Updated publication reference for PubMed record(s): 31337724. The formation of multimerized protein assemblies has emerged as a core component of immune signal amplification, yet the biochemical basis of this phenomenon remains unclear for many mammalian proteins within host defense pathways. The interferon-inducible protein 16 (IFI16) is a viral DNA sensor that oligomerizes upon binding to nuclear viral DNA and induces downstream antiviral responses. Here, we first generated oligomerization-incompetent IFI16 mutants that exhibit severely reduced ability to induce antiviral cytokine expression, suppress herpes simplex virus 1 (HSV-1) protein levels, and restrict viral progeny production. Using immunoaffinity purification and targeted mass spectrometry, we establish that oligomerization promotes IFI16 interactions with several proteins involved in transcriptional regulation, including PAF1C, UBTF, and ND10