Regulation of the high-affinity H+/peptide cotransporter in renal LLC-PK1 cells.
Di- and tripeptides and peptide mimetics such as beta-lactam antibiotics are efficiently reabsorbed from the tubular lumen by a high-affinity peptide transporter. We have recently identified and characterized this H+-coupled high-affinity peptide transport system in the porcine proximal tubular cell line LLC-PK1. Here we describe for the first time the regulation of the renal high-affinity peptide cotransporter at the cellular level. Uptake of 5 microM 3H-D-Phe-L-Ala into LLC-PK1 cells was significantly increased by lowering [Ca2+]in and decreased by increasing [Ca2+] in. Moreover, it was shown that the [Ca2+]in effects on peptide transport activity were dependent on Ca2+ entry from the extracellular site (e.g., via a store-regulated capacitative Ca2+ influx). Protein kinase C (PKC) was found to transmit the effects of [Ca2+]in on peptide transport. Although we demonstrate by pHin measurements that the PKC inhibitor staurosporine did decrease the transmembrane H+ gradient and consequently should have reduced the driving force for peptide uptake, the only effect on transport kinetics of 3H-D-Phe-L-Ala observed was a significant decrease in Km from 22.7+/-2.5 microM to 10.2+/-1.9 microM with no change in maximal velocity.
Wenzel
U
U
Institute of Nutritional Sciences, University of Giessen, Germany.
Diehl
D
D
Herget
M
M
Kuntz
S
S
Daniel
H
H
D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002470:Cell Survival; D004151:Dipeptides / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D007700:Kinetics; D018374:LLC-PK1 Cells; D011493:Protein Kinase C / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D019311:Staurosporine / Q000494:pharmacology; D013237:Stereoisomerism; D013449:Sulfonamides / Q000494:pharmacology; D013552:Swine; D027981:Symporters; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014268:Trifluoperazine / Q000494:pharmacology