Abstract
Cells of a continuous line of rat pheochromocytoma (PC12) were incubated with 125I-labeled beta nerve growth factor (beta NGF), and at given time intervals the cell nuclei were isolated by a procedure that used the detergent Triton X-100. NGF was detectable in the nucleus after 20 min and continued to accumulate in a linear fashion for several hours after the total binding to the cell had reached steady state. After 17 hr at 37 degrees C, about 60% of the NGF bound to the cell was in the nucleus, NGF was not translocated to the nucleus at 4 degrees C. When nuclei were purified from PC12 cells and incubated with 125I-labeled beta NGF, specific binding sites were found. Binding was saturable and consistent with two sites: a high-affinity site with a Kd of 0.08 nM (+/- nM) and a lower-affinity site with a Kd of 9.0 nM (+/- 2.0 nM). The receptors in the nucleus were shown to be localized to the nuclear membrane. Membrane-free chromatin did not bind NGF specifically. The translocation of NGF to the nucleus was accompanied by a commensurate decrease in the cell-surface binding capacity. In the nucleus, however, the receptor capacities of both sites were increased when PC12 cells were grown in the presence of NGF.