Toxoplasma: Molecular and Cellular Biology Chapter Abstracts
Chapter 1: The Life Cycle of Toxoplasma gondii
J. P. Dubey
Abstract
Infections by the protozoan parasite, Toxoplasma gondii, are widely prevalent worldwide in animals and human beings. Cats are the only definitive hosts for T. gondii and all other warm-blooded animals are intermediate hosts. Cats excrete the environmentally-resistant oocysts after ingesting any of the three infectious stages of T. gondii, tachyzoites, bradyzoites, and oocysts. Humans and other hosts become infected with T. gondii by ingesting uncooked infected meat, by ingesting food and water contaminated with oocysts, or transplacentally. Cats can excrete millions of oocysts after ingesting tissues of infected animals and oocysts can survive in the harsh environment for months. Contamination of sea water can occur from the oocysts washed off the land and marine mammals can acquire T. gondii infection and some die from toxoplasmosis.
Chapter 2: Ultrastructure
J.-F. Dubremetz and D. J. P. Ferguson
Abstract
Toxoplasma gondii belongs to the phylum Apicomplexa, and therefore shares a number of specific features with the other members of the phylum, that are especially prominent at the ultrastructural level, as already recognized more than 35 years ago (Scholtyseck and Mehlhorn, 1970; Vivier et al., 1970). The tachyzoite of T. gondii can actually be considered the paradigm of Apicomplexa invasive stages, as most of the structural, biological and molecular data concerning these "zoites" have been obtained using this organism as a model, even before its life cycle and phylogenetic links were discovered. Other stages, and especially those occurring in the definitive host have been less studied in many respects, but their ultrastructure is nevertheless very well known. The present chapter will summarize the main ultrastructural characteristics of the different stages of T. gondii, in both the intermediate and definitive hosts, including the tachyzoite and the other stages of asexual reproduction, and the sexual stages, that are typical of enteric coccidia.
Chapter 3: Epidemiology, Diagnostics and Chemotherapy
Eskild Petersen
Abstract
Toxoplasma gondii occur world-wide, but the incidence is higher in tropical areas and decreases with increasing latitude. Seroprevalence in Europe is high, up to 54% in Southern European countries and decrease with increasing latitude to 5% to 10%. The T. gondii seroprevalence of 15.8% in the age group 12-49 years in the United States. T. gondii is a common infection in South America and seroprevalence is high in people from poor socio-economic conditions probably due to water borne transmission and infection often takes place during childhood. Risk factors for infection with T. gondii is mainly through meat products and poor kitchen hygiene in Europe, and surface water in Brazil. T. gondii can be divided into three major genotypes of which type II dominate in Europe, and type I have been found overrepresented in patient with eye disease. The finding of Toxoplasma-specicifc IgM-antibodies do not necessarily mean an acute infection , and the major problem of diagnosis of T. gondii infection in pregnant women is the finding of Toxoplasma-specicifc IgM- and IgG-antobodies. Two-test strategies with IgM-capture assays and direct IgG assays followed by assay of the Toxoplasma-specific IgG-avidity ratio is presently the best option. T. gondii is treated with sulfadiazine and pyrimethamine, but there are only one randomized controlled trials and treatment efficacy is based on animal models and studies with historic controls. A randomized controlled trial found good effect on T. gondii eye disease of trimethoprim and sulfamethoiaxazol. In animal modles the best results for a new drug for treatment is atovaquone.
Chapter 4: Ocular Toxoplasmosis: Clinical Features, Pathology, Pathogenesis, Animal Models and Immune Responses
Fiona Roberts, Annie Kuo, Leigh Jones, Rima McLeod and Craig Roberts
Abstract
Ocular toxoplasmosis occurs during the acute acquired infection, during active congenital infection in utero and the newborn period and as a recrudescent sequela of both types of infection. It also occurs in immune compromised persons usually as reactivation disease.Destruction of the eye is secondary to parasite replication and the immune response to parasite growth may augment this destruction.Clinical features, pathogenesis, immunology and protective and pathogenic mechanisms, pathology, animal models for study of the disease,and methods of treatment are summarized and discussed.
Chapter 5: Congenital Toxoplasmosis
A.W. Pfaff, O. Liesenfeld and E. Candolfi
Abstract
Toxoplasma gondii is one of the few pathogens that regularly cross the placenta. The consequences of maternal infection during pregnancy depend largely on the timing. The risk of fetal infection continually increases with the duration of pregnancy. Conversely, early infection bears the greatest risk of severe sequelae for the unborn child. Brain and eye lesions are the most common consequences of in utero infection. Importantly, babies born without overt clinical symptoms are at risk of developing the disease during childhood or adolescence. These features explain why congenital toxoplasmosis continues to be a major health problem throughout the world. This chapter gives an overview of clinical disease, principles of diagnosis of maternal and fetal infection and treatment of congenital toxoplasmosis. The management and legal situation in different countries is discussed. Finally, the actual knowledge of immunologic control of materno-fetal transmission is presented.
Chapter 6: Innate Recognition and the Regulation of Protective Immunity to Toxoplasma gondii
Marion Pepper and Christopher A. Hunter
Abstract
Most infections with T. gondii are relatively asymptomatic as a consequence of the development of protective immunity that allows long-term control of this persistent organism. However, this pathogen can cause severe disease in immune competent individuals and is a major opportunistic infection in patients with primary or acquired defects in T cell function. The study of these events has led to the identification of the innate and adaptive components required to control T. gondii and highlighted the importance of cell-mediated immunity in resistance to intracellular pathogens. This infection has also provided a model to study general immuno-regulation and the balance between protective and pathological T cell responses. This chapter reviews our current understanding of immunity to T. gondii and outlines some of the prominent questions in this area of research.
Chapter 7: Mechanisms of Immune Evasion
Eric Y. Denkers, Chiang W. Lee, Leesun Kim and Barbara A. Butcher
Abstract
Toxoplasma gondii employs multiple strategies to avoid, deflect or subvert host defense mechanisms. The parasite avoids immunity through cyst formation and sequestration in immunoprivileged tissues as well as in the parasitophorous vacuole within the host cell. It deflects immunity through stage-specific antigen variation. Toxoplasma-induced suppression of immunity occurs through induction of anti-inflammatory mediators such as IL-10 and lipoxin A4. In infected cells, it is now evident that T. gondii directly subverts proinflammatory signaling cascades. Suppression of NFκB and MAPK signal transduction, and activation of anti-inflammatory transcription factor STAT3 have each recently emerged as targets of manipulation by the parasite. As a result, the parasite blocks many activities associated with activation in innate immune cells, including macrophages, dendritic cells and neutrophils. The many ways that Toxoplasma down-modulates immune responses reflects the sophistication of this parasite in its interactions with the host.
Chapter 8: Apoptosis and Its Impact on the Parasite-host Interaction
Carsten G. K. Lüder
Abstract
Apoptosis is a tightly regulated form of programmed cell death in multicellular organisms. With the unexpected observation of an apoptosis-like cell death in protozoans including Toxoplasma gondii it is now clear, however, that it is not confined to metazoans. In vertebrates, apoptosis fulfils essential roles in development, tissue turnover, regulation of immune responses and in innate as well as adaptive defence mechanisms against intracellular pathogens. Importantly, infection with T. gondii considerably modulates apoptosis exerting both pro- and anti-apoptotic activities within its host. During acute infection, apoptosis is triggered in leukocytes including activated T cells and this diminishes the anti-parasitic immune response but also avoids overwhelming immunopathology. The induction of exaggerated leukocyte apoptosis by T. gondii, on the other hand, may completely abrogate anti-parasitic immunity leading to life-threatening disease. T. gondii also inhibits apoptosis and this either affects uninfected bystander cells or parasite-infected host cells. In order to develop within its intracellular niche, T. gondii may indeed rely on direct countermeasures to shut down apoptotic cascades that would normally be triggered as a cellular suicide program in parasite-infected cells. Apoptosis is, however, also indirectly inhibited in uninfected bystander cells, e.g. in macrophages and granulocytes thereby sustaining the inflammatory response during acute toxoplasmosis. Apoptosis and its complex modulation during infection with T. gondii, thus, have a crucial impact on the parasite-host interaction and the pathogenesis of disease.
Chapter 9: Pathogenicity and Virulence in Toxoplasma gondii
Sonya Taylor, Asis Khan, Chunlei Su, and L. David Sibley
Abstract
Toxoplasma gondii is a widespread protozoan parasite that causes opportunistic disease in humans. T. gondii has a complex life cycle that alternates between haploid asexually dividing forms that infect a variety of hosts, and a sexual phase, which occurs exclusively in the intestinal epithelial cells of the cat. Despite the potential for sexual recombination, T. gondii maintains a highly clonal population structure consisting of three clonal lineages that predominate in North America and Europe. This unusual population structure may result from the recent adaptation for asexual oral infectivity, which bypasses the cat and allows direct transmission between intermediate hosts. Notably, type I strains are distinguished by their high mortality in the mouse model. Forward genetic approaches have been developed to allow mapping of complex phenotypes based on linkage analysis. Acute virulence in the type I lineage is tightly linked to a major locus on chromosome VIIa. Identification of genes that regulate virulence and pathogenicity will be aided by the recently completed genome and the assembly of a genome map based on genetic linkage analysis.
Chapter 10: Cellular Response to Infection
Ashley Fouts and John C. Boothroyd
Abstract
The high prevalence, worldwide distribution, and ability to persist in the face of a competent immune system all indicate that Toxoplasma gondii is very well adapted to life within a host. As an obligate intracellular parasite, these adaptations necessarily include mechanisms to co-opt the host cell for its own purposes. Much of the phenomenology associated with the intracellular niche has been known for decades. In recent years, however, some of the molecules that mediate a sustained interaction between host and parasite have begun to be identified and the way in which they promote intracellular survival have likewise begun to be uncovered. In this chapter, we will review the recent findings in the area of host response. In addition, some of the methods that are just beginning to be developed and that seem most likely to give further detail on these processes will also be discussed. We will focus on the best-studied stage, the tachyzoite, but what information exists on the other stages will also be briefly summarized.
Chapter 11: Genetics and Genome Organization of Toxoplasma gondii
Asis Khan, Sonya Taylor, Chunlei Su, L. David Sibley, Ian Paulsen, and James W. Ajioka
Abstract
Toxoplasma gondii is a member of the phylum Apicomplexa, a diverse group of early branching eukaryotes related to dinoflagellates and ciliates (See Chapter 12; Baldauf, 2003). The definitive host is the cat where the sexual cycle takes place in the intestinal epithelia and oocysts are shed in the feces where meiosis occurs to produce environmentally resistant sporozoites (See Chapter 1). The life cycle is unusual compared to closely related coccidians in that the parasite can transmit directly between secondary hosts. This has allowed clonal growth and expansion in the population where the vast majority of isolates in North America and Europe are dominated by three clonal lineages (Howe and Sibley, 1995). Moreover, the predominance of the three clonal lineages appears to have emerged from a population bottleneck about 10,000 years ago (Su et al., 2003) where the lineages appear to be derived from just a few related ancestral strains (Grigg et al., 2001). T. gondii in South America appear to be more divergent but a greater analysis of isolates is required. Genetic mapping and sequence analysis reveals a ~ 65 MB genome distributed across 14 chromosomes which is over twice the size of Plasmodium falciparum (Khan et al., 2005). This difference is due to higher predicted gene content, lower gene density and more introns per gene. The genome annotation shows that compared to P. falciparum, T. gondii retains many more enzymes in carbohydrate, lipid, amino acid and nucleic acid metabolic pathways (See Chapters 19 and 20).
Chapter 12: Evolution and Comparative Genomics of Toxoplasma gondii
Jessica Kissinger and Chih-Horng Kuo
Abstract
especially unicellular organisms, like protists and bacteria, many of which have no fossil record. Sequence data have shown that Toxoplasma gondii groups with other apicomplexan parasites and not surprisingly is most closely related to other coccidia. Analyses of currently available sequence data have revealed an evolutionary history for the Apicomplexa that includes the endosymbiosis of an alga, giving rise to the apicoplast organelle found in T. gondii and many other (but not all) apicomplexan parasites. Comparative sequence analyses have also revealed the presence of horizontal gene transfers from diverse sources into the T. gondii nuclear genome. Analyses of T. gondii expressed sequence tags (ESTs) and preliminary nuclear gene content have revealed the largest and most diverse gene content of any apicomplexan examined thus far.
Chapter 13: Population Genetics, Sex and the Emergence of Clonal Lines of Toxoplasma gondii
Michael E. Grigg
Abstract
The population biology of the cosmopolitan parasite Toxoplasma gondii is highly dependent on the pathogen's ability to propagate both clonally and sexually. This highly prevalent zoonosis utilizes its sexual cycle to reassort and produce new lines capable of emerging from wild niches to expand its host range and cause outbreaks. Highly successful clones then rapidly spread clonally to endemic levels worldwide.
Chapter 14: Manipulating the Toxoplasma Genome
Marc-Jan Gubbels, Jolly Mazumdar, Giel van Dooren, and Boris Striepen
Abstract
The parasitic protozoan Toxoplasma gondii has many hallmarks of a robust genetic model system: straightforward continuous culture, a fast generation time, standard codon usage, efficient transient (30%) and stable (1-5%) transfection, and a haploid genome carrying mostly single copy genes which now has been fully sequenced. The parasite's microscopically well-defined subcellular structure also makes it highly suitable for cell biological studies. It is therefore no surprise that Toxoplasma has attracted a continuously growing research community. Toxoplasma serves as a model system for medically important yet experimentally less or not tractable apicomplexan parasites like Plasmodium and Cryptosporidium, respectively. Research on T. gondii has produced exciting progress in our understanding of how apicomplexan parasites invade and modify cells to suit their needs, how they grow and replicate within these cells, and how they subvert the host's counter measures. Work using the Toxoplasma model has also contributed significantly to more general problems in cell biology especially in the areas of motility and organelle evolution and biogenesis. This chapter reviews the genetic approaches and techniques that drive this progress.
Chapter 15: Cell Cycle Control/Parasite Division
Michael W. White, Jay R. Radke, Magnolia Conde de Felipe, and Margaret Lehmann
Abstract
The virulence and developmental competency of Toxoplasma vary significantly between strain types (Grigg et al., 2001; Howe and Sibley, 1995; Sibley and Boothroyd, 1992; Sibley and Howe, 1996). The molecular basis for these differences is mostly unknown, but control of the replication cycle, the ability to form tissue cysts, as well as distinct differences in migration or the ability to stimulate the immune response have all been implicated (Saeij et al., 2005). Parasite burden plays a primary role in pathogenesis and may result from fast growth, or indirectly, when parasites fail to switch to the slow growing (or growth arrested) bradyzoite form. Tachyzoites from Type I strains proliferate unchecked in mice (until the death of the animal) (Radke and White, 1999; Sibley et al., 2002) and cell culture (5-6 h doubling) (Jerome et al., 1998) and do not readily form bradyzoites. The growth of Type II and III parasites is less vigorous, but these strains typically demonstrate some degree of cell cycle control after infection that always leads to bradyzoite development and cyst formation. These observations link parasite replication to the pathogenesis of disease (Jerome et al., 1998), yet we know few of the molecular details that define this relationship. In this chapter, we will initially summarize the available "tool-kit" (Section 2) and reagents developed over the last decade to investigate the Toxoplasma cell cycle. We will then discuss the major features of the known tachyzoite cell cycle (Section 3.1) and describe the order and timing of organelle replication and daughter formation (Section 3.2). We then outline the evidence for checkpoint control of chromosomal replication (Section 4.0), and finally, discuss observations that link cell cycle control to parasite development (Section 5.0).
Chapter 16: Toxoplasma Gene Regulation and Bradyzoite Development
Stanislas Tomavo and Louis M. Weiss
Abstract
Due to its central importance in disease pathogenesis, the biology of stage differentiation has been an active area of research and this topic is reviewed in the current chapter. Tachyzoites can transform into bradyzoites and visa versa depending on the environmental (i.e. host) conditions. Investigations into bradyzoite biology and this differentiation event have been accelerated by the development of in vitro techniques to study and produce bradyzoites as well as by the genetic tools that exist for the manipulation of T. gondii. Bradyzoite differentiation is coupled with a slowing of the cell cycle. Differentiation is a programmed response as it proceeds in a reproducible fashion when host cells are infected with sporozoites leading to the formation of tachyzoites and then terminating with the formation of bradyzoites. This programmed response can be altered by changes in the environment. Unfortunately, the genetic triggers and sensors for this differentiation response have yet to be identified. The development of a bradyzoite is a stress mediated differentiation response that leads to metabolic adaptations. Transcription of a whole set of bradyzoite specific genes occurs during differentiation in a coordinated fashion. These gene products include metabolic enzymes, surface antigens, secretory antigens (including rhoptry proteins) and cyst wall components. Many of the features of this differentiation event also have elements reminiscent of epigenetic phenomena described in other systems.
Chapter 17: Biology of Bradyzoites
Florence Dzierszinski and Laura J. Knoll
Abstract
Formation and maintenance of the bradyzoite cyst is essential for the lifecycle of Toxoplasma gondii. To understand this critical stage, many aspects of bradyzoite biology must be examined. In this chapter, we consider how the bradyzoite adapts to an encysted lifestyle by discussing its morphology and metabolism. We also explore bradyzoite tissue cyst development in vivo by analyzing different parasite strains, mouse models, the influence of host genetic background, and bradyzoite cyst rupture with parasite reactivation. Finally, we consider the long-term survival of cysts and the relationship between bradyzoite biology and host immunity. This discussion includes tissue tropism, the SRS family of surface antigens, and immune recognition or diversion. While the bradyzoite stage is challenging to study, a wealth of fascinating information will be learned as we dissect out its secrets.
Chapter 18: Chromatin Remodeling
William J. Sullivan, Jr.
Abstract
Once relegated to a mere supporting role, chromatin has recently taken center stage in performing the regulation of gene expression. Broadly defined as a complex of proteins and nucleic acid, chromatin has stepped into the limelight as a novel control mechanism for eukaryotic gene transcription. It has even been postulated that an epigenetic "code" emerges from the accumulation of certain patterns of post-translational modifications made to chromatin proteins. This chapter includes a primer on chromatin constitution and the enzymatic complexes capable of restructuring it in ways influencing gene expression. I will highlight some of the conserved, as well as the unique features, of chromatin remodeling in Toxoplasma gondii, with particular attention paid to how these processes orchestrate changes in the expressed genome that are pertinent to the parasite life cycle.
Chapter 19: Lipid Synthesis and Uptake
Isabelle Coppens
Abstract
Acquiring appropriate amounts of suitable lipid species to proper cell compartments is imperative for maintaining the various functions of biological membrane systems. During the intracellular development of Toxoplasma gondii, many membrane-containing organelles including the nucleus, endoplasmic reticulum, Golgi apparatus, mitochondria, apicoplast, dense granules, rhoptries, micronemes, acidocalcisomes, pellicular complex, and the ever-enlarging parasitophorous vacuolar membrane, are formed. Elaborate lipid metabolism and trafficking are required for T. gondii replication and persistence in their mammalian hosts. These parasites meet their high demand for the necessary lipid species through synthesis from metabolites produced de novo as well as through diversion of prefabricated molecules from host exogenous sources. Unique peculiarities in lipid biosynthetic pathways and in mechanisms of lipid trafficking from host cells to intravacuolar T. gondii are likely to provide us with important new therapeutic discoveries in the future.
Chapter 20: Nucleotides and Amino Acids
Barbara A. Fox, Kshitiz Chaudhary, and David J. Bzik
Abstract
Nucleotides and amino acids are of fundamental importance in the replication and development of Toxoplasma gondii. Nucleotides provide the key cellular energy source (ATP and GTP), and are particularly essential to the synthesis of DNA (dATP, dGTP, dCTP, dTTP) and the synthesis of RNA (dATP, dGTP, dCTP, dUTP) in rapidly replicating tachyzoites. Nucleotides are the precursors of more complex molecules such as folate. They also provide regulatory roles as intracellular second messengers such as cAMP, provide nucleotide based enzyme cofactors (NAD+, FMN, FAD), and control metabolic and gene regulation. In T. gondii, nucleotides are synthesized from small molecules and amino acids via pyrimidine biosynthetic pathways, and also are acquired from preformed host nucleobases and nucleosides via salvage pathways. The importance of nucleotide metabolism to T. gondii is illustrated by noting that several current clinical strategies for treating toxoplasmosis in humans are based on blocking the accumulation of nucleotides. The study of nucleotide and amino acid metabolism in T. gondii is a challenging area of biology due to the complexity of these parasite pathways as well as the obligatory presence of the mammalian host cell, which also possesses complex and active pathways in nucleotide and amino acid metabolism. Consequently, genetic studies involving the examination of T. gondii mutants, or parasite behavior in mutant host cells, have provided particularly valuable information. The investigation of nucleotide and amino acid metabolism in T. gondii has also resulted in the establishment of several important genetic selection models that are based on aspects of parasite nucleotide and amino acid metabolism.
Chapter 21: Protein Targeting to the Apicoplast
Marilyn Parsons Amy DeRocher, and Jean Feagin
Abstract
The apicoplast is a relict chloroplast that is present in Toxoplasma gondii and many other apicomplexans. Cytologic and phylogenetic analyses indicate that this organelle evolved via a secondary endosymbiotic event which partnered the progenitor apicomplexan with an algal cell containing the chloroplast ancestor of the apicoplast. Distinct from primary plastids, which are enveloped by two membranes, the apicoplast is bounded by multiple membranes. This chapter discusses how proteins encoded in the parasite nuclear genome are targeted to the lumen of the apicoplast. The first step is specified by an N-terminal signal sequence that directs the protein into the endoplasmic reticulum. Cleavage of the signal sequence reveals a transit peptide that directs the protein to the apicoplast. Current data suggest that trafficking bypasses the Golgi, hence distinguishing the apicoplast from other destinations in the T. gondii secretory system.
Chapter 22: The Metabolic Functions of the Mitochondrion and the Apicoplast
Frank Seeber and Dominique Soldati
Abstract
T. gondii, like most Apicomplexa, possesses a single endosymbiotic mitochondrion and a plastid-derived organelle called the apicoplast both of which host conserved as well as unanticipated metabolic pathways. In this chapter we give a short overview of the current knowledge of those pathways that have either been experimentally characterized to operate in these compartments, or where solid bioinformatical evidence gives clear indications for their localization in the respective organelles. Besides well-known tasks like the citric acid cycle or oxidative phosphorylation the mitochondrion also harbors unexpected pathways like the methylcitrate cycle for the detoxification of propionate. The plastid-derived pathways of fatty acid, isoprenoid biosynthesis and partial haem biosynthesis are the known major metabolic tasks in the apicoplast, but auxiliary pathways for the generation of reducing equivalents or co-factors in this organelle have shown surprising deviations from previous knowledge. These maintained metabolic functions of these two organelles are coming to light as rich sources of potential targets for anti-parasitic drug discovery.
Chapter 23: Microneme Protein Repertoire and Function
M.-H. Huynh and V.B. Carruthers
Abstract
Contents of the microneme organelles play essential roles in the survival and propagation of T. gondii parasites. Microneme proteins (MICs) are involved in gliding motility, host cell invasion, and virulence in the mouse model of disease. Almost all MICs are comprised of multiple adhesive motifs, and the diversity of MICs results from combining different numbers and types of modules. The proper synthesis, folding, trafficking, targeting, secretion, and proteolytic processing of these proteins are all critically important in the lytic cycle of this parasite. This chapter will focus on the conserved motifs of the MICs, which may have implications for the broad host cell range of Toxoplasma. We will also discuss trafficking of MICs through the secretory pathway and their secretion onto the parasite surface, as well as the cooperative functions of these proteins and complexes in cell invasion and virulence.
Chapter 24: Proteomic Analysis of the Rhoptry Organelles of Toxoplasma gondii
J.M. Wastling and P.J. Bradley
Abstract
Rhoptry organelles are part of the defining features of the phylum Apicomplexa. The contents and function of the rhoptries have been the focus of considerable attention in an effort to understand this most important of all apicomplexan characteristics: the ability to invade a host cell. Toxoplasma gondii is able to parasitize a wide variety of cells, during which these specialised secretory organelles play a central role. The recent development of proteomic methods for T. gondii and the availability of genome sequence to underpin a proteomic database represent an unprecedented opportunity to characterise protein composition and function in this parasite. For the rhoptry organelles specifically, this has enabled the identification of a large number of novel proteins and the subsequent discovery of a completely new class of rhoptry proteins which localise to the neck of the organelle. This rapid expansion in our understanding of the composition of rhoptries has led to new insights into their structure and function and set the foundation for a more comprehensive understanding of how they contribute to the process of invasion and intracellular survival in this parasite.
Chapter 25: Dense Granules of the Infectious Stages of Toxoplasma gondii: Their Central Role in the Host/Parasite Relationship
Corinne Mercier, Marie-France Cesbron-Delauw and David J.P. Ferguson
Abstract
The infectious forms (tachyzoite, bradyzoite, merozoite, sporozoite) of Toxoplasma gondii contain a variable number of dense granules. These granules are part of the apical complex and certain of the dense granule proteins are believed to be involved in the modification and function of the parasitophorous vacuole. In this chapter, the dense granules and their proteins will be described and discussed in the different stages relatively to their sub-cellular location and possible functions throughout the parasite cycle. In addition, the importance of dense granule proteins in the host immune response as well as their potential uses in the design of both new diagnostic reagents and a possible vaccine against toxoplasmosis will be reviewed.
Chapter 26: Calcium Homeostasis and Acidocalcisomes in Toxoplasma gondii
Silvia N.J. Moreno, Kildare Miranda and Wanderley de Souza
Abstract
Calcium ion (Ca2+) is used as a major signaling molecule in Toxoplasma gondii. Ca2+ is critical for conoid extrusion, microneme secretion, gliding motility, and invasion of host cells, and its cytosolic concentration is regulated by the concerted operation of a number of transporters present in the plasma membrane, endoplasmic reticulum, mitochondria, and acidocalcisomes. Recent findings have shed light on the function of these transporters and the roles that they play in cellular metabolism and have shown that acidocalcisomes, electron-dense acidic organelles rich in calcium and polyphosphate, are linked to several functions, including polyphosphate metabolism, and calcium homeostasis in T. gondii.
Chapter 27: Tubulin, Microtubules and Microtubule-Associated Structures in Toxoplasma gondii
Naomi S. Morrissette
Abstract
Toxoplasma gondii uses tubulin to build both conventional structures (spindles and flagella) and unconventional structures (conoid fibers are a nine-protofilament tubulin ribbon). Toxoplasma microtubules determine parasite shape and apical polarity and are essential to the sexual cycle which relies upon flagellated male gametes. Unlike most eukaryotes, Toxoplasma tachyzoites contain two independent microtubule organizing structures: the apical polar ring organizes subpellicular microtubules and centrioles are associated with spindle microtubules. This chapter describes the distinct properties of Toxoplasma tubulin and the unique structures and functions that rely upon Toxoplasma microtubules.
Chapter 28: Molecular Motors
Bernardo J. Foth and Dominique Soldati
Abstract
Molecular motors are biological engines that convert chemical energy into movement. Three types of motor molecules - myosins, kinesins, and dyneins - are ubiquitously found in eukaryotes and serve a multitude of fundamental biological functions such as locomotion, nuclear and cell division, and the intracellular transport of molecules, vesicles, and organelles. The almost complete genome sequence of Toxoplasma gondii reveals a repertoire of 11 myosin heavy chains, 15 kinesin heavy chains, and 10 dynein heavy chains, making this parasite the apicomplexan with the largest known inventory of all three motor types known to date. Of these, only a few T. gondii myosins have been experimentally investigated: TgMyoA for example has been shown to be indispensable for the gliding motility of tachyzoites, a form of cell locomotion that does not involve cellular shape change or cell protrusions and that is essential for the spreading through host tissues and for host cell invasion and egress, whereas two other myosins (TgMyoB/C) may be involved in parasite cell division. Here, we present an overview over all molecular motor heavy chains that can be gleaned from the T. gondii genome and summarize what is known about the few molecular motors on which research has been published. For the majority of T. gondii motor molecules that have not yet been studied experimentally we focus on their classification into existing classes and families and discuss possible functions based on this classification and on apparent protein domain structures. We conclude that T. gondii probably employs its diverse arsenal of molecular motors both for general functions like nuclear and cell division, flagellar movement, and organellar transport, as well as for apicomplexan-specific adaptations to their lifestyle as intracellular parasites such as gliding motility.
Chapter 29: Actin Dynamics and Motility in Apicomplexans
Jennifer L. Gordon, Nivedita Sahoo, Simren Mehta, L. David Sibley
Abstract
Actin-based motility controls substrate-dependent gliding, tissue migration, and cell invasion by apicomplexan parasites. Although motility depends on the action of a small myosin motor, the polymerization of new actin filaments controls the timing, and speed of motility. Additionally, motility is directional, implying that actin filaments, which are normally polarized, must also be aligned with the longitudinal axis of the parasite. Actin dynamics in apicomplexans are highly unusual and the majority of actin is kept in an unpolymerized state in resting cells. Actin polymerization occurs permissively yet results in short filaments that are inherently unstable. The relatively small component of actin-binding proteins in apicomplexans attests to the streamlined nature of actin-based motility in these cells. Structural differences in apicomplexan actin may render it ideally suited for rapid cycles of assembly and disassembly, which likely represents an important adaptation for gliding motility. The concerted action of this actin-based motility system drives forward motion by retrograde translocation of cell surface adhesins. This unique form of motility is responsible for pathogenesis and presents numerous potential targets for therapeutic intervention.
Chapter 30: Proteases of Toxoplasma gondii
Artemio Jongco and Kami Kim
Abstract
The protozoan parasite Toxoplasma gondii is an obligate intracellular organism that survives in a variety of hosts. It undergoes a complex life cycle and has a highly specialized series of unique secretory organelles that enable its successful invasion of host cells. The study of proteases of T. gondii is relatively recent, but ongoing studies from several laboratories suggest that proteases perform critical functions during the development of the parasite. It appears that regulated proteolysis of the secreted products is essential for efficient invasion by T. gondii. Proteases are implicated in regulation of biogenesis of secretory organelles as well as the assembly and disengagement of adhesive complexes secreted by micronemes. Proteases are important for assembly of the cytoskeleton, targeting to the apicoplast or mitochondria and other essential physiological processes. Several unique aspects of proteolysis are emerging that may eventually facilitate development of novel chemotherapeutic agents.
Chapter 31: The Toxoplasma gondii Parasitophorous Vacuole Membrane: Transactions at the Parasite Host Interface
Angela M. Martin and Anthony P. Sinai
Abstract
The obligate intracellular protozoan Toxoplasma gondii establishes its replication permissive niche within the infected host cell. This niche, the parasitophorous vacuole (PV), is delimited from the host cell cytoplasm by the PV membrane (PVM). In this chapter we highlight the roles of the PVM in the remodeling of host cell architecture, nutrient acquisition, the manipulation of signaling and touch upon the potential roles in the parasite developmental cycle. We further present the PVM as a unique and dynamic "organelle" found only within the infected cell where it is established outside the parent organism. Despite its importance little is known about the biology of the PVM. There has, however, been a recent renewal of interest in the PVM, the study of which has become more tractable with the application of both classical approaches as well as genomic and proteomic analyses. With this chapter we discuss the diverse activities associated with the PVM and present pressing questions that remain to be elucidated regarding this enigmatic organelle.
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