Hypoxia Inducible Factor 3α Plays a Critical Role in Alveolarization and Distal Epithelial Cell Differentiation during Mouse Lung Development

Lung development occurs under relative hypoxia and the most important oxygen-sensitive response pathway is driven by Hypoxia Inducible Factors (HIF). HIFs are heterodimeric transcription factors of an oxygen-sensitive subunit, HIF alpha, and a constitutively expressed subunit, HIF1 beta. HIF1 alpha and HIF2 alpha, encoded by two separate genes, contribute to the activation of hypoxia inducible genes. A third HIF alpha gene, HIF3 alpha, is subject to alternative promoter usage and splicing, leading to three major isoforms, HIF3 alpha, NEPAS and IPAS. HIF3 alpha gene products add to the complexity of the hypoxia response as they function as dominant negative inhibitors (IPAS) or weak transcriptional activators (HIF3 alpha/NEPAS). Previously, we and others have shown the importance of the Hif1 alpha and Hif2 alpha factors in lung development, and here we investigated the role of Hif3 alpha during pulmonary development. Therefore, HIF3 alpha was conditionally expressed in airway epithelial cells during gestation and although HIF3 alpha transgenic mice were born alive and appeared normal, their lungs showed clear abnormalities, including a post-pseudoglandular branching defect and a decreased number of alveoli. The HIF3 alpha expressing lungs displayed reduced numbers of Clara cells, alveolar epithelial type I and type II cells. As a result of HIF3 alpha expression, the level of Hif2 alpha was reduced, but that of Hif1 alpha was not affected. Two regulatory genes, Rarb, involved in alveologenesis, and Foxp2, a transcriptional repressor of the Clara cell specific Ccsp gene, were significantly upregulated in the HIF3 alpha expressing lungs. In addition, aberrant basal cells were observed distally as determined by the expression of Sox2 and p63. We show that Hif3 alpha binds a conserved HRE site in the Sox2 promoter and weakly transactivated a reporter construct containing the Sox2 promoter region. Moreover, Hif3 alpha affected the expression of genes not typically involved in the hypoxia response, providing evidence for a novel function of Hif3 alpha beyond the hypoxia response.