Leupeptin and ammonium chloride inhibit intrinsic factor mediated transcytosis of [57Co]cobalamin across polarized renal epithelial cells.
The [125I] intrinsic factor (IF) mediated transcytosis of [57Co]Cyanocobalamin (Cbl) by polarized opossum kidney cells was inhibited (greater than 80%) by preincubation of the cells with lysosomotropic agents leupeptin or ammonium chloride. Inhibition of Cbl transcytosis resulted in the intracellular accumulation of both [125I]IF (48 kDa) and [57Co]Cbl. Intracellular degradation of [125I]IF occurred during normal cellular transcytosis of [57Co]Cbl and in one h following internalization the major intracellular degradation products of IF were two polypeptides of Mr 29 kDa and 19 kDa. The size of the major degradation product of IF in the basolateral media was 10 kDa. Based on these results, we suggest that IF is internalized by the renal epithelial cells and is degraded by leupeptin-sensitive acid proteases during Cbl transcytosis.
Ramanujam
K S
KS
Department of Medicine, Medical College of Wisconsin, Milwaukee 53226.
Seetharam
S
S
Seetharam
B
B
D000643:Ammonium Chloride / Q000494:pharmacology*; D000818:Animals; D001345:Autoradiography; D001692:Biological Transport / Q000187:drug effects; D002478:Cells, Cultured; D002850:Chromatography, Gel; D003037:Cobalt Radioisotopes; D004591:Electrophoresis, Polyacrylamide Gel; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D007437:Intrinsic Factor / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D007457:Iodine Radioisotopes; D007668:Kidney / Q000378:metabolism*; D007976:Leupeptins / Q000494:pharmacology*; D008970:Molecular Weight; D009893:Opossums; D051381:Rats; D014805:Vitamin B 12 / Q000378:metabolism*
D003037:Cobalt Radioisotopes; D007457:Iodine Radioisotopes; D007976:Leupeptins; D000643:Ammonium Chloride; D007437:Intrinsic Factor; C032854:leupeptin; D014805:Vitamin B 12