G-Quadruplex Folding Enforces HIV-1 Transcriptional Latency via Nucleolin Recruitment

Despite antiretroviral therapy, the persistence of latently infected cellular reservoirs impedes HIV-1 eradication. G-quadruplexes (G4s) form in the HIV-1 LTR promoter, but their role has not been proven in infected cells. Combining virus reactivation, cell sorting and ChIP-qPCR, we demonstrate that G4s are folded in the transcriptionally silent LTR, while they are unfolded upon viral reactivation. These dynamics are orchestrated by different LTR interacting proteins. Specifically, nucleolin binds LTR G4s in the latent state, whereas the transcription factor SP1 binds the unfolded G-rich promoter in the active state. Nucleolin depletion induces reactivation, confirming its role in maintaining HIV latency. LTR G4 folding also correlates with low viral transcription in peripheral blood mononuclear cells from low-viremic patients living with HIV. These findings uncover G4s as molecular switches controlling HIV-1 latency, identify nucleolin as a key regulator, and indicate G4/nucleolin-targeting as potentially effective approaches to both a functional cure and virus eradication.