Collagen 6 is a Critical Component of Lung Structure andPulmonary Epithelial Cell Function

Basement membrane (BM) is a significant extracellular matrix (ECM) component, playing critical structural and signaling roles in the lung. Abnormal ECM composition and remodeling is associated with multiple respiratory diseases, including bronchopulmonary dysplasia (BPD), a chronic lung disease commonly affecting infants born before 29 weeks of gestation. We identified altered sequence and expression of BM gene, COL6A5 in association with BPD, and reduced collagen VI (COL6) protein levels in lung tissue from BPD patients. COL6 mutations cause a spectrum of muscular dystrophies, but its role in the lung is largely undetermined. We tested the hypothesis that COL6 plays a role in fundamental aspects of lung structure and pulmonary epithelial cell function. We examined the effects of COL6 loss in the lungs of a previously described Col6a1-/- mouse (Bonaldo et. al., 1998, Hum Mol Genet) finding numerous abnormalities, including spontaneous airspace enlargement. We observed reduced RNAseq gene expression signatures and staining of alveolar endothelium and alveolar epithelial type-II (ATII) cells in Col6a1-/- lungs. Airway epithelial cell density was increased in Col6a1-/- animals. Similarly, in vitro data using primary human lung epithelial cells (PHLE) and human lung epithelial cell lines (16HBE), shows increased steady-state cell density on matrices (Matrigel, a prototypical BM, and collagen I) not containing COL6. Characterization of cell-autonomous effects of COL6 revealed increased "wound-healing" rate after scratch injury, and cell-spreading on COL6 compared to other matrices. Importantly, combining COL6 with other matrices also enhanced cell-spreading and wound-healing. Inhibitor studies indicated activity of PI3K, Cdc42, and Rac1, likely downstream of integrin β1, significantly and specifically affect spreading on COL6. Constitutive activation of PI3K and Cdc42 was sufficient to enhance cell-spreading on other matrices to comparable levels as COL6. Pathway analysis of RNAseq data from WT and Col6a1-/- lung tissue identified reduced paxillin signaling, a known PI3K and Cdc42 regulator possibly involved in the response to COL6. In summary, we find that COL6 plays an important role in structural organization and epithelial cell function in the lung. COL6 loss mimics some of the pathophysiology of BPD, and may provide a useful model in its study.