Functional Impact of Collagens on the Activity Directed by the Promoter of the α5 Integrin Subunit Gene in Corneal Epithelial Cells

PURPOSE. The early step of corneal wound healing is characterized by the massive production of fibronectin (FN), whose secretion is progressively replaced by collagens from the basal membrane as wound healing proceeds. Here, we examined whether expression of the gene encoding the alpha 5 subunit from the FN-binding integrin alpha 5 beta 1 changes as corneal epithelial cells (CECs) are cultured in the presence of collagen type I (CI) or type IV (CIV). METHODS. Responsiveness of the alpha 5 gene toward collagen was determined by transfection of alpha 5 promoter/chloramphenicol acetyltransferase (CAT) plasmids into rabbit and human CECs cultured on BSA or collagens. Electrophoretic mobility shift assays and Western blots were used to monitor the transcription factors required for basal alpha 5 gene transcription in the presence of collagens. Gene profiling on microarrays was used to determine the impact of collagens on the patterns of genes expressed by CECs. RESULTS. All collagen types repressed the full-length alpha 5/CAT promoter activity in confluent CECs. A moderate increase was observed in subconfluent rabbit CECs grown on CIV but not on CI. These collagen-dependent regulatory influences also correlated with alterations in the transcription factors Sp1/Sp3, NFI, and AP-1 that ensure alpha 5 gene basal transcription. Microarray analyses revealed that CI more profoundly altered the pattern of genes expressed by human CECs than CIV. CONCLUSIONS. Collagens considerably suppressed alpha 5 gene expression in CECs, suggesting that during wound healing, they may interfere with the influence FN exerts on CECs by altering their adhesive and migratory properties through a mechanism involving a reduction in alpha 5 gene expression.