Differential effect of P75 coexpression on TrkA and TrkB response to ligands in neuroblastoma

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Neuroblastoma is a common childhood solid tumor derived from the neural crest and frequently arises in sympathetic ganglia or the adrenal medulla. TrkA, TrkB and TrkC are the receptors for nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), respectively. Favorable neuroblastomas usually express TrkA, whereas unfavorable, MYCN-amplified neuroblastomas usually express TrkB and BDNF. P75 (p75NTR, NGFR, TNFRSF16) binds all NGF-related neurotrophins with low affinity. One important function of p75 is to enhance the response of TrkA to NGF. Here we studied the consequences of p75 coexpression on TrkA and TrkB function in neuroblastoma cell lines. We stably transfected a p75 expression vector into our existing neuroblastoma transfectants: SY5Y cell line containing TrkA (p23A) and TrkB (BR6). Clones expressing TrkA (or TrkB) with or without p75 were exposed to different concentrations (0.1 to 100 ng/ml) of their specific ligand (NGF for TrkA, and BDNF for TrkB). Phosphorylation of TrkA or TrkB was studied by Western blotting, and increased cell number was assessed by MTT assay. In TrkA transfected cells, p75 coexpression increased TrkA phosphorylation by about 2 fold at higher concentrations of ligand, whereas basal TrkA phosphorylation in the absence of ligand showed only a modest increase with p75 coexpression. Furthermore, p75 did not alter cell proliferation in the absence of ligand. In TrkB transfected cells, TrkB phosphorylation in response to exogenous BDNF (1 to100 ng/ml) was similar with or without p75. However, p75 coexpression significantly increased basal TrkB phosphorylation at lower ligand concentrations (0 to 0.1 ng/ml). Phosphorylation of the downstream signaling intermediates, AKT and MAPK, exhibited a similar pattern with or without p75. In contrast to TrkA, p75 coexpression increased the proliferation of TrkB cells. At day 3 and day 5, there was a 2-fold increase in cell number for p75 expressing cells without ligand exposure (p<0.05). However, at very high concentrations of BDNF, cell number was similar with or without p75. In summary, p75 increases the maximal response to NGF at higher ligand concentrations in TrkA cells, whereas in TrkB cells, p75 increases basal phosphorylation and proliferation under lower concentrations of ligand. Thus, p75 plays alters the response of both TrkA and TrkB to ligand, but different ways.