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Mathematical modeling of Trypanosoma brucei transcription

ABSTRACT: Trypanosome RNA polymerase II transcription is polycistronic, individual mRNAs being excised by trans splicing and polyadenylation. In this study, we refined the previously published mathematical model for bloodstream form parasites and extended it to the procyclic form. We used the model, together with known mRNA half-lives, to predict the abundances of individual mRNAs, assuming rapid, unregulated mRNA processing; then we compared the results with measured mRNA abundances. Remarkably, the abundances of most mRNAs in procyclic forms are predicted quite well by the model, being largely explained by variations in mRNA decay rates and length. In bloodstream forms substantially more mRNAs are less abundant than predicted. We list mRNAs that are likely to show particularly slow or inefficient processing, either in both forms or with developmental regulation. We also measured ribosome occupancies of all mRNAs in trypanosomes grown in the same conditions as were used to measure mRNA turnover. In procyclic forms there was a weak positive correlation between ribosome density and mRNA half-life, suggesting cross-talk between translation and mRNA decay; ribosome density was related to the proportion of the mRNA on polysomes, indicating control of translation initiation. Ribosomal protein mRNAs in procyclics appeared to be exceptionally rapidly processed but poorly translated. Through this study, we conclude that lLevels of mRNAs in procyclic form trypanosomes are determined mainly by length and mRNA decay, with some control of precursor processing. In bloodstream forms variations in nuclear events play a larger role in transcriptome regulation, suggesting acquisition of new control mechanisms during adaptation to mammalian parasitism.

ORGANISM(S): Trypanosoma brucei

PROVIDER: GSE79208 | GEO | 2016/03/16

REPOSITORIES: GEO

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Project description:Trypanosoma brucei subspecies infect humans and animals in sub-Saharan Africa. This early diverging eukaryote shows many novel features in basic biological processes, including the use of polycistronic transcription to generate all protein-coding mRNAs. Therefore we hypothesized that translational control provides a means to tune gene expression during parasite development in mammalian and fly hosts. We used ribosome profiling to examine genome-wide protein production in animal-derived slender bloodstream forms and cultured procyclic (insect midgut) forms. About one-third of all CDSs showed statistically significant regulation of protein production between the two stages. Of these, more than two-thirds showed a change in translation efficiency, but few were controlled by this parameter alone. Ribosomal proteins were translated poorly, especially in animal-derived parasites. A disproportionate number of metabolic enzymes were up-regulated at the mRNA level in procyclic forms, as were variant surface glycoproteins in bloodstream forms. Comparison with cultured bloodstream forms from another strain identified stage-specific changes in protein production that transcend strain and growth conditions. Genes with upstream ORFs had a lower mean translation efficiency but no evidence was seen for involvement of these ORFs in stage-regulation. Ribosome profiling revealed that differences in the production of specific proteins in T. brucei slender bloodstream and procyclic forms are more common than anticipated from analysis of mRNA abundance. While in vivo and in vitro derived slender bloodstream forms of different strains are more similar to one another than to procyclic forms, they showed numerous differences at both the mRNA and protein production level. Ribosome profiling and mRNA libraries were constructed in triplicate from in vitro PCF and in vivo BF lifestages of theT. brucei Treu927 and in vitro T. brucei Lister427, to evaluate role of translational gene regulation

Project description:The infectious metacyclic forms of Trypanosoma brucei result from a complex development in the tsetse fly vThe infectious metacyclic forms of Trypanosoma brucei result from a complex development in the tsetse fly vector. When they infect mammals, they cause African sleeping sickness in humans. Due to scarcity of biological material and difficulties of the tsetse fly as an experimental system, very limited information is available concerning the gene expression profile of metacyclic Trapanosoma forms. We used an in vitro system based on expressing the RNA binding protein 6 (RBP6) to obtain infectious metacyclics and determined their protein and mRNA repertoires by mass-spectrometry (MS) based proteomics and mRNA sequencing (RNAseq) in comparison to non-infectious procyclic trypanosomes. This comparison showed that metacyclics are quiescent cells, and we propose this influences the choice of a monocistronic variant surface glycoprotein expression site. Metacyclics have a largely bloodstream-form type transcriptome, and thus are programmed to translate a bloodstream-form type proteome upon entry into the mammalian host and resumption of cell division. Genes encoding cell surface components showed the largest changes between procyclics and metacyclics, observed at both the transcript and protein levels. Genes encoding metabolic enzymes exhibited expression in metacyclics with features of both procyclic and bloodstream forms, suggesting that this intermediate-type metabolism is dictated by the availability of nutrients in the tsetse fly vector. ector. When they infect mammals, they cause African sleeping sickness in humans. Due to scarcity of biological material and difficulties of the tsetse fly as an experimental system, very limited information is available concerning the gene expression profile of metacyclic Trapanosoma forms. We used an in vitro system based on expressing the RNA binding protein 6 (RBP6) to obtain infectious metacyclics and determined their protein and mRNA repertoires by mass-spectrometry (MS) based proteomics and mRNA sequencing (RNAseq) in comparison to non-infectious procyclic trypanosomes. This comparison showed that metacyclics are quiescent cells, and we propose this influences the choice of a monocistronic variant surface glycoprotein expression site. Metacyclics have a largely bloodstream-form type transcriptome, and thus are programmed to translate a bloodstream-form type proteome upon entry into the mammalian host and resumption of cell division. Genes encoding cell surface components showed the largest changes between procyclics and metacyclics, observed at both the transcript and protein levels. Genes encoding metabolic enzymes exhibited expression in metacyclics with features of both procyclic and bloodstream forms, suggesting that this intermediate-type metabolism is dictated by the availability of nutrients in the tsetse fly vector.

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Mathematical modeling of Trypanosoma brucei transcription

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