The alarmin S100A9 is a calcium-binding protein known for its pro-inflammatory extracellular role in the immune response against pathogens in the form of a S100A8/S100A9 heterodimer called calprotectin. However, S100A9 was recently shown to exert an intracellular antiviral effect against HIV-1 infection of Langherans cells, a cutaneous- and mucosal-resident subset of dendritic cells. The calcium-dependent ability of S100A9 and calprotectin to bind various divalent cations is required for its known pro-inflammatory and antibacterial roles, and previous in vitro experiments point towards a role of S100A9-dependent magnesium chelation in HIV-1 reverse transcriptase (RT) inhibition. Our project aims to better characterize the role of calcium in S100A9-mediated HIV-1 restriction in immune myeloid cell subsets. Using a homemade S100A9 CRISPR-knockout THP-1 cellular model, we generated myeloid THP-1 cell lines stably expressing wild-type and calcium-binding deficient mutants of S100A9. Our results show that cells expressing calcium-binding mutants exhibit no antiviral effect against HIV-GFP pseudotyped lentivectors or wild-type HIV-1 infection compared to S100A9-WT expressing THP-1 cells. Additionally, the observed inhibition of HIV-RT enzymatic activity in-vitro by S100A9-WT protein is lost upon targeted calcium-binding site mutations. Finally, pharmacological increase of intracellular calcium stores into the cytosol with ionomycin shortly after virus entry overcomes S100A9-WT antiviral effect while having no effect on calcium-binding deficient mutants or when treatment is applied at later stages of the viral replication cycle. Together, these results suggest a calcium-dependent antiviral effect of alarmin S100A9 against HIV-1 in a human myeloid cell model.
Audience take away:
- These results characterize a new intracellular mechanism for the alarmin S100A9, highly expressed in myeloid cells, expanding the scope of its known functions beyond the well-described extracellular functions.
- Cellular antiviral factors can contribute to the discovery and development of therapeutical solutions against HIV-1.