SNT, a differentiation-specific target of neurotrophic factor-induced tyrosine kinase activity in neurons and PC12 cells.

Abstract

To elucidate the signal transduction mechanisms used by ligands that induce differentiation and the cessation of cell division, we utilized p13 uc1agarose, a reagent that binds p34C2/cdk2 By using thi re t identified a 78-to 90-kDa species in PC12 pheochromocytoma cells that is rapidly phosphorylated on tyrosine following treatment with the differentiation factors nerve growth factor (NGF) and fibroblast growth factor but not by the mitogens epidermal growth factor or insulin. This species, called SNT (suc-associated neurotrophic factor-induced tyrosine-phosphorylated target), was also phosphorylated on tyrosine in primary rat cortical neurons treated with the neurotrophic factors neurotrophin-3, brain-derived neurotrophic factor, and fibroblast growth factor but not in those treated with epidermal growth factor. In neuronal and fibroblast cells, where NGF can also act as a mitogen, SNT was tyrosine phosphorylated to a much greater extent during NGF-induced differentiation than during NGF-induced proliferation. SNT was phosphorylated in vitro on serine, threonine, and tyrosine in pl3sucl-agarose precipitates from NGF-treated PC12 cells, indicating that this protein may be a substrate of kinase activities associated with p13suc1-p34cdc21cdk2 complexes. In addition, SNT was associated predominantly with nuclear fractions following subcellular fractionation of NGF-treated PC12 cells. Finally, in PC12 cells, NGF-stimulated tyrosine phosphorylation of SNT was dependent on the levels of Trk tyrosine kinase activity and was constitutively induced by expression of pp6Ov<n?. However, Ras was not required for constitutive SNT tyrosine phosphorylation, suggesting that this protein functions distally to Trk and pp6OV but in a pathway parallel to that of Ras. SNT is the first identified specific target of differentiation factor-induced tyrosine kinase activity in neuronal cells.Survival and differentiation of neurons in the peripheral and central nervous systems is regulated by neurotrophic factors. These factors include the neurotrophin family, fibroblast growth factors (FGFs), and the interleukin-like factor ciliary neurotrophic factor (1,27,68,72). The neurotrophin family of factors includes nerve growth factor (NGF), brainderived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and 27,68). Neurotrophins promote the differentiation and survival of overlapping populations of neurons in culture and show distinct temporal and spatial expression patterns (30,33,43,46,48,51). Neurotrophins bind to the Trk family of receptor tyrosine kinases, which includes Trk, TrkB, and TrkC. NGF binds to Trk; BDNF, NT-3, and NT-4/5 bind to TrkB; and NT-3 binds to TrkC. NT-3 and NT-4/5 can also interact weakly with Trk (3,34,36,39,40,45,64,65).The best-characterized neurotrophic factor is NGF. NGF is required for differentiation and survival of sympathetic and some sensory neurons in the peripheral nervous system and for the cholinergic neurons of the basal forebrain (1, 59). NGF also promotes differentiation of the rat pheochromocytoma tum...

Nerve growth factor induced stimulation of Ras requires Trk interaction with Shc but does not involve phosphoinositide 3-OH kinase

“…Recently, a new docking protein has been characterized (Kouhara et al, 1997), FRS2, that is phosphorylated on tyrosine residues in response to FGF or NGF stimulation. FRS2 is thought to be closely related, but probably not identical to, SNT (Rabin et al, 1993). Upon NGF stimulation of Trk in PC12 cells, FRS2 becomes tyrosine phosphorylated and binds to Grb2/Sos.…”

FGF-receptor substrate 2 functions as a molecular sensor integrating external regulatory signals into the FGF pathway

“…In this report, we examined whether the adapter protein FRS2 is a potential substrate of crosstalk between the FGF pathway and other signaling pathways. As described previously, FRS2 undergoes strong PS/T in response to several extracellular stimuli [1,15,28]; however, a plausible explanation for the function of this phosphorylation was not reported. Recently, a negative feedback loop comprising FRS2 and MAPK/ERK has been described.…”

Functional Rac-1 and Nck signaling networks are required for FGF-2-induced DNA synthesis in MCF-7 cells

“…The second molecule involved in mediating Ras activation by FGFRs is SNT (Rabin et al, 1993), named also SLP (Wang et al, 1996) or more recently FRS-2 (Kouhara et al, 1997). As shown in Figure 3b (upper panel), FRS-2 tyrosine phosphorylation increased twofold after 5 min and sixfold after 60 min of FGF-2 stimulation in MCF-7 cells.…”