The progressive depletion of quiescent “bystander” CD4 T cells, which are nonpermissive to HIV infection, is a principal driver of the acquired immunodeficiency syndrome (AIDS). These cells undergo abortive infection characterized by the cytosolic accumulation of incomplete HIV reverse transcripts. These viral DNAs are sensed by an unidentified host sensor that triggers an innate immune response, leading to caspase-1 activation and pyroptosis. Using unbiased proteomic and targeted biochemical approaches, as well as two independent methods of lentiviral short hairpin RNA-mediated gene knockdown in primary CD4 T cells, we identify interferon-γ-inducible protein 16 (IFI16) as a host DNA sensor required for CD4 T cell death due to abortive HIV infection. These findings provide insights into a key host pathway that plays a central role in CD4 T cell depletion during disease progression to AIDS.