Hepatocyte growth factor-induced ectodomain shedding of cell adhesion molecule L1: role of the L1 cytoplasmic domain.
The L1 cell adhesion molecule and its soluble form are tumor-associated proteins and potential markers for tumor staging as well as targets for therapeutic intervention. Soluble L1 is produced by metalloprotease-mediated ectodomain shedding of L1. We investigated effects of hepatocyte growth factor (HGF), a growth factor shown to increase invasiveness of renal carcinoma cells, on ectodomain shedding of L1 from these cells. All of the tested L1-positive renal carcinoma cell lines released a 180-kDa form of L1 into the medium. In the presence of serum, addition of HGF led to a dose-dependent increase in L1 shedding with a maximum reached at 5 ng/ml. In contrast, L1 shedding was inhibited by glial cell line-derived neurotrophic factor (GDNF). The tyrosine kinase inhibitor Genistein reduced basal and HGF-stimulated L1 shedding, indicating that protein phosphorylation is involved. To investigate the role of the L1 intracellular domain, two mutants of the L1 cytoplasmic part were constructed. L1trun lacking the complete intracellular domain showed enhanced basal shedding. In a L1YH mutant, containing the mutation tyrosine 1229 to histidine that deletes the ankyrin binding motif of L1, basal shedding was reduced. Disruption of actin assembly by cytochalasin D also reduced shedding of L1. These results indicate that the cytoplasmic domain regulates basal shedding of L1, and association with the cytoskeleton through the L1 ankyrin binding site is involved. HGF stimulated L1 shedding in both mutants, indicating that receptor-mediated phosphorylation in the L1 cytoplasmic domain is not required for HGF-stimulated shedding.
Heiz
Monika
M
Center for Radiopharmaceutical Science ETH-PSI-USZ; Paul Scherrer Institute, CH-5232 Villigen, Switzerland.
Grünberg
Jürgen
J
Schubiger
P August
PA
Novak-Hofer
Ilse
I
D002292:Carcinoma, Renal Cell / Q000378:metabolism; D045744:Cell Line, Tumor; D019833:Genistein / Q000494:pharmacology; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D007680:Kidney Neoplasms / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000737:chemistry / Q000378:metabolism; D009414:Nerve Growth Factors / Q000494:pharmacology; D039842:Neural Cell Adhesion Molecule L1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019859:Proto-Oncogene Proteins c-met / Q000378:metabolism; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
C495039:GDNF protein, human; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D009363:Neoplasm Proteins; D009414:Nerve Growth Factors; D039842:Neural Cell Adhesion Molecule L1; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D017228:Hepatocyte Growth Factor; D019833:Genistein; D019859:Proto-Oncogene Proteins c-met; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases