Characterization and growth-dependent regulation of the nerve growth factor receptor gp140trk in rat C6 glioma cells.
The glioma cell line C6 was used to study the expression and growth-dependent regulation of the nerve growth factor (NGF) tyrosine kinase receptor gp140trk, which is the mature protein product of the trk proto-oncogene. Chemical cross-linking of 125I-NGF to C6 cells, followed by immunoprecipitation with polyclonal anti-NGF antibodies and separation by polyacrylamide gel electrophoresis, revealed the presence of 90-95 and 150 kDa species. Immunocytochemical staining of C6 cells with antibodies directed against either the low-affinity NGF receptor gp75NGFR or trk proto-oncogene products demonstrated a heterogeneous cellular distribution of both antigens. Brief treatment of C6 cells with NGF led to the tyrosine phosphorylation of 80, 110 and 140 kDa protein species, as detected on anti-phosphotyrosine Western blots. Similar molecular weight species were found with anti-Trk antibodies in the NGF-treated cells. Intracellular localization of Trk-like immunoreactivity in C6 cells released from a growth-arrested state indicated an initial immunostaining of the nuclear periphery, progressing to cytoplasmic vesicles and finally to the plasma membrane. These observations at the light microscopic level were confirmed using immunoelectron microscopy with the same anti-Trk antibodies, and showed clearly the trafficking of Trk-like immunostained particles from the endoplasmic reticulum to the plasmalemma. The cellular localization of trk gene products also appeared to depend on their glycosylation state. Such growth-dependent expression of NGF receptors on glial cells may be important in controlling autocrine regulatory processes of glia to NGF, which these cells produce.
Zanellato
A
A
Fidia Research Laboratories, Fidia S.p.A., Abano Terme, Italy.
Facci
L
L
Petrelli
L
L
Dal Toso
R
R
Skaper
S D
SD
D000818:Animals; D002460:Cell Line; D005910:Glioma / Q000378:metabolism*; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron; D008970:Molecular Weight; D009414:Nerve Growth Factors / Q000494:pharmacology*; D010750:Phosphoproteins / Q000096:biosynthesis / Q000302:isolation & purification; D011518:Proto-Oncogene Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000378:metabolism; D011519:Proto-Oncogenes; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000032:analysis / Q000096:biosynthesis* / Q000378:metabolism; D020917:Receptor, trkA; D017475:Receptors, Nerve Growth Factor / Q000032:analysis / Q000096:biosynthesis* / Q000378:metabolism; D014407:Tumor Cells, Cultured
D009414:Nerve Growth Factors; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D017475:Receptors, Nerve Growth Factor; D020794:Receptor Protein-Tyrosine Kinases; D020917:Receptor, trkA