (such as influenza); therefore, MERCs may also mediate mtDNA release in response to RNA viral infection. The central hypothesis is that MERCs regulate mtDNA release and coordinate dsRNA and DNA innate immune responses to amplify cellular antiviral defenses. Aim #1 examines whether MERCs stimulate mtDNA release during HSV-1 or EBV infection, and whether mtDNA release into the cytosol benefits the host cell or virus. Aim #2 builds upon Dr. Newman’s preliminary data that the mitochondrial protein MFN1 enhances innate immune responses to cytoplasmic DNA, and tests whether MFN1 complexes with two innate immune adaptors that sense DNA (STING) and dsRNA (MAVS) at MERCs to regulate antiviral defenses. Lastly, Aim #3 examines whether RNA viruses (Influenza A and SARS-CoV-2) disrupt MERCs, causing stalled mtDNA replication and release, and whether this enhances antiviral defenses. Successful completion of any aim will provide important insights into the regulation of antiviral defenses, possibly informing new therapeutic targets to limit viral infection. This research will also provide virology training to the candidate, and research on viral-mitochondrial interactions will be carried over to her own lab. This award will enable Dr. Newman to take advantage of virology and immunology expertise via her advisory committee (Drs. O’Shea and Kaech), as well as additional career development opportunities at the Salk Institute. This will aid her transition to an independent scientist specializing in the role of mitochondria within innate immune pathways, which is a rapidly expanding and important area of scientific research.