Dataset Information

Quantitative global proteomics of yeast PBP1 deletion mutants and their stress responses reveals glucose metabolism, translation and stress granule changes

ABSTRACT: The yeast protein PBP1 has been implicated in diverse pathways such as polyadenylation, translation, RNA-DNA hybrid formation, stress granule homeostasis, mitochondrial dysfunction, and TORC1 sequestration. Intriguingly, its deletion mitigates the toxicity of human neurodegeneration factors, but the molecular mechanisms of these effects are poorly understood. Here we performed label-free quantitative global proteomics to identify crucial downstream factors, comparing two PBP1 deletion strains (DB and SM) and two cell stress conditions (heat and NaN3). In all four analyses, downregulations of key bioenergetics enzymes (CIT1, SDH1, MLS1), cell wall mannoproteins (HSP150, PST1) and the prion protein RNQ1 as well as upregulations of the leucine biosynthesis enzyme LEU1 and the transcription factor TAF6 were documented. Consistently for both unstressed PBP1-deleted strains, over 2-fold dysregulations were documented for potential PBP1 interactors such as MKT1 or RPL39 and the stress granule component NRP1. Upregulation of the ribosomal biogenesis factor NOP10 was observed as in the mouse mutant. Consistently for both PBP1 deletion strains, heat stress triggered changes of the stress granule component GIS2 and several of its interactors.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: David Meierhofer

LAB HEAD: Georg Auburger

PROVIDER: PXD003868 | Pride | 2016-12-23

REPOSITORIES: Pride

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Quantitative Global Proteomics of Yeast PBP1 Deletion Mutants and Their Stress Responses Identifies Glucose Metabolism, Mitochondrial, and Stress Granule Changes.

Seidel Gunnar G Meierhofer David D Şen Nesli-Ece NE Guenther Anika A Krobitsch Sylvia S Auburger Georg G

Journal of proteome research 20161222 2

The yeast protein PBP1 is implicated in very diverse pathways. Intriguingly, its deletion mitigates the toxicity of human neurodegeneration factors. Here, we performed label-free quantitative global proteomics to identify crucial downstream factors, either without stress or under cell stress conditions (heat and NaN<sub>3</sub>). Compared to the wildtype BY4741 strain, PBP1 deletion always triggered downregulation of the key bioenergetics enzyme KGD2 and the prion protein RNQ1 as well as upregul ...[more]

PMID: 27966978

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Project description:Variant Surface Glycoproteins (VSG) coat parasitic African trypanosomes and underpin antigenic variation and immune evasion. This VSG is a super-abundant virulence factor that is subject to post-transcriptional gene expression controls mediated via the VSG 3’-untranslated region (3’-UTR). To identify positive VSG regulators in bloodstream form cells, we used prior genome-scale screening data to prioritise mRNA binding protein (mRBPs) knockdowns that phenocopy VSG mRNA knockdown, displaying both loss-of-fitness and pre-cytokinesis accumulation. The top three candidate VSG regulators were CFB2 (cyclin F-box protein 2, Tb927.1.4650), MKT1 (Tb927.6.4770) and PBP1 (polyadenylate binding-protein binding-protein, Tb927.8.4540). Notably, CFB2 was recently found to regulate VSG transcript stability, and all three proteins were found to associate with each other. We used data-independent acquisition for accurate label-free quantification and deep proteome coverage to quantify expression profiles following depletion of each mRBP. Only CFB2 knockdown significantly reduced VSG expression and the expression of a reporter under the control of a VSG 3’-untranslated region (3’-UTR). CFB2 knockdown also triggered depletion of cytoplasmic ribosomal proteins, consistent with translation arrest observed when VSG synthesis is blocked. In contrast, PBP1 knockdown triggered depletion of CFB2, MKT1, and other components of the PBP1-complex. Finally, all three knockdowns triggered depletion of cytokinesis initiation factors, consistent with the cytokinesis defect that was confirmed here for all three knockdowns. Thus, genome-scale knockdown datasets facilitate the triage and prioritisation of candidate regulators. Quantitative proteomic analysis confirms 3’-UTR dependent positive control of VSG expression by CFB2 and interactions with additional mRBPs. Our results also reveal connections between VSG expression control by CFB2, ribosomal protein expression, and cytokinesis.

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	BY_04_NaN3_1.raw	Raw
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