Osr1 orchestrates posterior second heart field cell migration for outflow tract formation.

Congenital heart defects, particularly those affecting the outflow tract (OFT), represent a significant cause of neonatal morbidity and mortality. The genetic and cellular mechanisms underlying OFT abnormalities, especially the migration of second heart field (SHF) cells, remain poorly understood. In this study, we identify two distinct migratory pathways for Osr1-expressing posterior SHF cells directed toward the OFT and inflow tract (IFT). We show that Osr1 regulates two key ligand-receptor signaling axes, Hedgehog-Ptch1 and Cxcl12-Cxcr7, both of which are essential for the directed migration of Osr1⁺ cells toward the OFT. We establish that Smo and Ackr3 (encoding CXCR7) are direct transcriptional targets of Osr1. Functional studies demonstrate that Osr1-Hh signaling governs SHF cell migration and OFT development, while Osr1-Cxcl12 signaling likely acts in synergy to support this process. These findings underscore the pivotal role of Osr1+ pSHF cells in directing heart pole development and reveal crucial ligand-receptor interactions involved in cell migration, potentially guiding future therapeutic targets for congenital heart defects.