Integrins in the Developing Lung

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PROJECT SUMMARY Human and murine lung development requires the coordinated efforts of the lung epithelium with the surrounding extracellular matrix (ECM), but the ECM-directed mechanisms that govern epithelial cell behavior in the lung remain undefined. In epithelial tissues, integrins serve as receptors for the basement membrane components collagen and laminins (LMs), with LMs being the most important ECM protein for lung organogenesis. Lung epithelial cells bind to LMs through integrins α3β1, α6β1, and α6β4. Recent studies reported that mutations in these integrins cause pulmonary hypoplasia or neonatal emphysema complicated by abnormal airways, suggesting they play a major role in human developmental lung diseases. To investigate the role of LM-binding integrins, we generated lung epithelial specific integrin deletions. Deletion of both the β1 and α6 subunits resulted in marked branching defects and early death. Deletion of α3 caused only minor airway branching disruption. β4 deficient mice exhibited normal branching but, surprisingly, were also perinatal lethal. The β4 histological examination was notable for proteinaceous material filling the airways and lack of cilia, similar to α6 deficient mice, suggesting their demise resulted from airway dysfunction. Pathway analysis of α6 deficient lung sequencing data revealed disruptions in BMP signaling, a critical pathway for airway branching. BMP receptor expression was increased in α6-null epithelial cells, but BMP target gene expression remained markedly reduced, implicating α6-containing integrins in regulation of the BMP pathway in the fetal lung. Consistent with loss of cilia in β4 deficient mice, β4-null epithelial cells exhibited reduced expression of transcription factors linked to MCC terminal differentiation. As a critical component of hemidesmosomes, α6β4 controls tight adhesion to the basement membrane and connects with the intracellular keratin intermediate filaments. Keratin also forms a support network apically for cilia, suggesting that α6β4 regulates keratin organization critical for terminal differentiation of MCCs in the lung. Taken together, these findings indicate that: 1) α6β1 is the principal integrin required for airway branching likely through BMP signaling and 2) α6β4 regulates terminal differentiation of MCCs. Based on preliminary data, we propose the hypothesis that α6-containing integrins are critical integrins for fetal lung development through regulation of BMP signaling during airway branching and terminal differentiation of multi-ciliated epithelial cells. AIM 1: Determine the mechanisms whereby α6-containing integrins regulate lung branching morphogenesis. AIM 2: Define the mechanisms whereby α6-containing integrins regulate BMP signaling during fetal lung development. AIM 3: Identify the role of α6β4 integrin in airway epithelial cell differentiation.