Context: Progesterone (P4)contributes to the maintenance of human pregnancy, in part by inhibiting activity of the human pro-labor genes CRH and cyclooxygenase-2 (COX-2). However, the molecular mechanisms underlying the action of P4 remain poorly defined. We have shown that in human placenta, the constitutively activated noncanonical nuclear factor (NF)-κB pathway positively regulates CRH and COX-2, which is further stimulated by glucocorticoid receptor signaling. Objective: We investigated the role of P4 receptor (PR) in the regulation of nuclear activity of v-rel avian reticuloendotheliosis viral oncogene homolog B (RelB)/NF-κB2 and, in turn, expression of placental CRH and COX-2. Methods: We used a variety of techniques including gene silencing, ectopic expression, chromatin immunoprecipitation, Western blot, quantitative RT-PCR, and immunohistochemical staining assays in human placental tissues and primary culture of human cytotrophoblast. Results: We identified PR isoform-A (PR-A) as the only isoform of PR produced in human placenta. PR-A levels were lower in term placenta than in midterm placenta. Depletion of PR-A by short interfering RNA derepressed inhibition of CRH and COX-2 by P4 and the synthetic progestin 17α- hydroxyprogesterone caproate. Overexpression of PR-A inhibited transcription of CRH and COX-2, which was further downregulated by treatment with P4 or 17α-hydroxyprogesterone caproate. Such an inhibition was mediated by a negative functional interaction of PR-A with the activity of RelB/NF-κB2. Conclusion: P4 inhibits the pro-labor genesCRHand COX-2 via PR-A repression of the noncanonical NF-κB signaling in human placenta. Characterization of these pathways may identify potential drug targets for prevention of preterm birth.