Distinct and Different Effects of the Oncogenes

Immature avian sympathetic neurons are able to proliferate in culture for a limited number of divisions albeit expressing several neuron-specific properties. The effect of avian retroviral transfer of oncogenes on proliferation and differentiation of sympathetic neurons was investigated. Primary cultures of 6-d-old quail sympathetic ganglia, consisting of 90% neuronal cells, were infected by Myelocytomatosis virus (MC29), which contains the oncogene v-myc, and by the v-src-containing Rous sarcoma virus (RSV). RSV infection, in contrast to findings in other cellular systems, resulted in a reduction of neuronal proliferation as determined by 3H-thymidine incorporation (50% of control 4 d after infection) and in increased morphological differentiation. This is reflected by increased neurite production, cell size, and expression of neurofilament protein. In addition, RSV-infected neurons, unlike uninfected cells, are able to survive in culture for time periods up to 14 d in the absence of added neurotrophic factors. In contrast, retroviral transfer of v-myc stimulated the proliferation of immature sympathetic neurons preserving many properties of uninfected cells. The neuron-specific cell surface antigen Q211 and the adrenergic marker enzyme tyrosine hydroxylase were maintained in MC29-infected cells and in the presence of chick embryo extract the cells could be propagated over several weeks and five passages. Within 7 d after infection, the number of Q211-positive neurons increased ~100-fold. These data demonstrate distinct and different effects of v-src and v-myc-containing retroviruses on proliferation and differentiation of sympathetic neurons: v-src transfer results in increased differentiation, whereas v-myc transfer maintains an immature status reflected by proliferation, immature morphology, and complex growth requirements. The possibility of expanding immature neuronal populations by transfer of v-myc will be of considerable importance for the molecular analysis of neuronal proliferation and differentiation. I N higher vertebrates the neurons of the central and peripheral nervous system are generated during embryonic or early postnatal days. During this period of neuronal birth, which extends over a period of several days for a specific neuronal cell type, a defined number of neurons is produced by proliferation and differentiation of neuronal precursor cells. The final number of neurons results from the process of selective survival which leads to the degeneration and death of large numbers of postmitotic neurons in many regions of the peripheral and central nervous system. The analysis of the control of neuronal proliferation during these early stages has been hampered by the lack of available tissue culture systems to investigate this process, and by the lack of markers to identify precursor cells, present only transiently and in limited numbers during development. We have recently demonstrated that neurons from 7-d-old (E7) ~ chick paravertebral sympathetic ganglia, identified by several neuronal markers, are able to proliferate in vitro for a limited number of divisions, and thus represent a suitable model to investigate proliferation and differentiation (Rohrer and Thoenen, 1987; Ernsberger et al., 1989a,b). Cells with similar properties have also been described in cultures of embryonic rat adrenal medulla (Anderson and Axel, 1986) and of embryonic rat sympathetic ganglia (DiCocco-Bloom and Black, 1988). We observed, however, that these cells cease to proliferate and start to degenerate after several days 1. Abbreviations used in this paper: CEE, chicken embryo extract; CEE chicken embryo fibroblast; CNTE ciliary neurotrophic factor; E, embryonic day; MC29, Myelocytomatosis virus; NF, neurofilament; NF-IR, NF immunoreactivity; NGF, nerve growth factor; RSV, Rous sarcoma virus; TH, tyrosine hydroxylase; TH-IR, TH immnnoreactivity. © The Rockefeller University Press, 0021-9525/90/06/2087/12 $2.00 The Journal of Cell Biology, Volume 110, June 199