P29-17. Constitutive expression of Siglec-1 by human circulating dendritic cell precursors enables HIV-1 replication and dissemination to CD4+ T cells

Nicolas Ruffin [1], Ester Gea-Mallorquí [1], Mabel Jouve [2], Flavien Brouiller [1], Aymeric Silvin [1,3], Peter See [3], Charles-Antoine Dutertre [3,4], Florent Ginhoux [3]*, Philippe Benaroch [1]*
Affiliates: [1] Institut Curie, PSL* Research University, INSERM U 932, F-75005, Paris, France. [2] UMR3215, Institut Curie, 75248 Paris Cedex 05, France. [3] Singapore Immunology Network (SIgN), A*STAR, 8A Biomedical Grove, Immunos Building, Level 4, Singapore 138648, Singapore. [4] Program in Emerging Infectious Disease, Duke-NUS Medical School, 8 College Road, 169857 Singapore. *These authors contributed equally to this work.

Background
Human dendritic cell (DC) lineage has been recently unraveled by high dimensional mapping revealing the existence of a discrete new population of blood circulating DC precursor (pre-DC). Among all blood DC subsets including cDC1, cDC2 and pDC, only pre-DC highly express the lectin-like receptor Siglec-1, a molecule known for its capacity to bind HIV-1. Whether this expression confers selective properties to pre-DC with regards to HIV-1 remains unknown.

Methods
We set up an isolation strategy of the four blood DC subsets (pre-DC, pDC, cDC1 and cDC2) based on their differential surface protein expression, by a combination of magnetic bead negative selection and flow cytometry sorting. Sorted cell populations were then exposed to CCR5- or CXCR4-tropic HIV-1 expressing GFP. HIV-1 capture and infection of dendritic cells were assessed by flow cytometry and electronic microscopy. Transmission of HIV-1 to activated primary CD4+ T cells was assessed following DC-T cell co-culture. In some experiments, TLR stimulation of the DC population was performed before HIV-1 capture or infection.

Results
We show that pre-DC are uniquely equipped among blood DC populations to promote HIV-1 replication and dissemination. Pre-DC stands out as the most susceptible DC population to infection by both HIV-1 CXCR4- and CCR5-tropic viral particles in a Siglec-1-dependent manner. In contrast to cDC2 cells for which HIV-1 assembly occurs at the plasma membrane, HIV-1-infected pre-DC produce and accumulate new viral progeny into internal compartments connected to the extracellular medium. HIV-1-infected pre-DC can transmit the virus in cis to activated CD4+ T cells. Upon activation with TLR-ligand, pre-DC become resistant to HIV-1 infection and switch to a replication-independent mechanism of virus transfer to activated primary T lymphocytes mediated by Siglec-1.

Conclusions
Beside their role in DC ontogeny, blood pre-DC possess stage-specific properties that HIV-1 may exploit for viral spreading and modulation of the immune response. As a result of constitutive Siglec-1 expression, blood pre-DC likely represent a pivotal HIV target cell during establishment of the infection and induction of the anti-viral immune response.