Salivary gland (SG) stem cells are the only cell population capable of extended growth in organotypic cultures, and thus they are considered a source for cell-based therapies aimed at SG regeneration. Studies in the mouse submandibular gland have identified only one population of tissue stem cells capable of salisphere formation in culture. These cells are actively dividing ductal cells that express epithelial progenitor markers keratin (K) 5/14 and normally function as lineage-restricted stem cells for differentiated ductal cells. In response to severe injury, however, these cells undergo a multipotency switch and contribute to regeneration of multiple cell lineages, including secretory units or acini. Little is known about the mechanism of cell renewal and regeneration in the other major SGs and whether comparable stem cell populations exist in the parotid (PG) and sublingual (SLG) glands. Using in vivo and ex vivo models, we show that both the PG and SLG contain a small population of K14-expressing ductal cells. Although they do not cycle frequently, K14-expressing ductal cells are the source of salisphere-forming cells in these glands. Long-term lineage tracing studies in adult mouse PGs showed a progenitor-progeny relationship between the K14-expressing ductal cells and the K19-expressing ductal cells in the striated ducts. In the SLGs, however, K14-expressing ductal cells did not generate a differentiated cell progeny for a 6-month period of observation and did not make a significant contribution to regeneration of gland after severe injury. These studies reveal the functional similarities and differences in tissue stem cells among the major SGs and have implications for developing strategies for SG regenerative therapies.