The clinical course and genetic defect in the PCFT gene in a 27-year-old woman with hereditary folate malabsorption

Abstract

We report 2 sequential homozygous mutations in the recently cloned proton-coupled folate transporter (PCFT) gene, resulting in the absence of this protein, in a 27-year-old woman with hereditary folate malabsorption, normal in all respects having completed higher education, who has been treated with parenteral 5-formyltetrahydrofolate since infancy.

Figure. Genetic and Functional analysis of PCFT mutation

(A) Pedigree of the family with HFM. The filled symbols indicate the patient with both PCFT alleles mutated. The half-filled symbols indicate the parents with one normal and one mutated PCFT allele. (B) Chromatograms of sequenced PCFT genomic DNA showing the homozygous GC>AA mutations in the patient and the heterozygous GC>AA mutations in her parents. (C) Genomic organization of PCFT showing the location of the GC>AA mutations. (D) Functional analysis of the patient’s mutated and wild-type PCFT cDNA transiently transfected into a Hela cell line that lack both genomic reduced folate carrier and PCFT expression Uptake of 0.5 μM [H³]5-methyltetrahydrofolate was assessed over two minutes at pH 5.5 as previously described . (E) Northern blot analysis of mRNA obtained from the Hela cells transiently transfected with the patient’s mutated or wild-type PCFT mRNA. (F) Western Blot analysis of whole cell lysate from the cells described in Figure E. PCFT proteins were tagged with hemagglutinin epitope inserted in the C-terminus and the blot was probed with anti-hemagglutinin antibody.