Project Grant R01AI183430

UNDERSTANDING HIV-1 PERSISTENCE AND HIV-1-INDUCED INFLAMMATION IN TISSUES - PROJECT SUMMARY DESPITE EFFECTIVE ANTIRETROVIRAL THERAPY (ART), HIV RESIDES IN INFECTED CELLS AS INTEGRATED PROVIRUSES AND PERSISTS LIFELONG. WHILE ART SUPPRESSES PLASMA VIRAL LOAD TO CLINICALLY UNDETECTABLE LEVELS, ART DOES NOT KILL THE EXISTING HIV-INFECTED CELLS. HIV-INDUCED CHRONIC IMMUNE ACTIVATION ACCELERATES AGING, INCREASES RISKS OF PREMATURE CARDIOVASCULAR DISEASES, AND INCREASES CANCER RISKS. IT WAS BELIEVED THAT HIV LATENTLY INFECTED CELLS, AS OPPOSED TO TRANSCRIPTIONALLY ACTIVE HIV-INFECTED CELLS, ARE THE MAJOR BARRIER TO CURE. TRANSCRIPTIONALLY ACTIVE HIV-INFECTED CELLS SHOULD PRESUMABLY DIE OF VIRAL CYTOPATHIC EFFECTS OR IMMUNE CLEARANCE AND THEREFORE WERE NOT THE MAJOR FOCUS OF RESEARCH. HOWEVER, RECENT STUDIES REVEALED THAT TRANSCRIPTIONALLY ACTIVE HIV-INFECTED CELLS CAN SURVIVE VIRAL CYTOPATHIC EFFECT, ESCAPE IMMUNE CLEARANCE, PERSIST, AND PROLIFERATE DESPITE SUPPRESSIVE ART. THIS IS BECAUSE ALTHOUGH ART INHIBITS VIRAL ENZYME FUNCTION, ART DOES NOT INHIBIT HIV LTR PROMOTER ACTIVITY. DESPITE SUPPRESSIVE ART, HIV LTR PROMOTER FIRES BURSTS OF VIRAL PROTEIN EXPRESSION IN PEOPLE LIVING WITH HIV UNDER SUPPRESSIVE ART AND INDUCE IMMUNE ACTIVATION. OUR CENTRAL HYPOTHESIS IS THAT TRANSCRIPTIONALLY ACTIVE HIV- INFECTED CD4+ T CELLS DRIVE CHRONIC IMMUNE ACTIVATION IN TISSUES. OUR GOAL IS TO EXAMINE HIV-INDUCED IMMUNE ACTIVATION AT THE SINGLE-CELL LEVEL. IDENTIFYING HIV-INFECTED CELLS FOR MECHANISTIC UNDERSTANDING IS EXTREMELY CHALLENGING BECAUSE OF THE RARITY AND HETEROGENEITY OF HIV-INFECTED CELLS. FIRST, ONLY 1 IN A MILLION CD4+ T CELLS HARBOR INFECTIOUS HIV PROVIRUS, WHILE AROUND 1000 PER MILLION CD4+ T CELLS HARBOR DEFECTIVE HIV PROVIRUS. THE REMAINING 99.9% OF CD4+ T CELLS ARE UNINFECTED. THEREFORE, BULK TRANSCRIPTOMIC APPROACHES MEASURE THE 99.9% OF UNINFECTED CELLS AND DO NOT REFLECT THE CELLULAR STATES OF HIV-INFECTED CELLS. SECOND, CD4+ T CELLS ARE EXTREMELY HETEROGENEOUS BECAUSE OF DISTINCT T CELL DIFFERENTIATION (SUCH AS NAIVE, CENTRAL MEMORY, AND EFFECTOR MEMORY), POLARIZATION (SUCH AS TH1, TH1, TH17, TFH, TREG), IMMUNE PROGRAMS (SUCH AS ACTIVATION, EXHAUSTION, AND CYTOKINE RESPONSES). THEREFORE, SINGLE-CELL MULTI-OMIC PROFILING IS NEEDED TO IDENTIFY THE RARE AND HETEROGENEOUS HIV-INFECTED CELLS. THIRD, THERE IS NO CELLULAR MARKERS THAT CAN SPECIFICALLY DISTINGUISH HIV-INFECTED CELLS FROM UNINFECTED CELLS. THEREFORE, UNDERSTANDING HIV PERSISTENCE IN VIVO HAS BEEN A MAJOR BARRIER IN THE FIELD. TO RESOLVE THE RARITY AND HETEROGENEITY OF HIV RESERVOIR, OUR APPROACH IS TO USE SINGLE-CELL MULTIOMIC APPROACHES TO PROFILE THE RARE HIV-INFECTED CELLS AT THE SINGLE CELL LEVEL AND EXAMINE THE HETEROGENEOUS INFECTED CELLS. WE PIONEERED SINGLECELL MULTI-OMIC PROFILING TO ADVANCE OUR UNDERSTANDING OF HIV RESERVOIR. USING SINGLE-CELL ECCITESEQ, WE USED HIV RNA EXPRESSION AS A SURROGATE TO IDENTIFY TRANSCRIPTIONALLY ACTIVE HIV-INFECTED CELLS AND THEIR SINGLE- CELL TRANSCRIPTOME LANDSCAPE. USING SINGLE-CELL DOGMASEQ, WE IDENTIFIED LATENT (HIV DNA+ RNA-) AND TRANSCRIPTIONALLY ACTIVE (HIV RNA+) HIV-INFECTED CELLS AND THEIR EPIGENETIC REGULATORS, TRANSCRIPTIONAL LANDSCAPE, AND SURFACE PROTEIN EXPRESSION. IN AIM 1, WE WILL EXAMINE THE IMMUNE ACTIVATION PROGRAMS IN HIV-INFECTED CELLS. IN AIM 2, WE WILL EXAMINE WHETHER HIV INFECTION INDUCES IMMUNE ACTIVATION IN UNINFECTED IMMUNE CELLS.