BEWO CHORIOCARCINOMA CELLS PRODUCE LAMININ-10

BeWo is a choriocarcinoma cell line that generates an extracellular ma trix (ECM) rich in laminin and is a useful model for human trophoblast . Immunofluorescence with monoclonal antibodies demonstrates that BeWo ECM contains laminin subunits beta 1 and gamma 1. Immunoprecipitation from conditioned medium shows that the cells secrete two distinct lam inin trimers both containing beta 1 and gamma 1 but with alpha subunit s of approx. 400 and 450 kDa. The culture medium also contains a speci es thought to be beta 1 gamma 1 dimer. Immunoprecipitation with monocl onal antibody 4C7, previously thought to recognize the alpha 1 subunit , isolates complexes containing only the smaller alpha subunit. A seco nd complex containing the larger alpha subunit along with beta 1, gamm a 1 and a 150 kDa polypeptide is precipitated from 4C7-depleted medium with an anti-(laminin 1) polyclonal antibody. Peptide sequencing demo nstrates that the 4C7-reactive species is alpha 5, which is present as two similarly sized polypeptides. mRNA species encoding laminin subun its alpha 1, alpha 5, beta 1, beta 2 and gamma 1 are all present in th e cells. These results demonstrate the secretion of a novel laminin is oform, laminin 10, the subunit composition of which is alpha 5 beta 1 gamma 1. Laminin 1 is also produced. No evidence for the secretion of beta(2)-containing laminin isoforms could be derived despite the prese nce of beta 2 mRNA. Analysis with reverse transcriptase-mediated PCR a lso showed the presence of laminin alpha 5 in first-trimester placenta and decidua.