Ligand-induced activation of epidermal growth factor receptor in intact rat mammary adenocarcinoma cells without detectable receptor phosphorylation.
Expression and function of epidermal growth factor receptor (EGFR) was investigated in a metastatic cell clone (MTLn3) derived from the 13762NF rat mammary adenocarcinoma. No receptor phosphorylation could be identified in intact cells or in membrane preparations, while EGF-dependent phosphorylation of substrates occurred in intact cells. Indications for active suppression of receptor phosphorylation came from the fact that EGFRs bound in immunocomplexes or associated with the cytoskeleton of detergent treated cells were able to undergo basal and EGF-induced phosphorylation in vitro. Cross-linking experiments with 125I-EGF, as well as [35S]methionine labeling followed by immunoprecipitation with receptor specific antibodies readily detected in MTLn3 cells the expected 170-kDa EGFR protein. In addition, two proteins with molecular masses of 420-480 and 95 kDa specifically bound 125I-EGF on intact MTLn3 and sparse cultures of A431 cells. Phosphorylation of the 420-480 kDa molecule could be identified in immunocomplexes of EGFRs isolated from MTLn3 and sparse A431 cells, but the 95-kDa receptor molecule was never phosphorylated. While the presence of alternative forms of EGFR in the highly metastatic cell clone MTLn3 was unexpected, our observations of inefficient receptor autophosphorylation are in agreement with other recent reports and suggest that in MTLn3 cells EGFR-mediated signal transduction can be an event independent from receptor autophosphorylation.
Lichtner
R B
RB
Department of Immunology and Genetics, German Cancer Research Center, Heidelberg.
Wiedemuth
M
M
Kittmann
A
A
Ullrich
A
A
Schirrmacher
V
V
Khazaie
K
K
D000230:Adenocarcinoma / Q000378:metabolism*; D000818:Animals; D001345:Autoradiography; D015152:Blotting, Northern; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D066246:ErbB Receptors / Q000378:metabolism*; D008024:Ligands; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D012334:RNA, Neoplasm / Q000378:metabolism; D051381:Rats; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism
D003432:Cross-Linking Reagents; D008024:Ligands; D012334:RNA, Neoplasm; D014443:Tyrosine; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases