Quantitative switch in integrin expression accompanies differentiation of F9 cells treated with retinoic acid

Abstract

F9 embryonal carcinoma cells resemble epithelial cells when in monolayer culture. After treatment with retinoic acid these cells differentiate into fibroblastic‐like cells in a sequence that has been modeled as the mammalian equivalent of the differentiation from stem cells of the inner cell mass to parietal endoderm. This study examined the changes in integrin subtypes that accompany retinoic acid‐induced differentiation of F9 cells. Although several integrins were found to be present on the surface of F9 cells and retinoic acid‐induced (RA) cells, the two dominant integrins were α3β1 and α5β1. Differentiation of F9 cells resulted in about 10‐ to 25‐fold increase in the amount of α3β1 integrin protein as measured by immunoprecipitation of cell surface labeled material. There was a corresponding several‐fold reduction of α5β1 protein. The concentration of α3 mRNA was about the same in F9 and RA cells while the concentration of α5 mRNA dropped several‐fold after retinoic acid treatment. Thus α3 regulation appeared to be largely posttranscriptional while the drop in α5 protein may have been a result of transcriptional down‐regulation. Quantitative measurement of adhesion suggested that most of the F9 and RA cell‐substrate adhesion to fibronectin or laminin is mediated by these integrins. They are the dominant integrins present, and antibodies to either these integrins or to the substrate blocked the adhesion. Despite the large switch in integrin subtype protein expression there was little difference between the two cell types in initial cell interactions when adhesive affinities were measured quantitatively. Also there was no difference between the two phenotypes in rate of initial adhesive strengthening. The phenotypic difference was first observed with later events in the attachment and spreading of the RA‐treated cells to the substrate. These results show that retinoic acid treatment alters the amounts of α5 and α3 integrin subunits during the F9 to RA phenotypic switch. The data show that these integrins are important in the cell‐substrate adhesion to fibronectin and laminin. They show, however, that the phenotypic changes observed with differentiation are not associated with the initial preferential adhesions to the substrate, but rather with consequences that alter the cytoskeletal architecture of the cell. © 1994 Wiley‐Liss, Inc. Copyright © 1994 Wiley‐Liss, Inc.