Genome-wide CRISPR/Cas Knockout Screen to Identify Proteins Involved in Silencing Unintegrated HIV-1 DNA.

Unintegrated HIV-1 DNA is efficiently epigenetically silenced in infected cells. Using an unbiased genetic screen, we identify all eight components of the host SMC5/6 complex as essential for this process. SMC5/6 first binds to chromatinized, unintegrated HIV-1 DNA and then triggers epigenetic silencing by inducing its SUMOylation. Inhibition of this SUMOylation step, either by point mutagenesis of the SMC5/6 component NSMCE2, a SUMO E3 ligase, or using the SUMOylation inhibitor TAK-981, blocks epigenetic silencing and rescues transcription from unintegrated HIV-1 DNA. Remarkably, we show that T cells lacking a functional SMC5/6 complex can support a spreading HIV-1 infection in vitro in the absence of integrase function. We further demonstrate that loss of SUMOylation results in fewer latent HIV-1 infections in primary human T cells, thus arguing that the epigenetic silencing of integration competent HIV-1 proviruses by the SMC5/6 complex prior to integration can give rise to latent HIV-1 infections.