of Anatomy and B/o/ogy, Ph//ade/ph/a, PennAylvdnrd 79 104 '151-nerve growth factor (NCF) was found to be internalized and translocated to the nucleus of SKBr5 breast carcinoma celis. The cytoplasm and chromatin isolated from nonmitotic cells accumulated two-and five-fold, respectively, inorc of I2'I-NGF than the cells undergoing mitosis. MAb 20.4 devclopcd against the NGF cell surface receptor immunoprecipitated the 80,000 M, receptor from plasma membrane and two protein species from the chromatin; 90,000 M, (major band) and 200,000 M, (minor band). In SKBr5 cells, binding of NCF to the chromatin did not affect synthesis of rRNA. Proliferation ofSKBr5 cells was slightly stimulated by NGF. In control melanoma A875 cells, which express the 230,000 M, chromatin receptor, NGF inhibited both rRNA synthesis and cell proliferation. We suggest that the 90,000 M, chromatin receptor expressed by SKBr5 cells repments a "nonactive," ligand-binding subunit of the high molecular wright receptor for NGF. The critical role of the chromatin receptor for NGF in rRNA-dependent cell proliferation is discussed. a 1 w W I I ~~-L I S ~, Inr.
Nerve growth factor (NGF) belongs to the family of neurotropins. Three groups of cells are targets for NGF: neural crest derivatives, central nervous system, and cells of nonneural origin, like mast cells. NGF exerts different effects in different cell types. NGF is essential for the survival and differentiation of sympathetic and sensory neurons, but not for their proliferation (Barde, 1989;Levi-Montalcini, 1987). In adrenal chromaffin cells, NGF stimulates mitotic response (Lillien and Claude, 1985), whereas in the rat adrenal pheochromocytoma cells, NGF inhibits proliferation and induces differentiation (Rudkin et al., 1989). The NGF receptor is also expressed by several human tumor cell lines. Two types of the cell surface (plasma membrane) receptors for NGF are known: the low molecular weight (M, 75,000) receptor, which is well characterized and cloned (Chao et al., 1986;Johnson et al., 1986); and the more recently identified, high molecular weight (M, 140,000) receptor (Kaplan et al., 1991;Klein et al., 1991; Nebreda et al., 1991). The 140,000 M, receptor represents a product of the trh protooncogene (p140tr'). It is believed that each of the receptors binds NGF with low affinity, whereas the heterodimer of both receptors binds NGF with high affinity (Hempstead et al., 1991). Klein et al. (1991) found that p140trh alone may also generate high affinity receptors. Most investigators suggest that only the high affinity receptors mediate NGF function. We have extensively tested several melanoma cell lines that express only the low molecular weight (M, 75,000), low affinity receptors (Chao et al., 1986) and found that melanoma cells internalize NGF and respond to NGF by inhibiting growth (Rakowicz-Szulczynska et al., 1988, 1989;Rakowicz-Szulczynska and Koprowski, 1989). The mechanism of melanoma cell growth inhibition by NGF involves intracellular
Methods
Cell lines SKBr5 breast carcinoma and WM164 and A875 melanoma cell lines were grown in minimum Eagle's mediumill5 (3:l) supplemented with 10% bovine se-