In BriefHIV evolved a minimalist but robust transcriptional circuit bypassing host regulatory checkpoints; however, the fragility of the circuit in the host phase (which primes HIV for activation) largely affects proviral transcription and fate. Highlights• The host and viral phases of the HIV transcriptional circuit have different functional requirements • HIV evolved a minimalist program to robustly bypass host cell regulatory checkpoints • A mathematical model reveals that the host phase is subject to transcriptional circuit fragility • Host transcriptional circuit fragility influences the viral feedback and latency reversal potential

Immune cell state alterations rewire HIV-1 gene expression, thereby influencing viral latency and reactivation, but the mechanisms are still unfolding. Here, using a screen approach on CD4+ T cell models of HIV-1 latency, we revealed Small Molecule Reactivators (SMOREs) with unique chemistries altering the CD4+ T cell state and consequently promoting latent HIV-1 transcription and reactivation through an unprecedented mechanism of action. SMOREs triggered rapid oxidative stress and activated a redox-responsive program composed of cell-signaling kinases (MEK-ERK axis) and atypical transcription factor (AP-1 and HIF-1α) cooperativity. SMOREs induced an unusual AP-1 phosphorylation signature to promote AP-1/HIF-1α binding to the latent HIV-1 proviral genome for its activation. Consistently, latent HIV-1 reactivation was compromised with pharmacologic inhibition of oxidative stress sensing or of cell-signaling kinases, and transcription factor’s loss of expression, thus functionally linking the host redox-responsive program to viral transcriptional rewiring. Notably, SMOREs induced the redox program in primary CD4+ T cells and reactivated latent HIV-1 in aviremic patient samples alone and in combination with known latency-reversing agents, thus providing physiological relevance. Our findings suggest that manipulation of redox-sensitive pathways could be exploited to alter the course of HIV-1 latency, thus rendering host cells responsive to help achieve a sterilizing cure.

Immune stimulation fuels cell signaling transcriptional programs inducing biological responses to eliminate virus infected cells. Yet, retroviruses that integrate into host cell chromatin, such as HIV1, coopt these programs to switch between latent and reactivated states; however, the regulatory mechanisms are still unfolding. Here, we implemented a functional screen leveraging HIV1 dependence on CD4+ T cell signaling transcriptional programs and discovered ADAP1 is an undescribed modulator of HIV1 proviral fate. Specifically, we report ADAP1 (ArfGAP with dual PH domain containing protein 1), a previously thought neuronal restricted factor, is an amplifier of select T cell signaling programs. Using complementary biochemical and cellular assays, we demonstrate ADAP1 inducibly interacts with the immune signalosome to directly stimulate KRAS GTPase activity thereby augmenting T cell signaling through targeted activation of the ERK/AP1 axis. Single cell transcriptomics analysis revealed loss of ADAP1 function blunts gene programs upon T cell stimulation consequently dampening latent HIV1 reactivation. Our combined experimental approach defines ADAP1 as an unexpected tuner of T cell programs coopted by HIV1 for latency escape.