Summary of Study ST002478

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001601. The data can be accessed directly via it's Project DOI: 10.21228/M8RH99 This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST002478
Study TitleThe effect of prions on cellular metabolism: The metabolic impact of the [RNQ+] prion and the native role of Rnq1p
Study SummaryWithin the field of amyloid and prion disease there is a need for a more comprehensive understanding of the fundamentals of disease biology. In order to facilitate the progression treatment and underpin comprehension of toxicity, fundamental understanding of the disruption to normal cellular biochemistry and trafficking is needed. Here, by removing the complex biochemistry of the brain, we have utilised known prion forming strains of Saccharomyces cerevisiae carrying different conformational variants of the Rnq1p to obtain Liquid Chromatography-Mass Spectrometry (LC-MS) metabolic profiles and identify key perturbations of prion presence. These studies reveal that prion containing [RNQ+] cells display a significant reduction in amino acid biosynthesis and distinct perturbations in sphingolipid metabolism, with significant downregulation in metabolites within these pathways. Moreover, that native Rnq1p downregulates ubiquinone biosynthesis pathways within cells, suggesting that Rnq1p may play a lipid/mevalonate-based cytoprotective role as a regulator of ubiquinone production. These findings contribute to the understanding of how prion proteins interact in vivo in both their prion and non-prion confirmations and indicate potential targets for the mitigation of these effects. . We demonstrate specific sphingolipid centred metabolic disruptions due to prion presence and give insight into a potential cytoprotective role of the native Rnq1 protein. This provides evidence of metabolic similarities between yeast and mammalian cells as a consequence of prion presence and establishes the application of metabolomics as a tool to investigate prion/amyloid-based phenomena.
Institute
Canterbury Christ Church University
Last NameHowell-Bray
First NameTyler
Address46 Canterbury Road, Kent
Emailt.l.howellbray@gmail.com
Phone07841631495
Submit Date2022-10-12
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailLC-MS
Release Date2023-02-26
Release Version1
Tyler Howell-Bray Tyler Howell-Bray
https://dx.doi.org/10.21228/M8RH99
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001601
Project DOI:doi: 10.21228/M8RH99
Project Title:The effect of prions on cellular metabolism: The metabolic impact of the [RNQ+] prion and the native role of Rnq1p
Project Summary:Within the field of amyloid and prion disease there is a need for a more comprehensive understanding of the fundamentals of disease biology. In order to facilitate the progression treatment and underpin comprehension of toxicity, fundamental understanding of the disruption to normal cellular biochemistry and trafficking is needed. Here, by removing the complex biochemistry of the brain, we have utilised known prion forming strains of Saccharomyces cerevisiae carrying different conformational variants of the Rnq1p to obtain Liquid Chromatography-Mass Spectrometry (LC-MS) metabolic profiles and identify key perturbations of prion presence. These studies reveal that prion containing [RNQ+] cells display a significant reduction in amino acid biosynthesis and distinct perturbations in sphingolipid metabolism, with significant downregulation in metabolites within these pathways. Moreover, that native Rnq1p downregulates ubiquinone biosynthesis pathways within cells, suggesting that Rnq1p may play a lipid/mevalonate-based cytoprotective role as a regulator of ubiquinone production. These findings contribute to the understanding of how prion proteins interact in vivo in both their prion and non-prion confirmations and indicate potential targets for the mitigation of these effects. . We demonstrate specific sphingolipid centred metabolic disruptions due to prion presence and give insight into a potential cytoprotective role of the native Rnq1 protein. This provides evidence of metabolic similarities between yeast and mammalian cells as a consequence of prion presence and establishes the application of metabolomics as a tool to investigate prion/amyloid-based phenomena.
Institute:Canterbury Christ Church University
Last Name:Howell-Bray
First Name:Tyler
Address:46 Canterbury Road, Kent
Email:t.l.howellbray@gmail.com
Phone:07841631495

Subject:

Subject ID:SU002568
Subject Type:Yeast
Subject Species:Saccharomyces cerevisiae

Factors:

Subject type: Yeast; Subject species: Saccharomyces cerevisiae (Factor headings shown in green)

mb_sample_id local_sample_id Treatment Ion mode
SA248069rnq-StressPIM50.2mM H202 negative
SA248070rnq-StressPIM60.2mM H202 negative
SA248071rnq-StressPIM40.2mM H202 negative
SA248072rnq-StressNIM30.2mM H202 negative
SA248073rnq-StressNIM60.2mM H202 negative
SA248074rnq-StressNIM20.2mM H202 negative
SA248075rnq-StressNIM10.2mM H202 negative
SA248076rnq-StressPIM20.2mM H202 negative
SA248077rnq-StressPIM10.2mM H202 negative
SA248078rnq-StressNIM50.2mM H202 negative
SA248079rnq-StressNIM40.2mM H202 negative
SA248080rnq-StressPIM30.2mM H202 negative
SA248057RNQ+StressNIM30.2mM H202 Positive
SA248058RNQ+StressNIM40.2mM H202 Positive
SA248059RNQ+StressNIM20.2mM H202 Positive
SA248060RNQ+StressNIM10.2mM H202 Positive
SA248061RNQ+StressNIM60.2mM H202 Positive
SA248062RNQ+StressNIM50.2mM H202 Positive
SA248063RNQ+StressPIM10.2mM H202 Positive
SA248064RNQ+StressPIM60.2mM H202 Positive
SA248065RNQ+StressPIM50.2mM H202 Positive
SA248066RNQ+StressPIM40.2mM H202 Positive
SA248067RNQ+StressPIM30.2mM H202 Positive
SA248068RNQ+StressPIM20.2mM H202 Positive
SA248093rnq-PIM6none negative
SA248094rnq-PIM5none negative
SA248095DeltaPIM1none negative
SA248096rnq-NIM1none negative
SA248097rnq-NIM2none negative
SA248098rnq-NIM3none negative
SA248099rnq-NIM4none negative
SA248100DeltaNIM6none negative
SA248101DeltaNIM5none negative
SA248102DeltaNIM2none negative
SA248103DeltaNIM3none negative
SA248104DeltaNIM4none negative
SA248105rnq-NIM5none negative
SA248106DeltaNIM1none negative
SA248107rnq-PIM1none negative
SA248108rnq-PIM2none negative
SA248109rnq-PIM3none negative
SA248110DeltaPIM6none negative
SA248111DeltaPIM5none negative
SA248112DeltaPIM2none negative
SA248113DeltaPIM3none negative
SA248114DeltaPIM4none negative
SA248115rnq-PIM4none negative
SA248081RNQ+PIM3none Positive
SA248082RNQ+PIM1none Positive
SA248083RNQ+PIM2none Positive
SA248084RNQ+NIM3none Positive
SA248085RNQ+PIM4none Positive
SA248086RNQ+PIM5none Positive
SA248087RNQ+PIM6none Positive
SA248088RNQ+NIM1none Positive
SA248089RNQ+NIM2none Positive
SA248090RNQ+NIM5none Positive
SA248091RNQ+NIM6none Positive
SA248092RNQ+NIM4none Positive
Showing results 1 to 59 of 59

Collection:

Collection ID:CO002561
Collection Summary:Strain and cultivation conditions The S. cerevisiae strain used in this study were derivatives of 74-D694 (MATa ade1-14(UGA) trp1-289(UAG) ura3-52 his3-∆200 leu2-3, 112) (Chernoff et al. 1993). Yeast harbouring [RNQ+] and knockout strain Δrnq were kind gifts from the Kent Fungal Group. Yeast were grown at 30°C with shaking at 180 rpm in synthetic complete (SC) media (2 % (w/v) glucose, 0.17 % Yeast Nitrogen Base (without amino acids, without ammonium sulphate), 0.5% ammonium sulphate, the appropriate concentration of yeast synthetic complete supplement mixture or synthetic complete drop-out media supplement). Transient growth on SC media containing 3mM guanidine hydrochloride (GdnHCl) was used as a curing agent in the media of S. cerevisiae cells that required a [prion-] status. Mild oxidative stresses were achieved by the addition of H2O2 (final concentration 0.2 mM) to the appropriate culture mediums. Cultures were grown using the filter culture method (as described by rabinowiz 2008)
Sample Type:Yeast cells
Storage Conditions:-80℃

Treatment:

Treatment ID:TR002580
Treatment Summary:Mild oxidative stresses were achieved by the addition of H2O2 (final concentration 0.2 mM)

Sample Preparation:

Sampleprep ID:SP002574
Sampleprep Summary:Metabolite sample preparation Analytical grade standards were supplied by Sigma Aldrich. Quenching was achieved by adaption of cold methanol protocol (56), via submersion of entire filter membrane. Metabolite extraction was performed on the resultant cell pellets using the boiling ethanol technique (57). Briefly, each tube was taken from the −80°C and 5 ml 75% (v/v) boiling ethanol was added (pre-heated). Each tube was immediately vortexed and placed in a water bath at 80°C. After 5 min each tube was cooled on ice for 3 min, followed by centrifugation (5000xg, 5 minutes, -20°C, precooled). Extracts were then stored at -80°C until further use. Immediately prior to mass spectrometry experimentation all extracts were concentrated by speed vacuum at 35° C for ≈ 3 hours. Following resuspension in 500µL of LC/MS grade water samples were lyophilised overnight. Lyophilised samples were then resuspended in 200µL of 0.1M formic acid, vortexed and loaded into vials.
Processing Storage Conditions:-80℃
Extraction Method:Boiling ethanol

Combined analysis:

Analysis ID AN004047
Analysis type MS
Chromatography type Reversed phase
Chromatography system ACQUITY SYNAPT G2-Si Mass Spectrometer
Column 1.7 µm C18 BEH column
MS Type ESI
MS instrument type QTOF
MS instrument name Waters Synapt-G2-Si
Ion Mode POSITIVE
Units m/z values

Chromatography:

Chromatography ID:CH002994
Instrument Name:ACQUITY SYNAPT G2-Si Mass Spectrometer
Column Name:1.7 µm C18 BEH column
Column Temperature:35
Flow Gradient:10-minute gradient from 0% to 50% acetonitrile (0.1% formic acid)
Flow Rate:500nL/min
Solvent A:95% Acetronitrile/5% water, 0.1% formic acid
Solvent B:100% water; 0.1% formic acid
Chromatography Type:Reversed phase

MS:

MS ID:MS003794
Analysis ID:AN004047
Instrument Name:Waters Synapt-G2-Si
Instrument Type:QTOF
MS Type:ESI
MS Comments:Data is labeled for negative or positive ionization mode
Ion Mode:POSITIVE
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