.alpha.1.beta.1 Integrin heterodimer functions as a dual laminin/collagen receptor in neural cells

Abstract

A monoclonal antibody (3A3) raised against a rat neural cell line (PC12) was shown previously to bind to the surfaces of these cells, inhibiting substratum adhesion. Immunochemical and other data indicated that the heterodimer recognized by 3A3 was a member of the integrin family of adhesive receptors and had a β 1 subunit. The relationship of the α subunit to other integrins was unknown. Here we show that 3A3 recognizes in rat tissues a heterodimer (~185 kDa, ~110 kDa; unreduced) that is electrophoretically and immunochemically indistinguishable from the antigen in PC12 cells. Immunoaffinity purification of the heterodimer from neonatal rats and protein microsequencing indicate that the α subunit is identical at 11 or 13 N-terminal residues with VLA-1, an integrin on human hematopoietic cells. Monoclonal antibody 3A3 inhibits the attachment of rat astrocytes to laminin or collagen but not to fibronectin or polylysine. These data suggest strongly that the integrin recognized by 3A3 is the rat homologue of VLA-1, i.e., α 1 β 1 , and that α 1 β 1 is a dual laminin/collagen receptor.Previous reports described attachment and neurite outgrowth of a neural cell line (PC12) on laminin, collagen, and other adhesive proteins Tomaselli et al., 1987Tomaselli et al., , 1988. These functional studies suggested that PC12 cells possess a dual laminin/collagen receptor . Subsequently, monoclonal antibody (mab) 1 3A3 generated against PC12 cells was found to inhibit adhesion to and neurite outgrowth on laminin and collagen . mab 3A3 immunoprecipitates two proteins of ~ 110 and ~ 185 kDa. First, functional and, later, immunochemical data suggested these proteins were members of the integrin family (Hynes, 1987;Buck & Horwitz, 1987;Ruoslahti & Pierschbacher, 1987) of heterodimeric adhesive receptors. Tomaselli et al. (1987Tomaselli et al. ( , 1988 . While the β subunit of this receptor has been identified as β 1 , the identity of the α subunit has not been determined. Since the α subunits in the β 1 integrin family largely determine the ligand binding specificity of each heterodimer (Hynes, 1987;Ruoslahti & Pierschbacher, 1987), it was important to ascertain whether the α subunit had a homologue among the several integrins identified in other species to which its functional properties could be related.Here we show that the heterodimer in PC12 cells recognized by mab 3A3 is expressed in rat tissues where it is electrophoretically and immunologically indistinguishable from that in PC12 cells. Having this rich source of receptor has permitted purication and microsequencing of the larger subunit. The amino acid sequence shows unambiguously that this is an integrin α subunit identical at 11 of the first 13 N-terminal amino acid residues with the α 1 subunit of human VLA-1, an integrin in human hematopoietic cells (Hemler, 1988). These data, together with those of others (Kramer & Marks, 1989;Ignatius & Reichardt, 1988; discussed below), argue strongly that the α 1 β 1 integrin is a dual laminin/collagen integrin is hematopoi...

A dual laminin/collagen receptor acts in peripheral nerve regeneration.

“…It has been shown elsewhere (25) that mAb 3A3 binds to a heterodimeric receptor complex (185 kDa, 125 kDa) of the integrin superfamily. mAb 3A3 appears to recognize the a subunit of this laminin/collagen receptor, which can be immunoprecipitated from detergent extracts not only of PC12 cells but also of embryonic rat neural tissues (26). We show here that (i) laminin coated onto the nerve tubes enhances sciatic nerve regeneration and (ii) mAb 3A3 inhibits regeneration.…”

“…They have been identified relatively recently; hence, the structural relationships among the different members are still being detailed. The smaller subunit of the 3A3 antigen cross-reacts in Western blots with several polyclonal antisera to the 131 subunit of the integrin superfamily (19,26,47). More recently, immunoaffinity purification of the a subunit (26) has made it possible to sequence the N-terminal 26 amino acid residues.…”

“…The smaller subunit of the 3A3 antigen cross-reacts in Western blots with several polyclonal antisera to the 131 subunit of the integrin superfamily (19,26,47). More recently, immunoaffinity purification of the a subunit (26) has made it possible to sequence the N-terminal 26 amino acid residues. These data identify the receptor unequivocally as a homolog (al, 131) of one of the very late antigens (VLA 1) (S.C., N. J. Tawil, M. Houde, R. Blacher, F. Esch, L. F. Reichardt, and D. C. Turner, unpublished data), which was first described in human lymphocytes and more recently in human cell lines (48,49).…”

Developmental and injury-induced expression of α1β1 and α6β1 integrins in the rat spinal cord

“…One possibility is that α1β1 integrin is involved in angiogenesis during development in addition to the proposed role in blood-spinal barrier formation. The main difference between α1β1 and α6β1 integrin is the binding of α1β1 integrin to collagen as well as laminin (Ignatius et al, 1990;Tawil et al, 1990). Collagen is expressed by developing blood vessels at P15 (Rafalowska et al, 2000).…”

“…Two predominant laminin-binding integrins are α1β1 (Ignatius et al, 1990;Tawil et al, 1990) and α6β1 (Sonnenberg et al, 1990). The α1β1 integrin can also bind collagen (preferentially collagen-IV; Ignatius et al, 1990;Tawil et al, 1990). The α1, α6 and β1 integrin subunits are expressed by the vasculature in rodent brain (Kloss et al, 1999;Milner and Campbell, 2002a).…”