Abstract
We previously demonstrated that blocking TGF-β with galunisertib, a safe, orally available small drug, reactivated latent SIV in vivo by shifting T cells toward a transitional effector phenotype. Here, we investigated the mechanisms underlying this effect using single-cell RNA sequencing, metabolic profiling, and high-dimensional spectral flow cytometry of samples from SIV-infected, antiretroviral therapy–treated (ART-treated) macaques before and after galunisertib. To characterize virus-transcribing, infected cells during ART, we developed a novel, sensitive SIV Transcripts Capture Assay (SCAP) that detected 127 SIV-expressing cells within lymph node single-cell transcriptome libraries. Galunisertib drove broad metabolic reprogramming in CD4+ T cells, with transcriptional upregulation of inflammatory and mitochondrial biosynthesis pathways, confirmed by Seahorse profiling. Metabolomics revealed increased energy metabolites and amino acids and enhanced metabolic flux without proliferation. SIV transcript–positive cells before galunisertib were metabolically quiescent compared with cells without detectable viral transcripts. After galunisertib, virus-expressing cells showed a dramatic metabolic activation, with upregulation of glycolysis, fatty acid metabolism, and TNF-α signaling. High-dimensional flow cytometry demonstrated effects beyond CD4+ T cells, including fewer tissue-resident memory T cells, but more inflammatory macrophages. In conclusion, SCAP represents a specific tool for characterizing rare SIV-infected cells transcribing virus during ART, and it reveals TGF-β as a key mediator of viral latency in vivo through metabolic suppression.
Authors
Romaila Abd-El-Raouf, Jakob Harrison-Gleason, Jinhee Kim, Ching Man Wai, Kayla L. Yerlioglu, Catarina Ananias-Saez, Alec Ksiazek, Jeffrey T. Poomkudy, Mariluz Araínga, Deepanwita Bose, Claudia Cicala, James Arthos, Francois J. Villinger, Ramon Lorenzo-Redondo, Elena Martinelli
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