The Laminin a Chains: Expression, Developmental Transitions, and Chromosomal Locations of a 1-5, Identification of Heterotrimeric Laminins 8–11, and Cloning of a Novel a 3 Isoform

Laminin trimers composed of a , b , and g chains are major components of basal laminae (BLs) throughout the body. To date, three a chains ( a 1–3) have been shown to assemble into at least seven heterotrimers (called laminins 1–7). Genes encoding two additional a chains ( a 4 and a 5) have been cloned, but little is known about their expression, and their protein products have not been identified. Here we generated antisera to recombinant a 4 and a 5 and used them to identify authentic proteins in tissue extracts. Immunoprecipitation and immunoblotting showed that a 4 and a 5 assemble into four novel laminin heterotrimers (laminins 8–11: a 4 b 1 g 1, a 4 b 2 g 1, a 5 b 1 g 1, and a 5 b 2 g 1, respectively). Using a panel of nucleotide and antibody probes, we surveyed the expression of a 1-5 in murine tissues. All five chains were expressed in both embryos and adults, but each was distributed in a distinct pattern at both RNA and protein levels. Overall, a 4 and a 5 exhibited the broadest patterns of expression, while expression of a 1 was the most restricted. Immunohistochemical analysis of kidney, lung, and heart showed that the a chains were confined to extracellular matrix and, with few exceptions, to BLs. All developing and adult BLs examined contained at least one a chain, all a chains were present in multiple BLs, and some BLs contained two or three a chains. Detailed analysis of developing kidney revealed that some individual BLs, including those of the tubule and glomerulus, changed in laminin chain composition as they matured, expressing up to three different a chains and two different b chains in an elaborate and dynamic progression. Interspecific backcross mapping of the five a chain genes revealed that they are distributed on four mouse chromosomes. Finally, we identified a novel full-length a 3 isoform encoded by the Lama3 gene, which was previously believed to encode only truncated chains. Together, these results reveal remarkable diversity in BL composition and complexity in BL development. L aminins are components of all basal laminae (BLs) 1 throughout the bodies of vertebrates and invertebrates. In mammals they play at least three essential roles. First, they are major structural elements of BLs, forming one of two self-assembling networks (the other is composed of the collagens IV) to which other glycoproteins and proteoglycans of the BL attach (for review see Yurchenco and O’Rear, 1994; Timpl, 1996). Second, they interact with cell surface components such as dystroglycan to attach cells to the extracellular matrix (for review see Henry and Campbell, 1996). Third, they are signaling molecules that interact with cellular receptors such as the integrins to convey morphogenetically important information to the cell’s interior (for review see Clark and Brugge, 1995; Mercurio, 1995; Yamada and Miyamoto, 1995). For example, laminin promotes myogenesis in skeletal muscle, outgrowth of neurites from central and peripheral neurons, and mesenchymal to epithelial transitions in kidney (Foster et al., 1987; Klein et al., 1988; Reichardt and Tomaselli, 1991; Vachon et al., 1996). Laminin was initially isolated from tumor cells as a heterotrimer of A, B1, and B2 subunits (Chung et al., 1979; Timpl et al., 1979), later renamed a 1, b 1, and g 1 (Burgeson et al., 1994). Molecular cloning revealed that the three subunits were encoded by distinct but homologous genes (Martin and Timpl, 1987). Subsequently, homologues of the a 1 chain (merosin, or a 2; Ehrig et al., 1990) and the b 1 chain (s-laminin, or b 2; Hunter et al., 1989 b ) were isolated, revealing a previously unsuspected heterogeneity of lamiPlease address all correspondence to Joshua R. Sanes, Department of Anatomy and Neurobiology, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110. Tel.: (314) 362-2507. Fax: