The VSG expression site of T. brucei contains several Expression Site Associated Genes which are expressed at high levels in a stage-specific manner. We have previously cloned and sequenced an ESAG which we named T- LR (for Trypanosome Leucine-rich Repeat) and others have called ESAG 8. The protein sequence predicted from T-LR contains two interesting domains. One has homology to the RING finger motif in regulatory proteins which bind DNA in a sequence-specific fashion, while the other is a leucine-rich repeat (LRR) sequence found in wide variety of prokaryotic and eukaryotic proteins involved in protein-protein interactions. Thus, it is likely that T-LR has an important regulatory role in parasite physiology. The overall aim of this project is to elucidate the role of T-LR and to identify the molecular mechanisms underlying its function. We also plan to identify and characterize other regulatory molecules with which T-LR may interact. We will express recombinant T-LR proteins in bacterial, yeast, insect or mammalian expression systems and produce specific antibodies against the potential nucleic acid- and protein-binding domains. The antibodies will be used to characterize the T-LR protein in terms of size, post- translational modification, and stage-specific expression and to determine its subcellular location. We will then identify molecules which interact with T-LR within the trypanosome cell. These nucleic acids and proteins will be characterized and their genes cloned. In addition, we will modulate the normal expression of the T-LR gene by knockout of the expressed T-LR gene in bloodstream forms and over-expression of T-LR in bloodstream and procyclic forms. Changes in trypanosome growth, morphology and expression of other specific genes will identify cellular processes involving T-LR. These studies are designed to identify new molecular processes that provide the basis for development of novel parasite control strategies. In addition, like the study of several other phenomena in T. brucei, they should also provide new insights into the fundamental molecular biology of all eukaryotic organisms.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29AI039086-04
Application #
2887111
Study Section
Tropical Medicine and Parasitology Study Section (TMP)
Program Officer
Fairfield, Alexandra
Project Start
1996-05-01
Project End
2001-04-30
Budget Start
1999-05-01
Budget End
2000-04-30
Support Year
4
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Seattle Biomedical Research Institute
Department
Type
DUNS #
City
Seattle
State
WA
Country
United States
Zip Code
98109