Dataset Information

The mitogen-activated protein kinase kinase 5 regulates the proliferation and biosynthetic processes in procyclic forms of Trypanosoma brucei

ABSTRACT: The pathogenic protozoan T. brucei alternates into distinct developmental stages in the mammalian and insect hosts. The mitogen-activated protein kinase (MAPK) signaling pathways transduce extracellular stimuli into a range of cellular responses, which ultimately lead to the adaptation to the external environment. Here, we combined a loss of function approach with stable isotope labelling with amino acids in cell culture (SILAC)-based mass spectrometry (MS) to investigate the role of the mitogen-activated kinase kinase 5 (MKK5) in T. brucei. The silencing of MKK5 significantly decreased the proliferation of procyclic forms of T. brucei. To shed light into the molecular alterations associated with this phenotype, we measured the total proteome and phosphoproteome of cells silenced for MKK5. In the total proteome, we observed a general decrease in proteins related to ribosome and translation as well as down-regulation of several components of the fatty acid biosynthesis pathway. In addition, we observed alterations in the protein levels and phosphorylation of key metabolic enzymes, which point toward a suppression of the oxidative metabolism. Taken together, our findings show that the silencing of MKK5 alters cell growth, energy metabolism, protein and fatty acid biosynthesis in procyclic T. brucei.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Trypanosoma Brucei

SUBMITTER: Fernanda Kugeratski

LAB HEAD: Fabricio Marchini

PROVIDER: PXD007910 | Pride | 2017-10-20

REPOSITORIES: Pride

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			Action	DRS
	141016_SEL_HF_FEK_R23_141016_Phospho_D_001.raw	Raw
	141016_SEL_HF_FEK_R23_141016_Phospho_D_002.raw	Raw
	141016_SEL_HF_FEK_R23_141016_Phospho_D_003.raw	Raw
	141016_SEL_HF_FEK_R23_141016_Phospho_E_001.raw	Raw
	141016_SEL_HF_FEK_R23_141016_Phospho_E_002.raw	Raw

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Publications

Mitogen-Activated Protein Kinase Kinase 5 Regulates Proliferation and Biosynthetic Processes in Procyclic Forms of Trypanosoma brucei.

Kugeratski Fernanda G FG Batista Michel M Lima Carla V de Paula CVP Neilson Lisa J LJ da Cunha Elizabeth Sousa ES de Godoy Lyris M LM Zanivan Sara S Krieger Marco A MA Marchini Fabricio K FK

Journal of proteome research 20171107 1

The pathogenic protozoan T. brucei alternates into distinct developmental stages in the mammalian and insect hosts. The mitogen-activated protein kinase (MAPK) signaling pathways transduce extracellular stimuli into a range of cellular responses, which ultimately lead to the adaptation to the external environment. Here, we combined a loss of function approach with stable isotope labeling with amino acids in cell culture (SILAC)-based mass spectrometry (MS) to investigate the role of the mitogen- ...[more]

PMID: 29043805

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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