Thirty years after the discovery of its production by activated macrophages, our appreciation of the diverse roles of nitric oxide (NO) continues to grow. Recent findings have not only expanded our understanding of the mechanisms controlling the expression of NO synthases (NOS) in innate and adaptive immune cells, but have also revealed new functions and modes of action of NO in the control and escape of infectious pathogens, in T and B cell differentiation, and in tumor defense. I discuss these findings, in the context of a comprehensive overview of the various sources and multiple reaction partners of NO, and of the regulation of NOS2 by micromilieu factors, antisense RNAs, and ‘unexpected' cytokines.