K-252b potentiation of neurotrophin-3 is trkA specific in cells lacking p75NTR.
K-252b potentiates the neurotrophic effects of neurotrophin-3 (NT-3) in primary cultures of rat central cholinergic and peripheral sensory neurons and in a rat pheochromocytoma PC12 cell line. The ligand and receptor specificity, and role of the low-affinity neurotrophin receptor (p75NTR) in the potentiation response induced by K-252b, are unknown. To address the issues of ligand and receptor specificity of K-252b potentiation, we have examined neurotrophin-induced DNA synthesis ([3H]-thymidine incorporation) in NIH3T3 cells expressing trkA, trkB, or trkC. Neither NT-3 nor K-252b alone could stimulate mitogenic activity in the trkA-overexpressing clone. However, coaddition of K-252b (EC50 of approximately 2 nM) with 10-100 ng/ml NT-3 led to incorporation of [3H]thymidine in trkA expressing cells to a level induced by optimal concentrations of nerve growth factor (NGF). The K-252b- and NT-3-induced [3H]thymidine incorporation correlated with an increase in the tyrosine autophosphorylation of the trkA receptor as well as tyrosine phosphorylation of trk-associated phospholipase C-gamma 1 and SH2-containing proteins. K-252b did not potentiate submaximal doses of NGF, or maximal doses of brain-derived neurotrophic factor (BDNF) or neurotrophin-4/5 (NT-4/ 5) in trkA-expressing cells. Furthermore, K-252b did not potentiate DNA synthesis by submaximal doses of BDNF, NT-4/5, or NT-3 in trkB- or trkC-expressing NIH3T3 cells, suggesting that the potentiation profile for K-252b was specific for NT-3 in trkA-expressing cells. We found no expression of p75NTR in the trk-expressing NIH3T3 cells. This is the first demonstration that K-252b potentiates a trkA-mediated biological nonneuronal response by NT-3 that occurs independent of p75NTR and appears to be both ligand and receptor specific.
Maroney
A C
AC
Cephalon Inc., West Chester, Pennsylvania 19380, USA.
Sanders
C
C
Neff
N T
NT
Dionne
C A
CA
D016475:3T3 Cells; D000818:Animals; D002227:Carbazoles / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D026121:Indole Alkaloids; D008024:Ligands; D051379:Mice; D008970:Molecular Weight; D009414:Nerve Growth Factors / Q000494:pharmacology*; D020933:Neurotrophin 3; D010766:Phosphorylation; D051381:Rats; D020917:Receptor, trkA / Q000235:genetics / Q000378:metabolism*; D017475:Receptors, Nerve Growth Factor / Q000378:metabolism* / Q000502:physiology; D013379:Substrate Specificity; D014162:Transfection; D014407:Tumor Cells, Cultured
D002227:Carbazoles; D004791:Enzyme Inhibitors; D026121:Indole Alkaloids; D008024:Ligands; D009414:Nerve Growth Factors; D020933:Neurotrophin 3; D017475:Receptors, Nerve Growth Factor; C049985:staurosporine aglycone; D020917:Receptor, trkA