BeWo choriocarcinoma cells produce laminin 10.
BeWo is a choriocarcinoma cell line that generates an extracellular matrix (ECM) rich in laminin and is a useful model for human trophoblast. Immunofluorescence with monoclonal antibodies demonstrates that BeWo ECM contains laminin subunits beta1 and gamma1. Immunoprecipitation from conditioned medium shows that the cells secrete two distinct laminin trimers both containing beta1 and gamma1 but with alpha subunits of approx. 400 and 450 kDa. The culture medium also contains a species thought to be beta1 gamma1 dimer. Immunoprecipitation with monoclonal antibody 4C7, previously thought to recognize the alpha1 subunit, isolates complexes containing only the smaller alpha subunit. A second complex containing the larger alpha subunit along with beta1, gamma1 and a 150 kDa polypeptide is precipitated from 4C7-depleted medium with an anti-(laminin 1) polyclonal antibody. Peptide sequencing demonstrates that the 4C7-reactive species is alpha5, which is present as two similarly sized polypeptides. mRNA species encoding laminin subunits alpha1, alpha5, beta1, beta2 and gamma1 are all present in the cells. These results demonstrate the secretion of a novel laminin isoform, laminin 10, the subunit composition of which is alpha5 beta1 gamma1. Laminin 1 is also produced. No evidence for the secretion of beta2-containing laminin isoforms could be derived despite the presence of beta2 mRNA. Analysis with reverse transcriptase-mediated PCR also showed the presence of laminin alpha5 in first-trimester placenta and decidua.
Church
H J
HJ
Department of Obstetrics and Gynaecology and School of Biological Sciences, Research Floor, St Mary's Hospital, Manchester M13 0JH, UK.
Aplin
J D
JD
D000911:Antibodies, Monoclonal / Q000378:metabolism; D002822:Choriocarcinoma / Q000737:chemistry*; D017077:Culture Media, Conditioned / Q000737:chemistry; D003656:Decidua / Q000737:chemistry; D005109:Extracellular Matrix / Q000737:chemistry; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D007797:Laminin / Q000737:chemistry*; D010920:Placenta / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000737:chemistry; D017421:Sequence Analysis; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D017077:Culture Media, Conditioned; D007797:Laminin; D012333:RNA, Messenger