Nef stabilizes actin to prevent HIV-1 sensing by RIG-I-like receptors

Sensing of viral pathogens by RIG-I-like receptors (RLRs) requires their priming via dephosphorylation mediated by the protein phosphatase 1 regulatory subunit 12C (R12C), which is activated upon virus-induced actin rearrangements. Here, we show that the HIV-1 accessory protein Nef disrupts R12C-mediated RLR priming, thereby preventing efficient viral sensing. HIV-1 variants containing single point mutations in Nef (F/R191A) that ablate its ability to bind the actin-modulating kinase PAK2 trigger increased interferon (IFN) responses in primary CD4+ T cells, macrophages, and dendritic cells. Neutralization of IFN suppressed innate immune activation and enhanced the replication of Nef-mutated HIV-1. We further demonstrate that HIV-1 encoding Nef F/R191A is sensed by MDA5 after proviral integration in an R12C-dependent manner. Mechanistically, PAK2 binding by Nef prevents the re-localization of R12C to MDA5 and its subsequent dephosphorylation. Our data identify Nef as an antagonist of actin-R12C-mediated RLR priming, enabling HIV-1 to escape immune control.