Wu, Chuanyue

ABSTRACT To study the biological role of α3β1 integrins in cell adhesion, migration, and in the deposition of extracellular matrix, we stably expressed the human α3 integrin subunit in the α4, α5 integrin deficient CHO cell line B2. The expression of α3β1 integrins enhanced cell adhesion on entactin (also known as nidogen), but not on fibronectin. Using recombinant GST-fusion proteins that span the entire length of the entactin molecule, we located cell adhesive activity to the G2 domain of entactin. These results suggest that the α3β1 integrin functions as an adhesion receptor interacting with the G2 domain of entactin. On the other hand, the expression of α3β1 integrins did not confer the ability to migrate on entactin. Strikingly, the expression of α3β1 dramatically increased the deposition of entactin and fibronectin into the pericellular matrix. This was accompanied by increased binding activity of the 29 kDa amino-terminal domain of fibronectin. Thus, similar to α5β1 integrins, α3β1 integrins can play an important role in modulating the assembly of pericellular matrices. However, unlike fibronectin deposition supported by α5β1, α3β1 supported fibronectin deposition into pericellular matrix was not inhibited by antibodies binding to the RGD containing cell adhesion domain of fibronectin, demonstrating that the two processes are mechanistically distinct. The role of α3β1 in pericellular matrix assembly potentially implicates this receptor in the assembly and/or recognition of entactin-containing pericellular matrices, an observation consistent with its apparent role in the renal glomerulus of the mammalian kidney.

ABSTRACT The α4β1 integrin has been suggested to play important roles in embryogenesis and pathogenesis of many diseases which involve both cell adhesion and cell migration. Previous studies using anti-α4β1 antibodies and fibronectin (Fn) fragments have suggested that α4β1 integrins may be involved in cell motility on Fn and vascular cell adhesion molecule-1 (VCAM-1). However, the cells used in these studies also express other Fn integrin receptors including α5β1 integrin, which is known to function in cell motility on Fn. To test whether α4β1 integrins mediate cell motility on Fn and VCAM-1 in the absence of α5β1 integrin, we expressed human α4 integrin in a Chinese hamster ovary (CHO) cell line that is deficient in α5β1 integrin (CHO B2). The parental α5 deficient CHO B2 cells were unable to adhere, spread or migrate on Fn, nor could they assemble a fibrillar Fn matrix. Expression of α4β1 integrin in the CHO B2 cells enabled the cells to adhere, spread and migrate on Fn and on VCAM-1 but not to assemble a fibrillar Fn matrix. The cellular processes mediated by the interaction of α4β1 with Fn or VCAM-1 were inhibited by the CS1 peptide derived from the major α4β1 binding site on Fn. These findings demonstrate that α4β1 integrins not only function as cell adhesion receptors but also as cell motility receptors for Fn and VCAM-1 independent of α5β1. Moreover, they reveal important functional differences between Fn binding integrins. The α4-positive, α5-negative CHO cells described in this report will be useful tools in studying the mechanism of molecular signalling during integrin mediated cellular processes.