Cloning and Functional Characterization of the Proton-Coupled Electrogenic Folate Transporter and Analysis of Its Expression in Retinal Cell Types
Open Access
- 1 November 2007
- journal article
- research article
- Published by Association for Research in Vision and Ophthalmology (ARVO)
- Vol. 48 (11), 5299-5305
- https://doi.org/10.1167/iovs.07-0288
Abstract
Purpose. In a prior study the cellular uptake of folate was investigated in the retina. Recently, a new proton-coupled folate transporter (PCFT) in human intestine was reported. In the present study, the expression of this novel transporter in the retina was determined, the mouse orthologue was cloned from retinal tissue, and its transport function was characterized. methods. RT-PCR and folate uptake measurements were used to detect the expression of PCFT in mouse retina and in retinal cell types. The expression of PCFT mRNA in intact retina was investigated by in situ hybridization. Mouse PCFT cDNA was cloned, and its transport characteristics were analyzed by electrophysiological methods after expression of the cloned transporter in Xenopus laevis oocytes. results. RT-PCR showed expression of PCFT mRNA in both neural retina and RPE eye cup. In situ hybridization detected PCFT mRNA in all retinal cell layers. Proton-coupled folate uptake was detectable in primary cultures of ganglion, Müller, and RPE cells of mouse retina and in RPE, ganglion, and Müller cells of human or rat origin. In X. laevis oocytes expressing the cloned mouse PCFT, folate and its derivatives methotrexate and 5-methyltetrahydrofolate induced H+-coupled inward currents with K t values of 1.2 ± 0.1, 4.6 ± 0.5, and 3.5 ± 0.8 μM, respectively. The transport process showed an H+-folate stoichiometry of 1:1, suggesting that PCFT transports the zwitterionic form of folate. conclusions. This is the first report on the expression of PCFT in the retina. All cell layers of the retina express this transporter. Mouse PCFT, cloned from retina, mediates H+-coupled electrogenic transport of folate and its derivatives.Keywords
This publication has 22 references indexed in Scilit:
- Functional Characterization of Human Proton-Coupled Folate Transporter/Heme Carrier Protein 1 Heterologously Expressed in Mammalian Cells as a Folate TransporterJournal of Pharmacology and Experimental Therapeutics, 2007
- Identification of an Intestinal Folate Transporter and the Molecular Basis for Hereditary Folate MalabsorptionCell, 2006
- Expression, Subcellular Localization, and Regulation of Sigma Receptor in Retinal Müller CellsPublished by Association for Research in Vision and Ophthalmology (ARVO) ,2006
- Haem carrier protein 1 (HCP1): Expression and functional studies in cultured cellsFEBS Letters, 2006
- The Intestinal Heme Transporter RevealedCell, 2005
- Identification of an Intestinal Heme TransporterCell, 2005
- Induction of Cystine-Glutamate Transporter xc−by Human Immunodeficiency Virus Type 1 Transactivator Protein Tat in Retinal Pigment EpitheliumPublished by Association for Research in Vision and Ophthalmology (ARVO) ,2004
- Folate receptor endocytosis and traffickingAdvanced Drug Delivery Reviews, 2004
- SLC19: the folate/thiamine transporter familyPflügers Archiv - European Journal of Physiology, 2004
- Functional coupling between a bafilomycin A1-sensitive proton pump and a probenecid-sensitive folate transporter in human placental choriocarcinoma cellsBiochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1994