Chemokines are key regulators of hematopoiesis and host defense. We report here that functional expression of the chemokine receptor CXCR4 on human immature CD34+ hematopoietic progenitors was increased as a result of sustained elevation in cellular cAMP by db-cAMP and prostaglandin E2. This effect of cAMP was specifically mediated by PKCδ activity. CXCR4 expression and PKCδ activation by cAMP were decreased after the inhibition of cAMP effector-Rap1 by Spa1 overexpression. Interference with the activation of Rac1, a downstream target of Rap1, prevented the cAMP-induced increase in PKCδ activity and CXCR4 levels. Functional manifestation of the effects of cAMP-elevating agents revealed an increased ability of human CD34+ cells to transmigrate the bone marrow (BM) endothelial layer and adhere to BM stroma in vitro, and it augmented the homing potential to the BM and spleens of immunodeficient mice in a Rac1- and a PKCδ-dependent manner. cAMP- and TNFα-stimulated pathways converged in PKCδ-activated CXCR4 expression and MMP-2/MMP-9 secretion. cAMP treatment had a beneficial effect on CD34+ cell survival in a PKCδ-mediated fashion. Taken together, our data reveal major roles for cAMP-induced PKCδ activation in signaling governing the motility and development of CD34+ cells.