Regulation of Nerve Growth Factor Receptor Gene Expression by Nerve Growth Factor in the Developing Peripheral Nervous System

Nerve growth factor (NGF) is a targetderived neurotrophic protein that promotes the survival and growth of developing sympathetic and sensory neurons. We have examined NGF receptor gene expression in these neurons after NGF administration. Northern blot and in situ hybridization analyses demonstrated that NGF given systemically to neonatal rats increased levels of NGF receptor mRNA in sympathetic neurons within the superior cervical ganglion. This increase was accompanied by a differential regulation of genes associated with neurotransmitter phenotype; tyrosine hydroxylase mRNA was increased, but neuropeptide Y mRNA was not. NGF receptor mRNA levels were also increased in L4-L5 dorsal root ganglia, although this mRNA was not expressed uniformly in sensory neurons of control or NGF-treated animals. Levels of Tod ot-mbulin mRNA, a marker of neuronal growth, also increased. In contrast to developing neurons, systemic NGF did not increase NGF receptor mRNA in nonneuronal cells of the sciatic nerve. To determine if NGF regulated NGF receptor gene expression at the transcriptional level, we examined PC12 cells. NGF treatment for 6 h increased NGF receptor mRNA fourfold; this increase was inhibited by cycloheximide. Nuclear run-off transcription assays demonstrated that the increase in steady-state NGF receptor mRNA levels was mediated at the transcriptional level. In contrast, although NGF treatment increased steady-state tyrosine hydroxylase mRNA levels, this effect was not blocked by cycloheximide, and was not due to increased transcription. These data raise the possibility that transcriptional regulation of NGF receptor gene expression by targetderived NGF could be a molecular mechanism for potentiating NGF's effects on neurons during developmental periods of neuronal competition and cell death. I NTERACTIONS between a developing peripheral neuron and its target organ are believed to partially determine the phenotypic fate of that neuron, and to play an important role in neuronal competition and cell death. Nerve growth factor (NGF) t is a target-derived neurotrophic factor involved in the survival and differentiation of developing sympathetic and neural crest-derived sensory neurons. NGF given systemically to neonatal rats promotes growth of sympathetic neurons (Levi-Montalcini and Booker, 1960a), and affects the neurotransmitter phenotype of both sensory and sympathetic neurons (Kessler and Black, 1980; Otten et al., 1980; Thoenen et al., 1971). Conversely, antibodies to NGF lead to the death of embryonic sensory neurons (Johnson et al., 1980; Aloe et al., 1981), and of neonatal or mature sympathetic neurons (Levi-Montalcini and Booker, 1960b; Angeletti et al., 1971; Gorin and Johnson, 1980). NGF synthesis in the target field of sympathetic neurons commences around the time of axonal contact (Davies et al., 1987). Together, these studies suggest that NGF plays an important role in regulating neuronal survival and differentiation. NGF mediates its actions by binding to the high-affinity 1. Abbreviations used in thispaper: DRG, dorsal root ganglia; NGF, nerve growth factor; SCG, superior cervical ganglia. form of the membrane-bound NGF receptor (Green et al., 1986). The low-affinity form of the NGF receptor (Sutter et al., 1979), which has been cloned (Johnson et al., 1986; Radeke et al., 1987), is believed to provide an essential component of the high-affinity receptor (Hosang and Shooter, 1985; Green and Greene, 1986), and is capable, when expressed in mutant PC12 cells, of restoring functional responses to NGF (Hempstead et al., 1989). Thus, the same gene product is believed to encode components of both the highand low-affinity binding sites, as well as a truncated form of the receptor (DiStefano and Johnson, 1988b). NGF receptor mRNA is expressed in both neural and nonneural tissues during the development of rodents and chickens (Ernfors et al., 1988; Large et al., 1989). Wyatt et al. (1990) recently demonstrated that the amount of NGF receptor on developing trigeminal neurons increased at approximately the same time as initial target contact. One explanation for this observation is that NGF may directly increase expression of the NGF receptor gene, a hypothesis supported by studies demonstrating that NGF administered in the cerebrospinal fluid increased NGF receptor mRNA in basal forebrain cholinergic neurons (Higgins et al., 1989; Cavicchioli et al., 1989), and that NGF increased receptor mRNA in cultures of adult sensory neurons (Lindsay et al., © The Rockefeller University Press, 0021-9525/91/01/303/10 $2.00 The Journal of Cell Biology, Volume 112, Number 2, January 1991 303-312 303 on O cber 8, 2017 jcb.rress.org D ow nladed fom 1990). Regulation of the number and density of NGF receptors on the surface of peripheral neurons by NGF in vivo could be a positive feedback mechanism that contributes to neuronal differentiation and survival. In this study, we tested whether exogenous, systemic NGF regulates the levels of NGF receptor mRNA in developing peripheral neurons during the period of neuronal competition and cell death. Results demonstrate that systemic NGF increased levels of NGF receptor mRNA in neonatal sympathetic and sensory neurons, but not in developing nonneuronal cells of sciatic nerve that also express NGF receptor mRNA. In PC12 cells, the NGF-mediated increases in NGF receptor mRNA occur at the transcriptional level with characteristics that implicate an immediate early gene product. This increase in NGF receptor gene expression was accompanied by increased expression of tyrosine hydroxylase and Totl ~tubulin mRNAs in sympathetic and sensory neurons, respectively. These data indicate that NGF increases transcription of the NGF receptor gene in developing peripheral neurons, and that this increase is coincident with other NGF-mediated changes in neuronal gene expression. Subsequent increases in levels of the high-affinity NGF receptor would provide a cellular mechanism for potentiating the effects of NGF on NGF-responsive neurons, and may indicate a role for target-derived NGF in neuronal competition