The presence of DNA in the cytoplasm of mammalian cells is perceived as a danger signal, alerting the host to the presence of microbial infection. In response to the detection of cytoplasmic DNA, the immune system mounts a programed response that involves the transcription of anti-viral genes such as type I interferons and production of inflammatory cytokines such as IL-1beta. The recent discovery of the cGAS-cGAMP second messenger pathway as well as IFI16 and additional sensors collectively provide critical insights into the molecular basis behind the sensing of cytoplasmic DNA. The insights obtained from these important discoveries could unveil new avenues to understand host-immunity, improve vaccine adjuvancy, and allow development of new treatments for inflammatory diseases associated with abberrant sensing of DNA.