16 research outputs found

    Widespread variation in transcript abundance within and across developmental stages of Trypanosoma brucei

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    <p>Abstract</p> <p>Background</p> <p><it>Trypanosoma brucei</it>, the causative agent of African sleeping sickness, undergoes a complex developmental cycle that takes place in mammalian and insect hosts and is accompanied by changes in metabolism and cellular morphology. While differences in mRNA expression have been described for many genes, genome-wide expression analyses have been largely lacking. Trypanosomatids represent a unique case in eukaryotes in that they transcribe protein-coding genes as large polycistronic units, and rarely regulate gene expression at the level of transcription initiation.</p> <p>Results</p> <p>Here we present a comprehensive analysis of mRNA expression in several stages of parasite development. Utilizing microarrays that have multiple copies of multiple probes for each gene, we were able to demonstrate with a high degree of statistical confidence that approximately one-fourth of genes show differences in mRNA expression levels in the stages examined. These include complex patterns of gene expression within gene families, including the large family of variant surface glycoproteins (VSGs) and their relatives, where we have identified a number of constitutively expressed family members. Furthermore, we were able to assess the relative abundance of all transcripts in each stage, identifying the genes that are either weakly or highly expressed. Very few genes show no evidence of expression.</p> <p>Conclusion</p> <p>Despite the lack of gene regulation at the level of transcription initiation, our results reveal extensive regulation of mRNA abundance associated with different life cycle and growth stages. In addition, analysis of variant surface glycoprotein gene expression reveals a more complex picture than previously thought. These data provide a valuable resource to the community of researchers studying this lethal agent.</p

    TriTrypDB: a functional genomic resource for the Trypanosomatidae

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    TriTrypDB (http://tritrypdb.org) is an integrated database providing access to genome-scale datasets for kinetoplastid parasites, and supporting a variety of complex queries driven by research and development needs. TriTrypDB is a collaborative project, utilizing the GUS/WDK computational infrastructure developed by the Eukaryotic Pathogen Bioinformatics Resource Center (EuPathDB.org) to integrate genome annotation and analyses from GeneDB and elsewhere with a wide variety of functional genomics datasets made available by members of the global research community, often pre-publication. Currently, TriTrypDB integrates datasets from Leishmania braziliensis, L. infantum, L. major, L. tarentolae, Trypanosoma brucei and T. cruzi. Users may examine individual genes or chromosomal spans in their genomic context, including syntenic alignments with other kinetoplastid organisms. Data within TriTrypDB can be interrogated utilizing a sophisticated search strategy system that enables a user to construct complex queries combining multiple data types. All search strategies are stored, allowing future access and integrated searches. ‘User Comments’ may be added to any gene page, enhancing available annotation; such comments become immediately searchable via the text search, and are forwarded to curators for incorporation into the reference annotation when appropriate

    Genome and phylogenetic analyses of Trypanosoma evansi reveal extensive similarity to T. brucei and multiple independent origins for dyskinetoplasty.

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    Two key biological features distinguish Trypanosoma evansi from the T. brucei group: independence from the tsetse fly as obligatory vector, and independence from the need for functional mitochondrial DNA (kinetoplast or kDNA). In an effort to better understand the molecular causes and consequences of these differences, we sequenced the genome of an akinetoplastic T. evansi strain from China and compared it to the T. b. brucei reference strain. The annotated T. evansi genome shows extensive similarity to the reference, with 94.9% of the predicted T. b. brucei coding sequences (CDS) having an ortholog in T. evansi, and 94.6% of the non-repetitive orthologs having a nucleotide identity of 95% or greater. Interestingly, several procyclin-associated genes (PAGs) were disrupted or not found in this T. evansi strain, suggesting a selective loss of function in the absence of the insect life-cycle stage. Surprisingly, orthologous sequences were found in T. evansi for all 978 nuclear CDS predicted to represent the mitochondrial proteome in T. brucei, although a small number of these may have lost functionality. Consistent with previous results, the F1FO-ATP synthase γ subunit was found to have an A281 deletion, which is involved in generation of a mitochondrial membrane potential in the absence of kDNA. Candidates for CDS that are absent from the reference genome were identified in supplementary de novo assemblies of T. evansi reads. Phylogenetic analyses show that the sequenced strain belongs to a dominant group of clonal T. evansi strains with worldwide distribution that also includes isolates classified as T. equiperdum. At least three other types of T. evansi or T. equiperdum have emerged independently. Overall, the elucidation of the T. evansi genome sequence reveals extensive similarity of T. brucei and supports the contention that T. evansi should be classified as a subspecies of T. brucei

    Diversity of a-type and b-type VSG between <i>T. b. brucei</i> TREU 927/4 and <i>T. evansi</i> STIB805 compared.

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    <p>Histograms showing a-type VSG (A.) or b-type VSG (B.) distributions of strain-specific clade size in <i>T. b. brucei</i> (black bars) and <i>T. evansi</i> (red bars) as defined by the phylogeny (see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003404#pntd.0003404.s010" target="_blank">S10 Fig</a>.). Frequency distributions of a-type VSG (C.) or b-type VSG (D.) synonymous (<i>Ks</i>) and non-synonymous (<i>Ka</i>) substitution rates per site, and the ω (<i>Ka/Ks</i>) for orthologous pairs of <i>VSG</i> (a-type n = 151; b-type n = 112), as defined by the phylogeny, in relation to values for unambiguous non-VSG orthologous pairs (n = 6331).</p

    Bayesian phylogeny of <i>Trypanozoon</i> isolates based on the dihydrolipoamide dehydrogenase gene (LipDH; Tb927.11.16730).

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    <p>Panel A shows a mid-point rooted tree based on an alignment of 32 unique LipDH haplotypes, assembled from sequences derived from 13 <i>T. b. brucei</i> (Tbb), 3 <i>T. b. gambiense</i> type 1 (Tbg1), 4 <i>T. b. rhodesiense</i> (Tbr), 15 <i>T. evansi</i> (Tev) and 5 <i>T. equiperdum</i> (Teq) samples. Scale units for the phylogeny are substitutions per site. The chart illustrates the distribution of each haplotype among samples from each <i>Trypanozoon</i> taxon. Phylogenetic analysis grouped all but three of the haplotype sequences into one of five major clusters with strong support (posterior probabilities ≥0.9), which are referred to as clades V, W, X, Y and Z. Eight unique Tev/Teq genotypes were found, as summarized in panel B. Discounting minor sequence differences (1–2 SNPs) these were reduced to four major genotypes based on the position of haplotypes in the tree, which mirrored the type of mutation (A281Δ, M282L, A273P, WT) in the C-termini of the ATP synthase subunit γ in these the samples.</p

    Pulse-field gel electrophoresis comparing chromosomes of <i>T. evansi</i> STIB805 (Tev) and <i>T. b. brucei</i> TREU 927/4 (927).

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    <p>While the sizes of megabase chromosomes are largely similar, differences in the intermediate and minichromosomes (825 kbp and smaller) are evident between <i>T. evansi</i> and <i>T. brucei</i>. Although <i>T. brucei</i> chromosomes I-XI were not unambiguously identified, labels to the left of the gel indicate bands consistent with the expected <i>T. brucei</i> chromosome sizes <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003404#pntd.0003404-Melville2" target="_blank">[46]</a>, as well as intermediate and minichromosomes. The signal at the top of the gel is from the well, as indicated.</p
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