Function of Wdfy4 in cross-presentation and immunity

Project Summary This application will investigate the molecular and cellular mechanisms underlying antigen cross-presentation by dendritic cells (DCs), with a major focus on type 1 classical DCs (cDC1) which play a major role in priming CD8 T cell responses that are critical for cancer immunotherapy. Cross-presentation involves the presentation of exogenous antigens by MHC-I molecules, which is predominantly facilitated by cDC1 in vivo. This application will address critical gaps in understanding the cross-presentation process and its implications for immune responses, leveraging several novel in vivo models that we have produced to elucidate the specific roles and mechanisms of cDC1 and cDC2 in antigen processing and presentation. A central facet of our proposal is the investigation of WDFY4, which we identified as critically important for cross-presentation by cDC1. Our preliminary data reveal a pivotal role in mediating CD8 T cell responses against viruses and tumors, and implicating it in autoimmunity contexts such as type I diabetes. The proposal aims to define the molecular basis of cross-presentation, challenging alternative models by providing rigorous data supporting vacuolar pathways for antigen processing that involve WDFY4. Specifically, our proposal challenges the models that rely on a cytosolic route, by providing both an explanation for the apparent dependence on the TAP transporter and evidence for TAP-independent cross-presentation. Aims of this proposal are twofold: firstly, we will rigorously test the hypothesis that WDFY4-dependent cross-presentation in cDC1 employs a vacuolar pathway, contrasting it with the cytosolic route and assessing the role of the TAP transporter in this process. Secondly, we will define the molecular interactions of WDFY4 with proteins that mediate its actions and determine whether they are involved in regulating vesicular trafficking, cargo sorting, or cytoskeletal dynamics, which are crucial for the cross-presentation of antigens. This will involve identifying and validating interacting proteins, determining their structural relationships with WDFY4, and elucidating their functional relevance to cross-presentation in vitro and in vivo. By challenging existing paradigms and employing cutting-edge genetic and molecular tools, this research promises to significantly advance our understanding of dendritic cell biology, particularly the cellular and molecular mechanisms of antigen processing and presentation. The outcomes are expected to enhance our understanding of immune activation and to have broad implications for the design of more effective cancer immunotherapies and interventions for autoimmune diseases.