Background: Kalinin, an anchoring filament-associated protein, is a member of the laminin family which is believed to play a role in the attachment of particular epithelial cells to the basement membrane.
Experimental design: A newborn mouse lung cDNA library was screened under low stringency conditions using as a probe a murine laminin beta 1 chain cDNA, an approach designed to identify novel laminin beta 1 chain homologs.
Results: We have identified a novel cDNA sequence comprising 4049 base pairs, with an open reading frame that potentially codes for 1168 amino acids, including a presumptive signal peptide. The gene was mapped to mouse chromosome 1. The amino acid sequence is homologous to laminin beta 1 and beta 2 chains, but is distinguished from these molecules by having a truncated short arm region. The sequence has considerable similarity to the human kalinin B1 chain (laminin beta 3), suggesting that it is murine beta 3 chain. Northern blot analysis demonstrated a single 4.0-kilobase species in the mouse newborn skin, lung, and Pam 212 cells. Immunolocalization studies using the 14-day embryo and adult mouse tissues demonstrated that the murine beta 3 chain is located at the dermal-epidermal junction of the skin and the basement membrane of the oral mucosa, oropharynx, and upper portions of the tracheobronchial tree. In situ hybridization studies of the mammary gland showed that the beta 3 mRNA chain was produced by the epithelial cells of the gland but not by the mesenchymal cells. Western blotting under nonreducing conditions using the antibody to beta 3 chain detected molecules of 400 and 440 kilodaltons (kDa) in the conditioned medium of human foreskin keratinocytes, A431 cells, and Pam 212 cells. Western blotting under reducing conditions detected only a single 140-kDa species, suggesting that beta 3 chain is present within disulfide-linked complexes. This 140-kDa species was contained in the kalinin complex immunoprecipitated by anti-kalinin B2 (laminin gamma 2) antibody from the conditioned medium of HT1080 cells, suggesting that both the beta 3 and gamma 2 chains are present in the multimeric complex.
Conclusions: Murine laminin beta 3 chain shows considerable homology to the human homolog and has a tightly restricted tissue distribution, confined to the basement membrane of cells in contact with the external environment. In vitro studies indicate that the beta 3 chain participates in trimer formation with the gamma 2 chain and possibly with other laminin chains as well.