Pure CD32+ CD4+ Cells Are Cytotoxic Memory CD4+ T Lymphocytes Highly Enriched for HIV-1 DNA

Abstract

The elusive viral reservoir is the major obstacle to an HIV-1 cure. Cell surface protein CD32 has been proposed to pinpoint cells with a very high proviral enrichment, potentially providing a tool for selective therapeutic targeting of the reservoir. However, this finding has been challenged by subsequent reports. In order to clarify the nature of CD32 expression on CD4+ T cells, here we designed a purification strategy allowing single-cell analysis using state-of-the-art technologies. The established multilevel sorting strategy allowed to characterize a rare (median 0.06% of CD4+ T cells) but bona fide CD32+ CD4+ T-cell population. In vitro cell stimulation could not lead to CD32 upregulation. Detailed phenotyping revealed that CD32+ T cells reside mostly in the memory T-cell compartment with heightened levels of acute and chronic activation markers. Single-cell RNA sequencing identified that CD32+ CD4+ T cells are mostly highly cytotoxic CD4+ cells. Importantly. in HIV-1 infected donors under suppressive antiretroviral therapy, we found that these cells are highly enriched for HIV-1 provirus, harbor a clear cytotoxic memory T-cell transcriptome, as well as TCR clonal enrichment. These findings indicate that CD32 remains a promising candidate marker of the HIV-1 reservoir and underline the importance of further investigation of this marker during latent infection.