Expression of novel 400-kDa laminin chains by mouse and bovine endothelial cells.
Laminin expression was studied in endothelial cells derived from different mouse tissues and primary cultures of bovine aortic endothelium (BAEC). Immunoprecipitation with polyclonal anti-laminin-1 revealed two 200-kDa chains, a novel 400-kDa chain and nidogen in all cells studied. Two-dimensional electrophoresis of laminin complexes immunoprecipitated from endothelial-cell-conditioned medium demonstrated that the 400-kDa chain was disulphide-linked to the laminin beta 1 and gamma 1 chains. In addition, rotary shadowing illustrated the secretion of molecules from the endothelial cells which were morphologically similar to laminin-1. Immunoblotting and Northern blot analysis confirmed the presence of beta 1 and gamma 1 polypeptides and the corresponding LamB1 and LamC1 mRNAs in all cells investigated. However, both polyclonal anti-laminin-1 and alpha 1-specific monoclonal antibodies failed to react with the 400-kDa polypeptide in immunoblots and immunoprecipitations. Similarly, the expression of the gene coding for alpha 1, LamA1, was not detected in the majority of cells by Northern blot analysis using three different cDNAs. Only in two cases, BAEC and a Simian-virus-40-transformed mouse endothelial cell line (SVEC), a 10-kb mRNA was detected by Northern analysis using a cDNA specific for the 3' coding region (LAC) of LamA1 mRNA. However, cDNAs specific for the central and 5' coding region of LamA1 mRNA did not show any reaction. Hybridization of LAC to BAEC and SVEC mRNA is consistent with the laminin-like nature of the 400-kDa chain expressed by these cells. The results demonstrate the existence of at least one and possibly two novel 400-kDa laminin chains which complex with beta 1 and gamma 1 and occur in mouse and bovine endothelial cell basement membranes.
Sorokin
L
L
Max-Planck-Society, Connective Tissue Research Unit, Erlangen, Germany.
Girg
W
W
Göpfert
T
T
Hallmann
R
R
Deutzmann
R
R
D000818:Animals; D000906:Antibodies; D000918:Antibody Specificity; D001485:Basement Membrane / Q000378:metabolism; D015152:Blotting, Northern; D002417:Cattle; D002460:Cell Line; D017077:Culture Media, Conditioned; D004591:Electrophoresis, Polyacrylamide Gel; D004727:Endothelium / Q000166:cytology / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D015870:Gene Expression; D007150:Immunohistochemistry; D007797:Laminin / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D051379:Mice; D008970:Molecular Weight; D009693:Nucleic Acid Hybridization; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000096:biosynthesis; D014407:Tumor Cells, Cultured
D000906:Antibodies; D017077:Culture Media, Conditioned; D007797:Laminin; D012333:RNA, Messenger