Molecular and Cell Biology of Pathogenic Protozoa

Kristin M. Hager     Assistant Professor Ph.D., University of Alabama-Birmingham Postdoctoral Fellowship, University of Pennsylvania School of Medicine

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What makes an organism a successful parasite? Our lab utilizes state of the art techniques such as time lapse video microscopy and advanced genetic analysis to study interactions between the parasite and host cell in order to answer this question. We believe that the protozoan parasite, Toxoplasma gondii, is spectacularly successful due to its ability to secrete proteins that allow it to interact with virtually *any* nucleated host cell during invasion and intracellular survival. A key step in protein secretion is the organisms' ability to synthesize and properly target these invasion/maintenance proteins to their respective organelles. Our laboratory is interested in dissecting the central steps involved in these phenomena and in general are interested in intracellular trafficking of proteins in protozoan parasites. Our current model system is T. gondii. It is an obligate intracellular parasite normally controlled by an active immune system. Unfortunately, there has been an alarming rise in the number of immunosupressed individuals such as HIV patients. Congenital toxoplasmosis is a major source of neurological birth defects. 

The early secretory pathway has several compelling features. Our laboratory is interested in events that occur between the endoplasmic reticulum (ER) and Golgi. In T. gondii , the apical end of the nuclear envelope appears to be the sole site of intense vesicle trafficking and recycling of proteins between the Golgi and ER (see Figure 1 and 1b) (Hager et al. 1999). It is likely that this region represents the organisms 'Achilles heel'; a bottle-neck through which all secretory proteins must process. The long-term goal of my lab is to determine what genes are expressed at this site, and characterize their role in regulating the trafficking of the specialized 'invasion/maintenance' proteins. Projects currently in progress are investigation of: the molecular and biochemical mechanisms of ER-Golgi recycling proteins using the receptor, Tg ERD2, investigation of the nature of these protein interactions with other components of the secretory apparatus in T. gondii by identifying specific vesicular coat proteins (COPI) such as Tg b COP, and determining the essential nature of the recycling genes for parasite viability. To achieve these ends, a variety of cell biological, molecular and genetic approaches will be used to characterize the recycling-specialized protein interactions. Genetically engineered Toxoplasma defective in sorting genes will be used to determine the importance of regulatory components in-vivo. We will use FACS analysis for a screen of mutants. These studies will further our understanding of the components and mechanisms involved in the Toxoplasma-host cell interactions and how they impinge on T. gondii pathogenesis.

Figure 1. T. gondii ultrastructure: two intracellular tachyzoites in a host cell.

Mi: micronemes, R: rhoptries, DG: dense granules, PV: parasitophorous vacuole, G: Golgi, N: nucleus, PVM: parasite vacuole membrane, ER: endoplasmic reticulum, Green star: Apicoplast, Rb: residual body, M: Mitochondria (note: these are host mitochondria, closely associated with the parasite membrane), Yellow star: apical face of nuclear envelope, see Figure 1b for close-up.

Figure 1b. Model for the apical end of nuclear envelope in T. gondii . Nucleus (N), Coated vesicle (C). Red arrows indicate movement of COPI coated vesicles. Green arrow head indicates movement of COPII coated vesicles. Light yellow receptor in Golgi: position of recycling HDEL-receptor encoded by Tg ERD2. Bright yellow star: apical end of nuclear envelope in T. gondii. Both electron micrographs courtesy of Dr. L. Tilney, Univ. Pennsylvania

Selected Publications

Hager, K.M., Pierce, M., Moore, R., Tytler, E., Esko, J., and S. Hajduk. 1994. Endocytosis of a cytotoxic human high density lipoprotein results in disruption of acidic intracellular vesicles and subsequent killing of African trypanosomes.  J. Cell Biol. 126:155-167.

Hager, K.M. and S. Hajduk. 1997.  Mechanism of resistance of African trypanosomes to cytotoxic human HDL.  Nature. 385: 823-825.

Hager, K.M., B. Striepen, L. Tilney and D.S. Roos.  1999. The nuclear envelope serves as an intermediary between the ER and the Golgi complex in the intracellular parasite Toxoplasma gondii.  J. Cell Science.  112: 2631-2638.

Roos, D.S., M.J. Crawford, R.G.K. Donald, L.M. Fohl, K.M. Hager, J.C. Kissinger, M.G.  Reynolds, B. Striepen, and W.J. Sullivan, Jr.  1999.  Transport and trafficking: Toxoplasma as a model for Plasmodium.  Novartis Fdn. Symp.  226.

Roos, D.S., J.A. Darling, M.G. Reynolds, K.M. Hager, B.S. Striepen and J.C. Kissinger. (1999) Toxoplasma as a model parasite: apicomplexan biochemistry, cell biology, molecular genetics, genomics and beyond. Biology of Parasitism. C. Tschudi and E. Pearce Editors. Kluwer Academic Publishers, Boston, Massachusettes. 

Shimamura M, K.M. Hager, and S.L. Hajduk. 2001.  The lysosomal targeting and intracellularmetabolism of trypanosome lytic factor by Trypanosoma brucei brucei.  Mol. Biochem. Parasitol. 115(2): 227-37.

Pfluger S.L., H.V. Goodson, J.M. Moran, C.J. Ruggiero, X. Ye, K.M. Emmons, and K.M. Hager. 2005.  A receptor for retrograde transport in the Apicomplexan parasite, Toxoplasma gondii. Eukaryotic Cell. 4(2): 432-42.

 

Smith S.S., S.L. Pfluger, E. E. Hjort, A.G. McArthur, and K.M. Hager. 2007.  Molecular evolution of the vesicle coat component bCOP in Toxoplasma gondii. Molecular Phylogeny Evolution. 44(3): 1284-1294.

Moran, J.M., Smith, S.S., and K.M. Hager. 2007. The apicomplexan parasite Toxoplasma gondii possesses a receptor for activated C kinase ortholog. Biochemical and Biophysical Research Communications. 363(3): 680-686

Hager K.M. and V. Caruthers.  2008.  ‘MAR’veling at parasite invasion.  Trends in Parasitology. 24(2):  51-54.

Eggleston, T.L., Fitzpatrick, E., and K.M. Hager. 2008. Parasitology as a teaching tool: isolation of apicomplexan cysts from store bought meat. Cell Biology Education-Life Sciences Education (CBE-LSE). 7(2008): 184-192.

Walker, M.E., Hjort, E.E., Hornick, J.E., Hinchcliffe, E.H., Archer, W., and K.M. Hager. 2008. Toxoplasma gondii actively remodels the host microtubule network. Microbe and Infection. In Press.