Trypanosomes: After The Genome Chapter Abstracts
Chapter 1
The Genome of Trypanosoma brucei
Christiane Hertz-Fowler, Hubert Renauld and Matthew Berriman
Abstract
July 2005 saw the culmination of a decade-long effort to unravel not only the genome sequence of Trypanosoma brucei, causative agent of human sleeping sickness and livestock trypanosomiasis in sub-Saharan Africa, but also the genomes of the related species Leishmania major and Trypanosoma cruzi. Not only has the genome told us a lot about the biology of the parasite, but - owing to an early data release policy - has also changed the way many research laboratories with an interest in the order Kinetoplastida approach their research. Here we outline how the sequencing project was undertaken and describe some of the major findings gleaned from the individual and comparative analyses of all three genome sequences.
Chapter 2
Reverse and Forward Genetics as Practical Approaches for Post-genome Studies
James C. Morris, Meredith T. Morris, Sylvia Y. Lee, William P. Toole, Clarice M. Seifert, and Alvaro Acosta-Serrano
Abstract
With the completion of the Trypanosoma brucei genome-sequencing project, the next major focus will be elucidation of the function of the gene complement. Here we discuss tools that have been developed for the characterization of single genes as well techniques for genome-wide assessment of gene function. The successful development of molecular tools for both reverse and forward genetics, as well as emerging technologies for the exploration of genome-wide gene expression, have led to the rise of the African trypanosome as a model genetic organism for the study of complex pathways, including development, pathogenesis, and signaling.
Chapter 3
The System of Genetic Exchange in Trypanosoma brucei and other Trypanosomatids
Annette MacLeod, Mike Turner and Andy Tait
Abstract
In this chapter, we discuss our current understanding of the systems of genetic exchange in trypanosomatids and the impact the recent genome projects have had on this area of research. We focus mainly on the details of Trypanosoma brucei as it is the most extensively studied of the 'trityps', but will also refer to a recently discovered novel mechanism of genetic exchange in T. cruzi and the apparent rarity of genetic exchange in Leishmania sp. The system of genetic exchange in Trypanosoma brucei has been known to exist since the late eighties when a genetic cross between different strains was carried out by co-transmission through the tsetse fly. We discuss the segregation of nuclear, chromosomal and kDNA markers and outline the two current models for the mechanism of genetic exchange. We also present how the completion of the genome project has allowed the identification of polymorphic micro and minisatellite markers distributed throughout the genome, which have been used to prove formally that meiosis, independent assortment and crossing over occur in this parasite, as would be predicted in a conventional Mendelian system. Such data have been used to construct the first genetic map of T. brucei, which opens up the use of genetic analysis, coupled with positional cloning and the genome sequence, as a tool to identify the genes involved in a range of traits relevant to the disease.
Chapter 4
Chromosome Structure and Dynamics
Hubert Renauld, John M. Kelly and David Horn
Abstract
In this chapter, we discuss the impact of the trypanosomatid genome sequencing projects on our understanding of chromosome structure and dynamics in these parasites. Focussing on the trypanosomes, Trypanosoma brucei and Trypanosoma cruzi, but with reference to the related kinetoplastid, Leishmania major, we first present an overview of the organisation of protein-coding and non-coding elements along the chromosomes. The discussion then progresses to the array of factors revealed by genome sequencing that are likely to influence chromosome dynamics during mitosis, but also more generally, through chromatin packaging and regulation. Finally, we present a forward view, detailing the types of experiments and research themes likely to reveal how chromosomes are organised and segregated.
Chapter 5
The Three R's of the Trypanosomatid Genomes: Replication, Recombination and Repair
Michele M. Klingbeil, Peter Burton, Rebecca Barnes and Richard McCulloch
Abstract
Replication, recombination and repair are universal processes that allow for the dissemination, rearrangement and maintenance of genomes. Each process is catalysed by complex protein machineries, which in some organisms appear to have been elaborated to provide overlapping activities that encode multiple, partially redundant, pathways. In addition, each of the three processes affect one another, either by competition for substrates or by acting together. To date, our understanding of the genes encoding replication, recombination and repair functions in Trypanosoma brucei and related trypanosomatids has been incomplete, despite the fact that they act in crucial processes such as antigenic variation of surface glycoproteins and maintenance of the highly unusual kinetoplast DNA network. The sequencing and annotation of the T. brucei, T. cruzi and L. major genomes represents a major step forward in this area. This chapter reviews the current understanding of the replication, recombination and repair machinery and considers how much more complete a picture of these crucial processes can now be drawn, as well as the impact that the sequencing projects have had toward understanding the unusual features of trypanosomatids that are a departure from typical eukaryotic biology.
Chapter 6
Transcription in Trypanosomes: A Different Means to the End
Arthur Günzl, Luc Vanhamme, and Peter J. Myler
Abstract
The recent publication of T. brucei genome has enabled a new wave of molecular and genetic experiments, which have resulted in a clearer picture of the transcription machinery in this organism. While trypanosomes possess most of the subunits common to the three RNA polymerases typically found in eukaryotes, a number of interesting differences exist, presumably reflecting their early divergence from the eukaryote crown lineage. Most notably, the absence of many class-specific RNAP subunits, and paucity of general transcription factors, suggests that the intricate, and varied, mechanisms for regulation of transcript initiation in higher eukaryotes arose after this divergence. It appears that the Tritryp transcriptional apparatus may be the most primitive yet identified in the Eukarya, containing perhaps a single shared set of GTFs for transcription initiation. Indeed, the RNAP I and II promoters described in T. brucei share structural similarities with the RNAP III type III promoters of higher eukaryotes, and some protein-coding genes are transcribed by RNAP I. The bi-directional RNAP II-mediated transcription initiation regions separating the long polycistronic clusters of protein-coding genes in trypanosomatids may represent ancestral, less sequence-specific, promoters mostly replaced in other eukaryotes by the archetypal TATA-containing promoters. Unlike most other organisms where transcriptional mechanisms plays a major role in regulation of mRNA levels, control of mRNA abundance in trypanosomes occurs mainly at the steps of RNA processing, stability and degradation. This reliance on posttranscriptional regulation of gene expression may explain the relative dearth of transcription factors and over-representation of RNA-binding proteins in the trypanosome genome.
Chapter 7
Post-Transcriptional Control of Gene Expression in African Trypanosomes
Edward F. Hendriks and Keith R. Matthews
Abstract
In trypanosomes genes are organised into polycistronic transcription units in which adjacent genes show differential patterns of expression. This genome organisation dictates that regulation operates primarily at the posttranscriptional level. This entails many potential regulatory steps, including pre-mRNA processing, mRNA stability, regulated control of mRNA translation and protein turnover, post translational modification, etc. In particular the genome of Trypanosoma brucei is rich in RNA binding proteins whose function is likely to be important in governing the patterns of gene expression characteristic of the different life cycle stages of the parasite. To date there is detailed experimental information on only a few regulated genes with the relative importance of the different steps in gene expression control still not fully clear for the majority of regulated genes. Moreover, the precise mechanisms by which the large set of regulatory RNA binding proteins effect gene expression in trypanosomes is almost completely unknown, this often being compounded by their lack of close homology to RNA binding proteins in other organisms. In this chapter, the experimental evidence for posttranscriptional gene expression control is reviewed and potential molecular components involved in this divined using the recently completed genome sequence data. This provides a blueprint of the components likely to be involved in controlling gene expression. The interactions between particular RNA binding proteins, their RNA targets and the regulatory machineries involved remain to be understood.
Chapter 8
Cell Structure, Cell Division and Cell Cycle
Tansy C. Hammarton, Bill Wickstead and Paul G. McKean
Abstract
In this chapter we consider how the completion of the T. brucei genome sequencing project has furthered our molecular understanding of trypanosome cell structure, cell division processes and cell cycle regulatory mechanisms. Analysis of the 'in silico trypanosome' is highly informative, both in identifying structures/pathways conserved in other eukaryotes, but also in discerning significant omissions in the genetic repertoire of the African trypanosome. However, the large number of genes that are trypanosomatid specific (many of which may have cytoskeletal/cell cycle regulatory roles) emphasises the enormous challenges that lie ahead for the trypanosome field. In order to place this in silico analysis into a biological framework, we begin by providing a detailed description of the T. brucei cell and outline key morphogenetic events that occur during the cell cycle. It is worth mentioning at the outset that although some of these descriptions are based on cytological evidence pre-dating the genome sequencing projects, most of the recent studies cited below benefited significantly from earlier releases of T. brucei sequence information.
Chapter 9
Intracellular Transport Systems in Trypanosomes: Function, Evolution and Virulence
Markus Engstler, James D. Bangs and Mark C. Field
Abstract
The surface of pathogenic trypanosomatids represents the host-parasite interface, and characterization of the surface molecules and their functions has yielded major insights into mechanisms of disease progression, immune modulation and immune evasion. Additionally trypanosomatids place a particularly heavy emphasis onto use of the GPI-anchor as a means for membrane attachment of proteins, glycoconjugates and glycolipids. Synthesis, degradation and maintenance of the surface are a function of the endomembrane system. Recent advances have begun to unravel the complexity of the trypanosome trafficking system, how it is regulated and the mechanisms that underpin protein sorting. Availability of complete genome data now allows molecular level comparisons between the TriTryps and their hosts, identifying unique aspects to trafficking pathways in trypansomes, as well as providing potential for more rational experimental manipulation of transport systems. These aspects together with our current state of knowledge of the trypanosome trafficking system are discussed.
Chapter 10
Comparison and Evolution of the Surface Architecture of Trypanosomatid Parasites
Alvaro Acosta-Serrano, Clyde Hutchinson, Ernesto S. Nakayasu, Igor C. Almeida, and Mark Carrington
Abstract
The surface glycocalyx of Trypanosoma and Leishmania parasites is composed of a variety of abundant glycosylphosphatidylinositol (GPI)-anchored glycoconjugates. The valuable information released by the different genome projects has allowed the identification of novel families of surface proteins as well as the identification of new members of already described families of surface molecules. The recent development of powerful genetic tools has allowed researchers to determine the biological importance of these molecules especially in relation to virulence and adaptability to different host environments. In this review, we discuss what influence has the genome sequences had on our knowledge of the molecular architecture of these cell surfaces and how each of them may have evolved.
Chapter 11
Antigenic Variation in Trypanosoma brucei
Etienne Pays, Didier Salmon, Liam J. Morrison, Lucio Marcello and J. David Barry
Abstract
During its life cycle, Trypanosoma brucei is continuously covered with a homogeneous and dense protein coat. In the bloodstream form this coat is made of Variant Surface Glycoprotein (VSG), which triggers efficient antibody responses but is changed repeatedly, thereby prolonging infection. In the insect stage procyclic form the coat is made of another glycoprotein termed procyclin, several isoforms of which are expressed sequentially. Provision of the genome sequence has advanced our views on the processes, controls and biological significance of these changes of surface coats. In particular, the identification now of a significant fraction of the VSG gene archive has allowed better understanding of the programming and variability of the antigen repertoire of the parasite, and has highlighted the importance of segmental gene conversion in these processes. The genome sequence has also added to our understanding of the crucial involvement of the telomeric VSG expression sites in the generation of VSG-based proteins associated with adaptation of the parasite to different hosts.
Chapter 12
Comparative Genomics of Trypanosome Metabolism
Michael L. Ginger, Alan H. Fairlamb and Fred R. Opperdoes
Abstract
Over the last fifty years, the metabolism of Trypanosoma brucei and Trypanosoma cruzi has been the subject of many extensive biochemical investigations, but the recent completion of two trypanosome genome sequencing projects now provides a more complete insight in the full metabolic capacities of these two trypanosomatids. The genes required for various pathways of carbohydrate metabolism, including glycolysis and the hexose-monophosphate pathway, are all present, but a comparison between African and American trypanosomes indicates that only T. cruzi contains a considerable number of genes encoding bacterial-type kinases with predicted specificity for various sugars, other than glucose. The enzymes encoded by these genes all contain targeting signals for import into the glycosomes, suggesting that in American trypanosomes the glycosomes have adapted to the breakdown of a wide variety of sugars. In T. brucei the carbohydrates that can provide carbon for energy metabolism are likely to be limited to glucose, fructose and mannose. No evidence in either trypanosome species was found for the presence of either a functional glyoxylate cycle or uric acid cycle. Both Trypanosoma species are capable of synthesising and oxidising fatty acids. Consistent with previous biochemical studies, the capacity of trypanosomes for general lipid synthesis is, in comparison with many parasitic protozoa, impressive. Most amino acids, apart from the aromatic ones, can be oxidised by the two organisms, but amino-acid synthesis is generally limited to the so-called non-essential amino acids. T. cruzi, but not T. brucei, is able to utilise histidine as an energy source. The assembly of the mitochondrial respiratory chain, which is required for oxidative phosphorylation and efficient metabolism of amino acid and fatty acid carbon sources, appears to be a balance between conserved and unique biochemical processes. Finally, the presence of numerous genes of bacterial ancestry indicates that horizontal gene transfer has played an important role in shaping the trypanosomatids metabolic capacities.
Current Books: