Regulation of reduced-folate transporter-1 in retinal pigment epithelial cells by folate

Hany Naggar, Tracy K. Van Ells, Vadivel Ganapathy, Sylvia B Smith

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

Purpose: Reduced-folate transporter-1 (RFT-1), a typical transport protein with 12 membrane-spanning domains, transports reduced-fblates, such as N 5-methyltetrahydrofolate (MTF), the predominant circulating form of folate. RFT-1 is localized to the RPE apical membrane and transports folate from RPE to photoreceptor cells. We asked whether RFT-1 activity in RPE is altered under high folate conditions. Materials and methods: ARPE-19 cells were cultured 24, 48, or 72 h in medium containing either 0.5 nM, 5.0 nM, or 2.26 μM MTF, and the activity of RFT-1 was assessed by determining the uptake of N 5-MTF. Semiquantitative reverse transcription-polymerase chain reaction and Western blot analysis were used to study RFT-1 gene and protein expression. Results: Cells treated for 72 h with 2.26 μM MTF showed a significant (40%) decrease in MTF uptake compared to cells exposed to 0.5 nM or 5 nM MTF. The effect of high concentrations of folate on RFT-1 activity was specific. Kinetic analysis showed that folate-induced attenuation of RFT-1 activity was associated with a decrease in the maximal velocity of the transporter, but no change in the substrate affinity. Steady-state levels of RFT-1 mRNA and protein decreased significantly in the presence of excess folate. Conclusions: Excess folate levels downregulate RFT-1 in RPE. This study represents the first molecular analysis of the regulation of RFT-1 by folate in RPE and reveals attenuation of the activity and expression of a folate transport protein under conditions of high levels of folate.

Original languageEnglish (US)
Pages (from-to)35-44
Number of pages10
JournalCurrent Eye Research
Volume30
Issue number1
DOIs
StatePublished - Jan 1 2005

Keywords

  • ARPE-19 cells
  • Cell culture
  • Folate
  • Reduced-folate transporter
  • Retinal pigment epithelium

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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