Merosin is synthesized by thyroid cells in primary culture irrespective of cellular organization.
The in vitro synthesis and deposition of laminin family glycoproteins were studied using primary porcine thyroid cells cultured as monolayers or in follicles. The latter organization mimics the in vivo state of these polarized epithelial cells. In both cell systems a trimeric molecule was immunoprecipitated by using polyclonal antibodies against EHS-laminin. When the cells were fully polarized the protein was found at the basal pole of cells, irrespective of their organization. However, this molecule was different from laminin purified from a traditional source, the murine Engelbreth-Holm-Swarm (EHS) tumor. Thyroid cell laminin was composed of two light chains, analogous to EHS B1 and B2, and a disulfide-bonded heavy chain not found in EHS-laminin. The heavy chain was first synthesized as a 380 kDa polypeptide, then rapidly cleaved to a doublet of 350-380 kDa, which was subsequently found in both cell extracts and conditioned culture media. This thyroid laminin variant was compared with merosin, another variant found in the basement membranes of trophoblast, Schwann cells, striated muscle and liver. The heavy chain (M) of merosin shows homology to EHS-laminin heavy chain at the C-terminal domain, and is usually found as two polypeptides of 80 kDa and 300 kDa (Ehrig K., Leivo I., Argraves W. S., Ruoslahti E. and Engvall E. Proc. Nat. Acad. Sci. 87, 3264-3268, 1990). mRNA of the M chain was identified by RT-PCR in freshly isolated thyrocytes as well as in 6-day-old cultured thyroid cells. Furthermore, both the classical laminin heavy chain and the 350 kDa variant were detected by immunoblotting and immunofluorescence in the thyroid gland in vivo. All these results suggest strongly that merosin is a basement membrane component of thyroid cells in vivo and in vitro.
Andre
F
F
INSERM U 270, Faculté de Médecine Secteur Nord, Marseille, France.
Filippi
P
P
Feracci
H
H
D000818:Animals; D001345:Autoradiography; D001483:Base Sequence; D015139:Blotting, Southern; D002250:Carbon Radioisotopes; D002460:Cell Line; D002478:Cells, Cultured; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D007150:Immunohistochemistry; D007797:Laminin / Q000096:biosynthesis* / Q000302:isolation & purification; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012313:RNA / Q000302:isolation & purification / Q000378:metabolism; D013552:Swine; D013961:Thyroid Gland / Q000166:cytology* / Q000378:metabolism* / Q000648:ultrastructure
D002250:Carbon Radioisotopes; D017931:DNA Primers; D007797:Laminin; D046911:Macromolecular Substances; D012313:RNA