Mutation of the human neu protein facilitates down-modulation by monoclonal antibodies.
Amplification and overexpression of the neu gene have been found in several human adenocarcinomas. We have obtained monoclonal antibodies to the human neu protein by immunizing a Balb/c mouse with a Balb/c cell line expressing the human neu gene by transfection. The monoclonal antibodies reacted with neu protein on intact cells by immunofluorescence and immunoprecipitated neu in metabolically labeled cells, also in the presence of tunicamycin. We tested possible down-modulating effects of these monoclonal antibodies on SKBR-3 mammary tumor cells, which express high levels of wild-type human neu protein. We also used NIH3T3 cells transfected with either a normal or a mutated human neu gene, encoding a protein with a valine to glutamic acid substitution in the transmembrane domain. Down-modulation of the normal cell-surface neu protein was inefficient. In contrast, the antibodies induced 50-65% down-modulation in NIH3T3 cells expressing the mutated human neu protein and could inhibit these cells to form colonies in soft agar. We propose that these differences are due to changed aggregation properties of the point-mutated protein.
van Leeuwen
F
F
Division of Molecular Biology, Netherlands Cancer Institute, Amsterdam.
van de Vijver
M J
MJ
Lomans
J
J
van Deemter
L
L
Jenster
G
G
Akiyama
T
T
Yamamoto
T
T
Nusse
R
R
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002448:Cell Adhesion; D002455:Cell Division; D002460:Cell Line; D002478:Cells, Cultured; D002999:Clone Cells; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008325:Mammary Neoplasms, Experimental; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D009154:Mutation; D010957:Plasmids; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000276:immunology; D011519:Proto-Oncogenes; D018719:Erb-b2 Receptor Tyrosine Kinases
D000911:Antibodies, Monoclonal; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases