BREFELDIN-A INHIBITS PHOSPHATE-TRANSPORT IN OPOSSUM KIDNEY-CELLS

Brefeldin A (BFA) is a fungal metabolite that blocks the transport processes between the endoplasmic reticulum and the Golgi apparatus. In the present study, we have tested the effect of BFA on phosphate transport in a kidney epithelial cell line, opossum kidney (OK) cells. Electron microscopy showed that exposure of OK cells to BFA caused a rapid and reversible disorganization of Golgi apparatus. Addition of BFA also caused a time (2-8 h)- and dose (1-10 mug/ml)-dependent inhibition of Na+-dependent cell phosphate uptake. The inhibition of cell phosphate uptake by BFA was reversible and was associated with a decrease in the maximum velocity of phosphate transport. Both the inhibition and the stimulation of cell phosphate uptake by parathyroid hormone and insulin, respectively, were not affected by BFA. BFA at 1 mug/ml concentration did not affect protein synthesis as determined by [H-3]leucine incorporation but diminished the adaptive increase in cell phosphate uptake in response to 2 or 8 h of incubation in nominally phosphate-free medium. On the other hand, inhibition of protein synthesis by cycloheximide (5 muM) abolished the adaptive increase in cell phosphate uptake in response to 8 but not 2 h of incubation in nominally phosphate-free medium, indicating the existence of an early response to phosphate deprivation, which does not require new protein synthesis but is sensitive to the effect of BFA. In summary, results of these studies show that, in OK cells, BFA inhibits phosphate uptake and curtails the adaptive response to phosphate deprivation. These results suggest the participation of protein transport processes out of the Golgi apparatus in the maintenance of phosphate uptake and the response to phosphate deprivation.