Null mutation of the Gαi2 trimeric G protein results in a discrete and profound mucosal disorder, including inflammatory bowel disease (IBD), attenuation of IL-10 expression, and immune function polarized to Th1 activity. Genetic and adoptive transfer experiments have established a role for B cells and IL-10 in mucosal immunologic homeostasis and IBD resistance. In this study, we addressed the hypothesis that Gαi2 is required for the development of IL-10-producing B cells. Gαi2−/− mice were reduced in the relative abundance of marginal zone (MZ), transitional type 2 (T2), and B-1a B cells and significantly increased in follicular mature and B-1b B cells. Reconstitution of RAG2−/− mice with Gαi2−/− bone marrow induced an IBD-like colitis and a deficiency in absolute numbers of MZ, T2, and B-1 B cells. Thus, the Gαi2−/− genotype in colitis susceptibility and B cell development involved a cis effect within the hemopoietic compartment. In vitro, the B cell population of Gαi2−/− mice was functionally deficient in LPS-induced proliferation and IL-10 production, consistent with the exclusive capacity of T2 and MZ cell subpopulations for LPS responsiveness. In vivo, Gαi2−/− mice were selectively impaired for the IgM response to T-independent type II, consistent with the relative depletion of MZ and peritoneal B-1 subpopulations. Collectively, these results reveal a selective role for Gαi2 in MZ and B-1 B cell development. Disorders of this Gαi2-dependent process in B cell development may represent a mechanism for IBD susceptibility.