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Cell biology of the human proton‐coupled folate transporter (hPCFT) in renal epithelial MDCK cells
Author(s) -
Subramanian Veedamali Subbian,
Marchant Jonathan S,
Said Hamid M
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1156.2
Subject(s) - transporter , microbiology and biotechnology , transmembrane protein , solute carrier family , membrane transport protein , transmembrane domain , transport protein , cell , function (biology) , biochemistry , chemistry , biology , amino acid , receptor , gene
The human proton‐coupled folate transporter (hPCFT) is a recently identified folate transporter. Little is known about the structure‐function relationship of the different domains of this transporter, in particular which regions are important for folate transport as well as targeting of the transporter to the apical cell surface. Our aim in this study was to investigate the role of the COOH‐terminal domain and a well conserved sequence separating transmembrane (TM) domains TM2 and TM3 (DXXGRR; amino acids 109–114) to be important for transport function. Using live cell (MDCK) imaging approaches, we demonstrate that a hPCFT‐YFP construct is functionally expressed at the apical membrane domain and is localized differentially to the human reduced folate carrier (hRFC). In addition, the predicted COOH‐terminal of the hPCFT is not essential for apical targeting or functionality, and that mutations that ablate a consensus beta‐turn sequence separating predicted TM2 and TM3 abolished apical 3 H‐folic acid uptake (due to retention in ER). Further, cell surface delivery of hPCFT is disrupted by microtubule depolymerization, and by overexpression of p50. These data presents information regarding structure‐function and membrane targeting of the hPCFT polypeptide, as well as the mechanisms that control its steady state expression in renal epithelial cells. [Supported by grants from the NIH and the DVA].