ABSTRACT Immune activation is a hallmark of chronic HIV infection that is associated with increased risk of comorbidity and mortality. Prior studies of cannabinoids (CAN) show that CAN has immunomodulatory effects in the HIV- uninfected population. Given the elevated inflammatory state in chronic HIV-infected individuals and CAN’s possible anti-inflammatory effects, it has been suggested that CAN may reduce inflammation in HIV disease. However, studies of CAN’s ability to reduce inflammation in the setting of HIV-infection are contradictory and the underlying mechanism of CAN’s effects in the setting of HIV infection remains poorly understood. Our preliminary data in vivo in humans administered Δ-9 tetrahydrocannabinol (THC), the principal active constituent of CAN, demonstrate that the immunomodulatory effects of CAN differ by cell type. Thus, it is critical to understand the precise mechanisms of THC in HIV infection in specific cell types. In this application, we hypothesize that THC alters the immunogenome in a cell type-specific fashion and alters cytokine production via epigenetic regulatory mechanisms and that these alterations differ between HIV-infected and HIV-uninfected host genomes. To test these hypotheses, we propose defining the epigenomic and transcriptomic alterations at single cell resolution in peripheral blood mononuclear cells by administering THC to humans with and without HIV infection. Specifically, we will 1) identify the cell type-specific mRNA alterations on immune genes and serum cytokine alterations by acute THC in HIV-infected and HIV-uninfected individuals; 2) define the underlying epigenetic mechanisms responsible for acute THC-modulated gene expression; 3) identify epigenetic mechanisms regulating gene expression and serum cytokine levels for chronic cannabis use in HIV-infected men and women. Using a well-controlled THC challenge paradigm and rigorous study design (i.e. in vivo and in vitro, within and between subjects, functional profiling, included both sexes, evaluation of acute and chronic effects of the drug), we expect to identify cell-type specific genes whose expression is affected by THC and are associated with chromatin and DNA methylation modifications that lead to cytokine alteration as a functional outcome. The project will fill knowledge gaps in our understanding of THC’s effect on immune and inflammatory functions, whether in vivo changes are consistent with those observed in vitro, and whether these changes depend on HIV status. The in-depth understanding of gene regulatory mechanisms that may result in cell type and cytokine abnormalities will provide novel insights on whether CAN protects against inflammatory disease progression in HIV-infected individuals and inform treatment options.