Growth phase regulation of the main folate transporter of Leishmania infantum and its role in methotrexate resistance.
The protozoan parasite Leishmania relies on the uptake of folate and pterin from the environment to meet its nutritional requirements. We show here that a novel gene (folate transporter 1 (FT1)) deleted in a Leishmania infantum methotrexate-resistant mutant corresponds to the main folate transporter (K(m), 410 nM). FT1 was established as the main folate transporter by both gene transfection and by targeted gene deletion. Modulation of the expression of FT1 by these manipulations altered the susceptibility of Leishmania cells to methotrexate. Folate transport was stage-regulated with higher activity in the logarithmic phase and less in the stationary phase. FT1 fused to green fluorescent protein led to the observation that FT1 was located in the plasma membrane in the logarithmic phase but was retargeted to an intracellular organelle followed by a degradation of the protein in stationary phase. Leishmania has several folate transporters with different characteristics, and the growth stage-related activity of at least one transporter is regulated post-translationally.
Richard
Dave
D
Centre de Recherche en Infectiologie du Centre de Recherche du Centre Hospitalier and Division de Microbiologie, Faculté de Médecine, Université Laval, Québec G1V 4G2, Canada.
Leprohon
Philippe
P
Drummelsmith
Jolyne
J
Ouellette
Marc
M
D000595:Amino Acid Sequence; D000818:Animals; D027321:Anion Transport Proteins / Q000235:genetics* / Q000502:physiology*; D004351:Drug Resistance / Q000235:genetics*; D005492:Folic Acid / Q000378:metabolism; D017353:Gene Deletion; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins / Q000235:genetics; D018314:Leishmania infantum / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development*; D008727:Methotrexate / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins; D014162:Transfection
D027321:Anion Transport Proteins; D011993:Recombinant Fusion Proteins; C497355:folate transporter 1, Leishmania infantum; C485192:green fluorescent protein, Aequorea victoria; D049452:Green Fluorescent Proteins; D005492:Folic Acid; D008727:Methotrexate