Nucleic acids (NAs) represent one of the most important classes of molecules recognized by the innate immune system. However, NAs are not limited to pathogens, but are also present within the host. As such, the immune system has evolved an elaborate set of pathogen recognition receptors (PRRs) that employ various strategies to recognize distinct types of NAs, while reliably distinguishing between self and nonself. The here‐employed strategies encompass the positioning of NA‐sensing PRRs in certain subcellular compartments that potentially come in contact with pathogens but not host NAs, the existence of counterregulatory measures that keep endogenous NAs below a certain threshold, and also the specific identification of certain nonself patterns. Here, we review recent advances in the molecular mechanisms of NA recognition by TLRs, RLRs, and the cGAS–STING axis. We highlight the differences in NA‐PRR interfaces that confer specificity and selectivity toward an NA ligand, as well as the NA‐dependent induced conformational changes required for signal transduction. Detection of nonself nucleic acids (NAs) by cells of the innate immune system plays a pivotal role in pathogen defense, but also in the context of sterile inflammatory diseases. Here, we discuss our current knowledge of the molecular mechanisms of the major NA‐sensing pathways that are operational in the human system.