Properties of the carcinoma-associated antigen MH 99/KS 1/4 in normal and transformed human keratinocytes: regulation of synthesis, molecular cross-linking and ultrastructural localization.
A 38-kDa cell-surface glycoprotein defined by monoclonal antibody MH 99 is markedly increased in many epithelial tumours. In normal human skin, it is a characteristic marker for germ-cell phenotypic tissues. Although the gene encoding the MH 99 antigen has recently been cloned, and several histological and biochemical studies have been performed, the biological function of this interesting antigen still remains unknown. In the present study, we examined the synthesis of MH 99 in keratinocyte populations showing different in vitro differentiation capacity. Normal keratinocytes, spontaneously immortalized keratinocytes (cell line HaCaT), three SV-40-transformed keratinocyte lines (130, 425, and HaSV), and two squamous cell carcinoma lines (SCL-1 and SCL-2), were compared. Radioimmunoprecipitation revealed the highest levels of synthesis in cell populations with the least differentiation. This was paralleled by an increase of MH 99 synthesis in normal keratinocytes cultured in low concentrations of Ca2+ and by an increase of MH 99 synthesis during subculture of normal keratinocytes. Both phenomena were paralleled by an opposite behaviour of a differentiation marker. Molecular cross-linking and subsequent immunoprecipitation led to a decrease of the MH 99 signal, but an increase of a high molecular weight protein signal was seen. After cleavage of the crosslinker, the MH 99 signal reappeared, whereas the signal of the large protein remained unchanged. Thus, the MH 99 antigen may be associated with a high molecular weight protein on the cell surface, supporting the suggestion of a receptor-like function. Phosphorylation of the molecule could not be detected. Immunoelectron microscopy revealed homogeneous distribution on the cell surface, but cells of the same culture exhibited clear differences in their MH 99 expression. A concept for MH 99 regulation in normal and transformed human keratinocyte populations in vitro is proposed, showing that the synthesis of MH 99 is inversely correlated with cell differentiation. The association with a high molecular weight protein supports the suggestion that the MH 99 antigen interacts with other molecules.
Schön
M P
MP
Department of Dermatology, University Medical Center Steglitz, Free University of Berlin, Germany.
Schön
M
M
Klein
C E
CE
Kaufmann
R
R
Herzberg
F
F
Schmidt
R
R
Orfanos
C E
CE
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm / Q000378:metabolism*; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000276:immunology*; D002454:Cell Differentiation / Q000276:immunology; D002461:Cell Line, Transformed; D006801:Humans; D015603:Keratinocytes / Q000276:immunology*; D016253:Microscopy, Immunoelectron; D012878:Skin Neoplasms / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm