Mapping of the functional domains of TOR, an adaptor protein regulating protein trafficking in Leishmania.Leishmania Leishmania /Leish·ma·nia/ (lesh-ma´ne-ah) a genus of parasitic protozoa, including several species pathogenic for humans. In some classifications, organisms are placed in four complexes comprising species and subspecies: L. are pathogenic protozoa of the family Trypanosomatidae. This parasite is found in the tropical and subtropical regions on every continent except for Australia and Antarctica. Leishmania cause Leishmaniasis leishmaniasis (lēsh'mənī`əsĭs), any of a group of tropical diseases caused by parasitic protozoans of the genus Leishmania. and is transmitted by the bite of sand flies. It is estimated that 12 million people are infected worldwide and hundreds of thousands to millions of individuals are newly infected each year. Excessive morbidity, occassional mortality and difficulty/cost of treatment with the older drugs have encouraged the search for newer anti-Leishmania compounds. Drug resistance to the older drugs has also hampered treatment.
While investigating how Leishmania becomes resistant to toxic purine nucleosides, we identified and cloned the TOR gene [1,2]. TOR is ah atypically multidrug resistance protein that was originally discovered by its ability, when present in high levels, to render Leishmania resistant to toxic nucleosides. Subsequent research showed that it also elicited resistance to a wide variety of structurally diverse toxins, among them many of the clinically important anti-Leishmania compounds. TOR is found in at least two regions in Leishmania: the mitochondria and the Golgi/trans Golgi network. In the Golgi/trans Golgi network, TOR acts as a traffic regulator and influences the amount of purine transporters which reach the plasma membrane .
Truncation mutants expressed in Leishmania and the yeast Saccharomyces Saccharomyces: see yeast. and far western blots showed that the minimal domain extending from Met31 to Ser204 was responsible for interacting with TOTs (i.e., targets of TOR). Expression of the C terminal half of TOR as a "quasi" dominant-negative mutant showed that it also exhibited biological activity. Deletion of a putative clathrin binding domain in the C terminal half by site specific mutagenesis abolished TOR's ability to internalize the adenosine transporter. Far western experiments showed that the C terminal half of TOR also interacted with several of the Rab proteins. Furthermore, high level ectopic expression of these Rab proteins in Leishmania acted as a dominant-negative mutant and impaired the ability of TOR to internalize the trafficking of the adenosine permease permease /per·me·ase/ (per´me-as) former name for transport protein.
An enzyme that promotes the passage of a substance across a cell membrane. . Saccharomyces Rab4 and Rab5 null mutants were no longer able to support the intemalization of the Leishmania adenosine permease by TOR.
These data indicate that TOR is an adaptor protein interacting with TOTs at one end and multiple components of the vesicle vesicle /ves·i·cle/ (ves´i-k'l)
1. a small bladder or sac containing liquid.
2. a small circumscribed elevation of the epidermis containing a serous fluid; a small blister. trafficking apparatus at the other end. TOR appears to be the progenitor of a new class of endocytotic accessory proteins. In contrast to the other heterotetrameric and monomeric adaptor protein classes, TOR is a homotetrameric protein.
[1.] Detke, S. (1997) Identification of a transcription factor like protein at the TOR locus in Leishmania mexicana amazonensis. Mol. Biochem. Parasitol. 90:505-511.
[2.] Kerby, B.R., and Detke, S. (1993) Reduced purine accumulation is encoded on an amplified DNA in Leishmania mexicana amazonensis resistant to toxic nucleosides. Mol. Biochem. Parasitol. 60:171-185.
[3.] Detke, S. (2007) TOR-induced resistance to toxic adenosine analogs in Leishmania brought about by the internalization and degradation of the adenosine permease. Exp. Cell Res. 313:1963-1978.
Department of Biochemistry & Molecular Biology, University of North Dakota, Grand Forks, North Dakota
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