PMID:23483992

Citation	Biswas, A, Elmatari, D, Rothman, J, LaMunyon, CW and Said, HM (2013) Identification and functional characterization of the Caenorhabditis elegans riboflavin transporters rft-1 and rft-2. PLoS ONE 8:e58190
Abstract	Two potential orthologs of the human riboflavin transporter 3 (hRFVT3) were identified in the C. elegans genome, Y47D7A.16 and Y47D7A.14, which share 33.7 and 30.5% identity, respectively, with hRFVT3. The genes are tandemly arranged, and we assign them the names rft-1 (for Y47D7A.16) and rft-2 (for Y47D7A.14). Functional characterization of the coding sequences in a heterologous expression system demonstrated that both were specific riboflavin transporters, although the rft-1 encoded protein had greater transport activity. A more detailed examination of rft-1 showed its transport of riboflavin to have an acidic pH dependence, saturability (apparent Km = 1.4 ± 0.5 µM), inhibition by riboflavin analogues, and Na(+) independence. The expression of rft-1 mRNA was relatively higher in young larvae than in adults, and mRNA expression dropped in response to RF supplementation. Knocking down the two transporters individually via RNA interference resulted in a severe loss of fertility that was compounded in a double knockdown. Transcriptional fusions constructed with two fluorophores (rft-1::GFP, and rft-2::mCherry) indicated that rft-1 is expressed in the intestine and a small subset of neuronal support cells along the entire length of the animal. Expression of rft-2 is localized mainly to the intestine and pharynx. We also observed a drop in the expression of the two reporters in animals that were maintained in high riboflavin levels. These results report for the first time the identification of two riboflavin transporters in C. elegans and demonstrate their expression and importance to metabolic function in worms. Absence of transporter function renders worms sterile, making them useful in understanding human disease associated with mutations in hRFVT3.
Links	PubMed PMC3590142 Online version:10.1371/journal.pone.0058190
Keywords	Analysis of Variance; Animals; Base Sequence; Biological Transport/genetics; Caenorhabditis elegans/genetics; Caenorhabditis elegans/metabolism; Caenorhabditis elegans Proteins/genetics; Caenorhabditis elegans Proteins/metabolism; Cloning, Molecular; DNA Primers/genetics; Green Fluorescent Proteins/metabolism; Humans; Hydrogen-Ion Concentration; Intestines/metabolism; Larva/metabolism; Luminescent Proteins/metabolism; Membrane Transport Proteins/genetics; Membrane Transport Proteins/metabolism; Molecular Sequence Data; Nerve Tissue Proteins/genetics; RNA Interference; Real-Time Polymerase Chain Reaction; Riboflavin/metabolism; Sequence Analysis, DNA; Sequence Homology
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
CAEEL:Q3LFN0		GO:0032217: riboflavin transporter activity	ECO:0000314:		F		Figure 2 shows riboflavin uptake in rft-1 clones compared to control cells.	complete CACAO 8548
CAEEL:Q3LFN0		GO:0009792: embryo development ending in birth or egg hatching	ECO:0000315:		P		Figure 4 shows the effects of RNAi targeting the rft-1 transcripts on eggs.	complete CACAO 8600
CAEEL:Q3LFN0	involved_in	GO:0009792: embryo development ending in birth or egg hatching	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete
CAEEL:Q3LFN0	enables	GO:0032217: riboflavin transmembrane transporter activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
CAEEL:Q95XZ2		GO:0032217: riboflavin transporter activity	ECO:0000314:		F		Figure 2 shows rft-1 and rft-2 clones riboflavin uptake activity against control cells.	complete CACAO 8549
CAEEL:Q95XZ2	enables	GO:0032217: riboflavin transmembrane transporter activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete

References

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