African trypanosomes are the causative agents of sleeping sickness in man and of nagana in domestic livestocks in tropical Africa. Their development in host blood depends entirely on glycolysis in the microbody organelles, the glyscosomes, for energy. In Trypanosoma brucei, all the glycolytic enzymes in the organelle can be crosslinked together without loss of activities. There is, however, little substrate channeling among the crosslinked enzymes. The glysocomal membrane compartmentalizes the glycolytic process, its selective permeabilities to sugar phosphates, ADP and NAD+ performs unique functions, such as synthesis of intraglycosomal ATP from exogeneous ADP and alpha-glycerol phosphate. The glycosomal membrane and two integral membrane proteins of 26K and 24K were recently purified. An in vitro assay of insertion of precursor proteins into glycosome has been established. The process is dependent on ATP and inhibitable by suramin. The chemically synthesized C-terminal peptide of glycosomal 3-phosphoglycerate kinase (PGK) postulated as essential for PGK insertion, was, however, without effect. Glycosomal hexokinase, glycerol kinase and phosphoglucoisomerase were purified to homogeneity. They and phosphoenolpyruvate carboxykinase and malate dehydrogenase of the procyclic form glycosome will be partly sequenced for gene cloning. The cloned genes will be sequenced, transcribed and translated in vitro for the insertion assay. Antibodies generated against the purified enzymes will be used to monitor precursor insertions. Monoclonal antibodies to glycosomal membrane proteins will be made and tested against protein insertion. A recent indication that polyamine deficiency may trigger transformation of T. brucei from long-slender to short-stumpy form prompted us to clone and sequence the T. brucei ornithine decarboxylase (ODC) gene. The gene was transcribed and translated in vitro and the protein product identified by ODC antibody. The in vitro transformation of T. brucei will be established, and polyamines and ODC inhibitors will be tested. Cloned ODC genes and ODC antibodies will monitor the expression of ODC during transformation. Similarly, the genes and antibodies of the glycolytic enzymes will examine the mechanisms of genetic control of glycosomal composition during T. brucei transformation. By understanding the regulation and mechanism of glycosomal biogenesis one can try to interfere and confuse these processes in the parasite for antitrypanosomal therapy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI021786-07
Application #
3132150
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Project Start
1984-12-01
Project End
1992-11-30
Budget Start
1990-12-01
Budget End
1991-11-30
Support Year
7
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Pharmacy
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Hu, Huiqing; Gourguechon, Stéphane; Wang, Ching C et al. (2016) The G1 Cyclin-dependent Kinase CRK1 in Trypanosoma brucei Regulates Anterograde Protein Transport by Phosphorylating the COPII Subunit Sec31. J Biol Chem 291:15527-39
Bessat, Mohamed; Knudsen, Giselle; Burlingame, Alma L et al. (2013) A minimal anaphase promoting complex/cyclosome (APC/C) in Trypanosoma brucei. PLoS One 8:e59258
Garlapati, Srinivas; Saraiya, Ashesh A; Wang, Ching C (2011) A La autoantigen homologue is required for the internal ribosome entry site mediated translation of giardiavirus. PLoS One 6:e18263
Sun, Lu; Wang, Ching C (2011) The structural basis of localizing polo-like kinase to the flagellum attachment zone in Trypanosoma brucei. PLoS One 6:e27303
Li, Zhi; Umeyama, Takashi; Li, Ziyin et al. (2010) Polo-like kinase guides cytokinesis in Trypanosoma brucei through an indirect means. Eukaryot Cell 9:705-16
Li, Ziyin; Umeyama, Takashi; Wang, C C (2009) The Aurora Kinase in Trypanosoma brucei plays distinctive roles in metaphase-anaphase transition and cytokinetic initiation. PLoS Pathog 5:e1000575
Gourguechon, Stephane; Wang, Ching C (2009) CRK9 contributes to regulation of mitosis and cytokinesis in the procyclic form of Trypanosoma brucei. BMC Cell Biol 10:68
Li, Ziyin; Lee, Ju Huck; Chu, Feixia et al. (2008) Identification of a novel chromosomal passenger complex and its unique localization during cytokinesis in Trypanosoma brucei. PLoS One 3:e2354
Li, Ziyin; Umeyama, Takashi; Wang, Ching C (2008) The chromosomal passenger complex and a mitotic kinesin interact with the Tousled-like kinase in trypanosomes to regulate mitosis and cytokinesis. PLoS One 3:e3814
Li, Ziyin; Lindsay, Megan E; Motyka, Shawn A et al. (2008) Identification of a bacterial-like HslVU protease in the mitochondria of Trypanosoma brucei and its role in mitochondrial DNA replication. PLoS Pathog 4:e1000048

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