The Na+/Pi-cotransporter of OK cells: reaction and tentative identification with N-acetylimidazole

Abstract

Using an established renal epithelial cell line (OK cells) the effect of the amino-acid side-chain modifying reagent N-acetylimidazole (NAI) upon the sodium-dependent transport of phosphate (Pi) was investigated. After an incubation with 10 mM NAI for 20 min, cellular Na+/Pi uptake was inhibited by 70%. The presence of 5 mM Pi protected this transport function from being affected by NAI by 80 to 100%. Since the presence of sulfate was unable to protect the Na+/Pi transport inactivation by NAI and since the presence of Pi did not affect NAI inhibition of other transport systems, it is suggested that NAI interacts with the Pi transporter directly. The protective effect of Pi was used as a criterion to identify Pi-protectable [3H]NAI labelling of OK cell plasma membrane proteins. Pi protection was observed in four molecular mass regions: 31, 53, 104 and 176 kDa. Since the incorporation of [3H]NAI into these proteins was also affected by parathyroid hormone at 10(-10) M, it is concluded that the identified proteins represent possible candidates for the renal Na+/Pi cotransporter.