Internalisation of short-chain isoprenyl diphosphates by chromaffin cells from bovine adrenal medulla

The uptake of FPP and GGPP was studied using primary cultures of chromaffin cells as an experimental model. The translocation of the isoprenyl diphosphates was shown to be both dose- and time-dependent. After incubation and a series of thorough washing cycles the residual radioactivity could only be released by either detergent or alkali treatment endorsing true internalisation rather than sorption onto the external cell surface. Preliminary experiments suggest that the isoprenyl diphosphates are internalised-at least for the greater part-without a preceeding dephosphorylation step. The translocation process does not seem to follow regular Michaelis-Menten kinetics. Upon varying pH (pH-range 6-8) or temperature (0 degree-40 degrees C) of the incubation medium only moderate changes in internalisation rate were observed. The metabolic poisons (CN-, F-, chloroquine) administered were without any significant effect, suggesting the transport phenomenon not to be energy dependent nor to proceed via receptor-mediated endocytosis. Upon inclusion of metallic ions in the test medium a pronounced increase in uptake was registered for Zn(2+)-ions, while most other cations did not cause any significant alterations. From the amino acid modifying agents screened sulphydryl reagents (NEM, PCMB, IA) resulted into a moderate unexpected elevation of uptake. An interesting feature was the activatory effect on the uptake of the diphosphorylated derivatives elicited by the corresponding free alcohols farnesol and geranylgeraniol; shorter and longer chain alcohols were without effect. In order to assess the specificity of the transport system a number of (pyro)phosphorylated compounds were included in the incubation medium indicating dolichylmonophosphate and isopentenylpyrophosphate to promote the uptake.