Defining roles and mechanisms of class II MHC antigen presentation in human endothelial cells across barrier tissues - Project Summary/Abstract Non-immune cells in external antigen-exposed tissues participate in immune responses through antigen presentation. In such barrier tissues, non-immune, structural cells, like endothelial cells (ECs), directly engage T cells, which then proliferate and differentiate into distinct functional subsets that can influence vascular diseases. To alleviate such pathologies, it is important to understand how ECs, which differ structurally and transcriptionally from professional antigen presenting cells, are capable of exogenous antigen presentation on class II major histocompatibility complex MHC II. Though the mechanism and is unclear, it is known that the inflammatory cytokine, interferon- (IFN-) induces extensive transcriptional changes and promotes MHC II presentation in ECs. So, my central hypothesis is that genes upregulated by cytokine interferon- (IFN-) enable MHC II antigen presentation and direct tissue-specific interactions with T cells. I aim to elucidate the molecular mechanisms of antigen presentation by ECs and profile the subsequent local, tissue-specific adaptive immune responses, ultimately identifying therapeutic targets for vascular pathologies. In this proposal, I will profile antigen-experienced memory CD4+ T cell responses to IFN--activated ECs from the barrier tissues of human skin, lung, and intestine presenting foreign antigen on MHC II, and identify the IFN- -stimulated genes (ISGs) required for ECs to express MHC II. In Aim 1, I will characterize the tissue-specific, IFN--driven CD4+ T cell responses to surface MHC II expression on ECs from barrier (skin, lung, and intestine) and non-barrier (spleen) tissues. To profile CD4+ T cell responses, I will co-culture the ECs with CD4+ T cells in an in vitro model of transplantation and assess the T cell subset differentiation and proliferation to distinguish the resulting physiological immune responses. In Aim 2, I will conduct a forward genetics screen to determine the ISGs required for MHC II presentation in ECs. I will introduce my lab’s unique library of ISGs into skin ECs, separate the cells by their antigen presenting phenotype, and sequence the barcoded genes to identify which deleted ISGs were essential. Finally, I will investigate the mechanism of each identified ISG. Together, this novel investigation will clarify the heterogenous T cell responses to semi-professional antigen presentation and reveal the cellular and molecular mechanisms that underlie them. My findings will identify targets that can be used to develop therapies that mediate transplant rejection and other vascular diseases. My long-term goal is to be an independent investigator in vascular immunology, leveraging the immunomodulatory potential of non-immune cells to regulate immune and vascular pathologies. With the guidance of my mentors, my research training plan will advance my technical skills as a researcher and develop the interpersonal skills in teaching and scientific communication required for my professional future.
