Leishmania parasites are protozoa that cause devastating diseases in much of the tropical and subtropical world. Current drug treatments for these parasites are deficient in many respects and improved therapies founded upon the basic biochemistry of the parasites are urgently required. In this proposal we will study the role of three membrane transporters, those for glucose, myo-inositol and nucleosides, in the biology of Leishmania parasites. 1) Glucose is a major source of metabolic energy in the promastigote stage of the parasite that lives in the insect vector. In one set of experiments, the function of the Pro-1 glucose transporter genes in vivo will be studied by knocking out these genes by targeted gene replacement and testing the phenotype of the resulting Pro- 1 null mutants. In a second set of experiments, a subcellular targeting signal that exists within the flagellar isoform of Pro-1 but not within the plasma membrane isoform of Pro-1 will be identified. This will be done by mutagenizing the one domain of the flagellar isoform that differs from the plasma membrane isoform and determining which amino acids are required to target the protein to the flagellum. These experiments will begin to define a novel pathway for subcellular targeting of membrane proteins. 2) Myo-inositol is an important building block for many of the cell surface components of Leishmania parasites. In one aim of this proposal, site-directed mutagenesis will be used to identify -amino acids in the myo-inositol transporter that allow this permease to concentrate myo-inositol inside the parasite by co-transporting a proton down its electrochemical gradient. These experiments will begin a detailed structure-function analysis of this model proton symporter. 3) Purines are essential nutrients that can not be synthesized by Leishmania parasites. The final specific aim of this proposal is to clone a gene encoding a Leishmania nucleoside transporter, a membrane permease responsible for salvaging purine nucleosides from the host. This nucleoside transporter gene will be functionally expressed, and its DNA sequence will be determined to predict the amino acid sequence of the transporter. Nucleoside transporters are of considerable pharmacological importance, because they mediate the uptake of cytotoxic nucleoside analogs that are candidates for novel anti-parasite drug therapies. These experiments will initiate a detailed molecular study of these biochemically and pharmacologically important transporters in these purine auxotrophic parasites.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI025920-11
Application #
2671913
Study Section
Tropical Medicine and Parasitology Study Section (TMP)
Project Start
1991-09-01
Project End
2001-07-31
Budget Start
1998-08-01
Budget End
1999-07-31
Support Year
11
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Oregon Health and Science University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
009584210
City
Portland
State
OR
Country
United States
Zip Code
97239
Landfear, Scott M; Tran, Khoa D; Sanchez, Marco A (2015) Flagellar membrane proteins in kinetoplastid parasites. IUBMB Life 67:668-76
Rodriguez-Contreras, Dayana; Aslan, Hamide; Feng, Xiuhong et al. (2015) Regulation and biological function of a flagellar glucose transporter in Leishmania mexicana: a potential glucose sensor. FASEB J 29:11-24
Rodriguez-Contreras, Dayana; Landfear, Scott M (2014) Transporters, channels and receptors in flagella. Channels (Austin) 8:477-8
Tran, Khoa D; Rodriguez-Contreras, Dayana; Vieira, Danielle P et al. (2013) KHARON1 mediates flagellar targeting of a glucose transporter in Leishmania mexicana and is critical for viability of infectious intracellular amastigotes. J Biol Chem 288:22721-33
Feng, Xiuhong; Rodriguez-Contreras, Dayana; Polley, Tamsen et al. (2013) 'Transient' genetic suppression facilitates generation of hexose transporter null mutants in Leishmania mexicana. Mol Microbiol 87:412-29
Tran, Khoa D; Rodriguez-Contreras, Dayana; Shinde, Ujwal et al. (2012) Both sequence and context are important for flagellar targeting of a glucose transporter. J Cell Sci 125:3293-8
Blume, Martin; Hliscs, Marion; Rodriguez-Contreras, Dayana et al. (2011) A constitutive pan-hexose permease for the Plasmodium life cycle and transgenic models for screening of antimalarial sugar analogs. FASEB J 25:1218-29
Vince, James E; Tull, Dedreia; Landfear, Scott et al. (2011) Lysosomal degradation of Leishmania hexose and inositol transporters is regulated in a stage-, nutrient- and ubiquitin-dependent manner. Int J Parasitol 41:791-800
Feng, Xiuhong; Feistel, Torben; Buffalo, Cosmo et al. (2011) Remodeling of protein and mRNA expression in Leishmania mexicana induced by deletion of glucose transporter genes. Mol Biochem Parasitol 175:39-48
Landfear, S M (2010) Transporters for drug delivery and as drug targets in parasitic protozoa. Clin Pharmacol Ther 87:122-5

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