Mucus-secreting cells of the stomach epithelium provide a protective barrier against damage that might result from bacterial colonization or other stimuli. Impaired barrier function contributes to chronic inflammation and cancer. Knock-out mice for the epithelium-specific transcription factor Spdef (also called Pdef) have defects in terminal differentiation of intestinal and bronchial secretory cells. We sought to determine the physiologic function of Spdef in the stomach, another site of significant levels of Spdef expression. We used in situ hybridization and immunohistochemistry to localize Spdef-expressing cells in the mouse stomach; targeted gene disruption to generate mice lacking Spdef; and histologic, immunologic, and transcriptional profiling approaches to determine the requirements of Spdef in stomach epithelial homeostasis. In wild-type mice, Spdef RNA and protein are expressed predominantly in mucous gland cells of the antrum and in mucous neck cells of the glandular corpus. Within 1.5 years, nearly half of homozygous mutant mice developed profound mucosal hyperplasia of the gastric antrum. Submucosal infiltration of inflammatory cells preceded antral hyperplasia by several weeks. The absence of Spdef impaired terminal maturation of antral mucous gland cells, as reflected in reduced expression of Muc6 and Tff2 and reduced numbers of secretory granules. Antral gene expression abnormalities overlapped significantly with those in Spdef(-/-) colon, including genes implicated in secretory granule traffic and functions. Spdef is required for terminal maturation of antral mucous gland cells to protect animals from gastric inflammation and resulting hyperplasia. These requirements parallel Spdef functions in secretory intestinal cells and suggest a common molecular mechanism for maturation of gastrointestinal secretory lineages.