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Y. Dun, P.D. Prasad, B. Mysona, T.K. Van Ells, V. Ganapathy, S.B. Smith; Analysis of Folate Receptor (FR) Alpha in Ganglion and Müller Cells and FR Delta in RPE . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2969.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Folate, a water–soluble vitamin essential for DNA, RNA and protein synthesis, is required for cell survival. Lack of folate is implicated in optic neuropathy, in which papillo–macular bundle fibers of the optic nerve are lost. Recently, we characterized the transport of folate in RPE cells and reported the polarized distribution of two folate transport proteins, reduced–folate transporter–1 (RFT–1) and folate receptor α (FRα). Little is known about folate uptake mechanisms in other retinal cells, particularly ganglion cells, which are affected by lack of folate; or Müller cells, which support neuronal cells. We hypothesized that FRα would be present and functional in ganglion and Müller cells. We also investigated whether FRΔ, the newest FR, was present in RPE cells. Methods: RGC–5, rMC–1 and primary mouse Müller cells were used for RT–PCR, western blot analysis and immunohistochemistry to study FRα expression. ARPE–19 cells were used to study expression of FRΔ. Results: RT–PCR analysis of RNA isolated from rMC–1 and RGC–5 cells amplified the expected 638 bp product using primers specific for rat FRα. Immunoblotting using a commercially available antibody for FRα detected a band of the appropriate size (∼30 kD) consistent with a positive detection of FRα. Immunohistochemistry in RGC–5, rMC–1 and primary Müller cells showed positive detection of FRα. RT–PCR analysis of RNA isolated from ARPE–19 cells using primers specific for FRΔ amplified the expected 604 bp product. The PCR product was cloned into pGEM–T easy vector, sequenced to confirm its identity and then used to screen a mouse spleen cDNA library. A positive clone was identified which was purified to homogeneity by secondary screening and confirmed by nucleotide sequencing. The cloned mouse Folbp3 (the murine ortholog of FRΔ) is 1,130 bp long and has an open reading frame of 735 bp, encoding a protein of 244 amino acids. Conclusions: FRα is present in ganglion and Müller cells; FRΔ is present in RPE cells. This is the first comprehensive investigation of FRs in retinal cells. Future studies will determine the role played by these FRs in the uptake of folate and factors that may compromise their function.
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