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This Article Full Text Full Text (PDF) All Versions of this Article: 581/2/457    most recent jphysiol.2007.129171v1 Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Bose, S. Articles by Seetharam, B. Search for Related Content PubMed PubMed Citation Articles by Bose, S. Articles by Seetharam, B. Related Collections Cellular

Division of Gastroenterology and Hepatology, Departments of
¹ Medicine
² Biochemistry, Medical College of Wisconsin and Veterans Administration Medical Center, Milwaukee, Wisconsin 53295, USA

Cells that are metabolically active and in a high degree of differentiation and proliferation require cobalamin (Cbl: vitamin B₁₂) and they obtain it from the circulation bound to transcobalamin (TC) via the transcobalamin receptor (TC-R). This study has investigated the plasma membrane dynamics of TC-R expression in polarized human intestinal epithelial Caco-2 cells using techniques of pulse-chase labelling, domain-specific biotinylation and cell fractionation. Endogenously synthesized TC-R turned over with a half-life (T_1/2) of 8 h following its delivery to the basolateral plasma membrane (BLM). The T_1/2 of BLM delivery was 15 min and TC-R delivered to the BLM was endocytosed and subsequently degraded by leupeptin-sensitive proteases. However, about 15% of TC-R endocytosed from the BLM was transcytosed (T_1/2, 45 min) to the apical membranes (BBM) where it underwent endocytosis and was degraded. TC-R delivery to both BLM and BBM was inhibited by Brefeldin A and tunicamycin, but not by wortmannin or leupeptin. Colchicine inhibited TC-R delivery to BBM, but not BLM. At steady state, apical TC-R was associated with megalin and both these proteins were enriched in an intracellular compartment which also contained Rab5 and transferrin receptor. These results indicate that following rapid delivery to both plasma membrane domains of Caco-2 cells, TC-R undergoes constitutive endocytosis and degradation by leupeptin-sensitive proteases. TC-R expressed in apical BBM complexes with megalin during its transcytosis from the BLM.

(Received 25 January 2007; accepted after revision 5 March 2007; first published online 8 March 2007)
Corresponding author B. Seetharam: Zablocki VA Medical Center, 5000 West National Avenue, Research 151, Bldg 70C, Milwaukee, WI 53295, USA. Email: seethara{at}mcw.edu