The Jun N-terminal kinase (JNK) pathway is an evolutionary conserved kinase cascade best known for its roles during stress-induced apoptosis and tumor progression. Recent findings, however, have identified new roles for this pleiotropic pathway in stem cells during regenerative responses and in cellular plasticity. Here, we provide an overview of recent findings about the new roles of JNK signaling in stem cell biology using two well-established models: the testis and the intestine. We highlight the pathway's roles in processes such as proliferation, death, self-renewal and reprogramming, and discuss the known parallels between flies and mammals.