The parasite
Trypanosoma brucei
is the causative agent of human and animal African trypanosomiasis (sleeping sickness). Trypanosomiasis, which affects humans and cattle, is fatal if untreated.
The alternative sigma factor RpoS regulates transcription of over 1000 genes in Escherichia coli in response to many different stresses. RpoS levels rise continuously after exposure to stress, and the consequences of changing levels of RpoS for the temporal patterns of expression of RpoS-regulated genes has not been described. We measured RpoS levels at various times during the entry to stationary phase, or in response to high osmolarity or low temperature, and found that the time required to reach maximum levels varied by several hours. We quantified the transcriptome across these stresses using RNA-seq. The number of differentially expressed genes differed among stresses, with 1379 DE genes were identified in in stationary phase, 633 in high osmolarity, and 302 in cold shock. To quantify the timing of gene expression, we fit sigmoid or double sigmoid models to differentially expressed genes in each stress. During the entry into stationary phase, genes whose expression rose earlier tended to be those that had been found to respond most strongly to low levels of RpoS. The timing of individual genes' expression was not correlated across stresses. Taken together, our results demonstrate E. coli activates RpoS with different timing in response to different stresses, which in turn generates a unique pattern of timing of the transcription response to each stress.
The eukaryotic protozoan parasite Trypanosoma brucei, spp. is transmitted by the tsetse fly to both humans and animals, where it causes a fatal disease called African trypanosomiasis. While the parasite lacks canonical DNA sequence specific transcription factors, it does possess histones, histone modifications, and proteins that write, erase, and read histone marks. Chemical inhibition of chromatin interacting bromodomain proteins has previously been shown to perturb bloodstream specific trypanosome processes, including silencing of the Variant Surface Glycoprotein genes (VSGs) and immune evasion. Transcriptomic changes that occur in bromodomain inhibited bloodstream parasites mirror many of the changes that occur as parasites developmentally progress from the bloodstream to the insect stage. We performed RNA-seq timecourses to determine the effects of chemical bromodomain inhibition in insect stage parasites using the compound I-BET151. We found that treatment with I-BET151 causes large changes in the transcriptome of insect stage parasites, and also perturbs silencing of VSG genes. The transcriptomes of bromodomain inhibited parasites share some features with early metacyclic stage parasites in the fly salivary gland, implicating bromodomain proteins as important for regulating transcript levels for developmentally relevant genes. However, the downregulation of surface procyclin protein that typically accompanies developmental progression is absent in bromodomain inhibited insect stage parasites. We conclude that chemical modulation of bromodomain proteins causes widespread transcriptomic changes in multiple trypanosome life cycle stages. Understanding the gene regulatory processes that facilitate transcriptome remodeling in this highly diverged eukaryote may shed light on how these mechanisms evolved.
The disease African trypanosomiasis imposes a severe human and economic burden for communities in sub-Saharan Africa. The parasite that causes the disease is transmitted to the bloodstream of a human or ungulate via the tsetse fly.
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