Follicular helper T cells (Tfh) are believed to be major contributors to the viral reservoir that persists in
HIV-infected individuals, even when on antiretroviral therapy. We will explore the novel hypothesis that cytokines
IL-10 and TGF-b, which play important roles in lymph node biology and are upregulated post HIV infection, are
critically important for the formation of such a reservoir and represent potential targets for intervention.
Specifically, we hypothesize that IL-10/TGF-b promote the differentiation of HIV-susceptible Tfh cells, with
reduced antiviral defenses, that favor integration of intact proviruses into open chromatin gene targets
downstream of IL-10/TGF-b signaling pathways. We will examine multiple aspects of this model, including how
IL-10 and TGF-b suppress the intrinsic antiviral machinery in Tfh cells and how chromatin accessibility in genes
downstream of IL10/TGF-b signaling, promoted by STAT3/SMADs activation, allows infecting virions to integrate
preferentially in these open loci. As a result of these mechanisms, infected Tfh cells provide a reservoir of actively
transcribing intact proviruses, even under ART.
In Aim 1, we will investigate these hypothesized roles of IL-10 and TGF-b in Tfh differentiation and HIV
integration/transcription using biobanked samples from lymph nodes of HIV-infected individuals at different
stages post HIV infection (i.e. Fiebig IV/V, untreated chronic infection, and ART-treated). In Aim 2, we will
mechanistically validate our model, using tools such as CRISPR gene knockouts in ex vivo tonsil isolated CD4
T cells, to examine how the major pathways identified in Aim 1 contribute to the differentiation of Tfh cells, the
suppression of anti-viral machinery, and the promotion of integration of intact proviruses into active transcription
sites. Finally, in Aim 3, we will take advantage of lymph node specimens available from a cohort of rhesus
macaques that previously controlled SIV replication post-ART interruption (VL<1000 cps/mL), subsequent to in
vivo blockade of the IL-10 and PD-1 pathways. We will dissect mechanistically how this treatment led to SIV viral
DNA decay in their LNs.
This proposal has several innovative aspects. Additionally to the available biobanked human and
macaque samples, we will use cutting-edge methods, including multiome, MIP-seq, multiplexed confocal imaging
with vDNA/RNA scope, and spatial transcriptomics. The project builds on our expertise with HIV pathogenesis,
Tfh biology, and the modulation of immune pathways through both in vitro models and in vivo interventions in
rhesus macaques. Additionally, we have the support of strong Co-Investigators with expertise in virology
(Vandekerckhove), tissue imaging (Petrovas), and data analysis, integration and interpretation (Kamaleswaran
and Sekaly). With such multidisciplinary approaches, synergies across the Aims, and a highly collaborative group
of established and early-stage investigators (proposing PI), we are confident that this project will lead to important
discoveries about immune regulation in the LN milieu and its impact on the HIV tissue reservoir.
