Defining critical roles for ZBP1 in PANoptosis utilizing a novel genetic tool for disease modeling and therapeutic development

Innate immunity and innate immune cell death provide a critical first line of defense against disease. However, excess cell death leads to pathological inflammation. ZBP1 is an innate immune sensor that is central to this balance between defense and inflammation as a driver of inflammatory lytic cell death, PANoptosis. Activation of ZBP1-dependent PANoptosis downstream of diverse triggers has roles in both host defense and disease pathology, making ZBP1 an attractive therapeutic target. Therefore, understanding the distinct roles of ZBP1 in different cell types, organ systems, and tissues is critical to identify therapeutic strategies. Although ZBP1 regulates PANoptosis in multiple cell types, there are limited tools to interrogate its function in a cell type-specific manner. Here, we report the generation of a Zbp1-floxed mouse line (Zbp1fl/fl) for investigation of ZBP1 in distinct cell populations. We crossed Zbp1fl/fl mice to LysMcre mice to selectively deplete Zbp1 from the myeloid compartment, which did not alter immune homeostasis. Bone marrow-derived macrophages (BMDMs) from Zbp1fl/fl mice had normal ZBP1 expression and PANoptosis activation, while those from Zbp1fl/flLysMcre mice exhibited markedly reduced ZBP1 expression and were biochemically and functionally protected from ZBP1-driven PANoptosis; these effects were validated using known triggers of the ZBP1-PANoptosome--IAV, nuclear export inhibition plus IFN, and ethanol. These findings demonstrate this new Zbp1fl/fl mouse as a versatile tool that can be utilized with a variety of Cre-drivers to study ZBP1 in a wide array of distinct cell types. Given the critical role of ZBP1 in disease, this tool will inform the development of therapeutic strategies.