Dataset Information

Identification of positive and negative regulators in the stepwise developmental progression towards infectivity in Trypanosoma brucei.

ABSTRACT: Trypanosoma brucei is a protozoan parasite that causes important human and livestock diseases in sub-Saharan Africa. By overexpressing a single RNA-binding protein, RBP6, in non-infectious procyclics trypanosomes, we previously recapitulated in vitro the events occurring in the tsetse fly vector, namely the development of epimastigotes and infectious, quiescent metacyclic parasites. To identify genes involved in this developmental progression, we individually targeted 86 transcripts by RNAi in the RBP6 overexpression cell line and assessed the loss-of-function phenotypes on repositioning the kinetoplast, an organelle that contains the mitochondrial genome, the expression of BARP or brucei alanine rich protein, a marker for epimastigotes, and metacyclic variant surface glycoprotein. This screen identified 22 genes that positively or negatively regulate the stepwise progression towards infectivity at different stages. Two previously uncharacterized putative nucleic acid binding proteins emerged as potent regulators, namely the cold shock domain-containing proteins CSD1 and CSD2. RNA-Seq data from a selected group of cell lines further revealed that the components of gene expression regulatory networks identified in this study affected the abundance of a subset of transcripts in very similar fashion. Finally, our data suggest a considerable overlap between the genes that regulate the formation of stumpy bloodstream form trypanosomes and the genes that govern the development of metacyclic form parasites.

SUBMITTER: Toh JY

PROVIDER: S-EPMC7952579 | biostudies-literature | 2021 Mar

REPOSITORIES: biostudies-literature

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Identification of positive and negative regulators in the stepwise developmental progression towards infectivity in Trypanosoma brucei.

Toh Justin Y JY Nkouawa Agathe A Sánchez Saúl Rojas SR Shi Huafang H Kolev Nikolay G NG Tschudi Christian C

Scientific reports 20210311 1

Trypanosoma brucei is a protozoan parasite that causes important human and livestock diseases in sub-Saharan Africa. By overexpressing a single RNA-binding protein, RBP6, in non-infectious procyclics trypanosomes, we previously recapitulated in vitro the events occurring in the tsetse fly vector, namely the development of epimastigotes and infectious, quiescent metacyclic parasites. To identify genes involved in this developmental progression, we individually targeted 86 transcripts by RNAi in t ...[more]

PMID: 33707699

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Project description:Trypanosoma brucei ssp., unicellular parasites causing human and animal trypanosomiasis, are transmitted between mammals by tsetse flies. Periodic changes in variant surface glycoproteins (VSG), which form the parasite coat in the mammal, allow them to evade the host immune response. Different isolates of T. brucei show heterogeneity in their repertoires of VSG genes and have single nucleotide polymorphisms and indels that can impact on genome editing. T. brucei brucei EATRO1125 (AnTaR1 serodeme) is an isolate that is used increasingly often because it is pleomorphic in mammals and fly transmissible, two characteristics that have been lost by the most commonly used laboratory stocks. We present a genome assembly of EATRO1125, including contigs for the intermediate chromosomes and minichromosomes that serve as repositories of VSG genes. In addition, de novo transcriptome assemblies were performed using Illumina sequences from tsetse-derived trypanosomes. Reads of 150 bases enabled closely related members of multigene families to be discriminated. This revealed that the transcriptome of midgut-derived parasites is dynamic, starting with the expression of high affinity hexose transporters and glycolytic enzymes and then switching to proline uptake and catabolism. These changes resemble the transition from early to late procyclic forms in culture. Further metabolic reprogramming, including upregulation of tricarboxylic acid cycle enzymes, occurs in the proventriculus. Many transcripts upregulated in the salivary glands encode surface proteins, among them 7 metacyclic VSGs, multiple BARPs and GCS1/HAP2, a marker for gametes. A novel family of transmembrane proteins, containing polythreonine stretches that are predicted to be O-glycosylation sites, was also identified. Finally, RNA-Seq data were used to create an optimised annotation file with 5' and 3' untranslated regions accurately mapped for 9302 genes. We anticipate that this will be of use in identifying transcripts obtained by single cell sequencing technologies.

Dataset Information

Identification of positive and negative regulators in the stepwise developmental progression towards infectivity in Trypanosoma brucei.

Publications

Identification of positive and negative regulators in the stepwise developmental progression towards infectivity in Trypanosoma brucei.

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