Summary of Study ST003174

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 PR001819. The data can be accessed directly via it's Project DOI: 10.21228/M8KQ72 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

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 IDST003174
Study TitleO-GlcNAcase activity maintains stress granules for proximity-enhanced ATP production to ensure recovery from stress (Part 3)
Study SummaryAccurate disassembly of stress granules (SGs) after environmental stimuli release is essential for cells to maintain homeostasis , which requires ATP-consuming processes. However, the molecular mechanism whereby regulation of SGs programmatically disassembly and ATP restoration remain poorly understood in mammalian cells. Here we found that defect of OGA in cells leads to aggregates formation, severe autophagy and eventually apoptosis during stress recovery. OGA, which localized in SGs, had no effect on SGs formation but could protect SGs from rapid disassembly during stress recovery stage. Then the SGs localized glycolysis-related enzymes were reserved and concentrated in SGs during stress release for ATP production in a proximity manner, which was vital to guarantee cells resistant to stress and survival during recovery. Finally, supplementation of ATP to OGA knockdown cells during stress recovery significantly rescue cell from aggregates, autophagy and apoptosis. Together, these results describe a brand new mechanism on how OGA regulates the programmed disassembly of stress granules and restoration of ATP to safeguard cell viability in a very precisely programmed process, whose rate is rigorous regulated. This is a continuation of study ST002927 and ST002936 where an in vitro reaction was performed with 13C-glucose on purified stress granules, to validate the conclusion.
Institute
Zhejiang University
DepartmentLife Sciences Institute
LaboratoryShixian Lin
Last NameChen
First NameYulin
AddressLife Sciences Institute, Zhejiang University, Hangzhou, Zhejiang province, China
Emailychen209@qq.com
Phone18868107794
Submit Date2024-04-16
Raw Data AvailableYes
Raw Data File Type(s)mzML, wiff
Analysis Type DetailLC-MS
Release Date2024-04-30
Release Version1
Yulin Chen Yulin Chen
https://dx.doi.org/10.21228/M8KQ72
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001819
Project DOI:doi: 10.21228/M8KQ72
Project Title:O-GlcNAcase activity maintains stress granules for proximity-enhanced ATP production to ensure recovery from stress
Project Summary:Accurate disassembly of stress granules (SGs) after environmental stimuli release is essential for cells to maintain homeostasis , which requires ATP-consuming processes. However, the molecular mechanism whereby regulation of SGs programmatically disassembly and ATP restoration remain poorly understood in mammalian cells. Here we found that defect of OGA in cells leads to aggregates formation, severe autophagy and eventually apoptosis during stress recovery. OGA, which localized in SGs, had no effect on SGs formation but could protect SGs from rapid disassembly during stress recovery stage. Then the SGs localized glycolysis-related enzymes were reserved and concentrated in SGs during stress release for ATP production in a proximity manner, which was vital to guarantee cells resistant to stress and survival during recovery. Finally, supplementation of ATP to OGA knockdown cells during stress recovery significantly rescue cell from aggregates, autophagy and apoptosis. Together, these results describe a brand new mechanism on how OGA regulates the programmed disassembly of stress granules and restoration of ATP to safeguard cell viability in a very precisely programmed process, whose rate is rigorous regulated.
Institute:Zhejiang University
Department:Life Sciences Institute
Laboratory:Shixian Lin
Last Name:Chen
First Name:Yulin
Address:Life Sciences Institute, Zhejiang University, Hangzhou, Zhejiang province, China
Email:ychen209@qq.com
Phone:18868107794

Subject:

Subject ID:SU003293
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606

Factors:

Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Sample source Factor
SA343191C1cultured cells 0hr
SA343192C3cultured cells 0hr
SA343193C2cultured cells 0hr
SA343194C6cultured cells 1hr
SA343195C5cultured cells 1hr
SA343196C4cultured cells 1hr
SA343197C9cultured cells 3hr
SA343198C8cultured cells 3hr
SA343199C7cultured cells 3hr
SA343200QC-2cultured cells QC
SA343201QC-3cultured cells QC
SA343202QC-1cultured cells QC
Showing results 1 to 12 of 12

Collection:

Collection ID:CO003286
Collection Summary:Care was taken to quench cells quickly to minimize oxidation and degradation.
Sample Type:in vitro reaction

Treatment:

Treatment ID:TR003302
Treatment Summary:Stress granules purified from HeLa cells were treated with 13C-Glucose for 0, 1, 3 hr reaction.

Sample Preparation:

Sampleprep ID:SP003300
Sampleprep Summary:Cells without stress treatment or treated with 0.6 M sorbitol stress for 1.5 h were scraped off and centrifuged to collect cell pellets. The cell pellets were thawed on ice, resuspended in stress granule lysis buffer (50 mM Tris, 100 mM K2OAc, 2 mM MgOAc, 0.5 mM DTT, 50 μg/mL heparin, 0.5% NP-40, 0.02% antifoam B, proteinase inhibitor cocktail, pH 7.4) and lysis by syringe. Lysates were centrifuged at 1000 × g for 5 min to collect supernatants. Supernatants were centrifuge at 18000 × g for 20 min to collect pellets and the pellets were washed once with stress granule lysis buffer. Then, pellets were resuspended in stress granule lysis buffer and centrifuged at 850 × g for 2 min to collect supernatants containing stress granule. Then NAD+, ATP and 13C-glucose were added for reaction at 37 'C for 0, 1 and 3 hr. The reaction was quenched by 3-fold methanol and centrifuge to collect the supernant for analysis.

Combined analysis:

Analysis ID AN005209 AN005210
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Shimadzu 20AD Shimadzu 20AD
Column Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um) Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um)
MS Type ESI ESI
MS instrument type QTRAP QTRAP
MS instrument name ABI Sciex Triple Quad 5500+ ABI Sciex Triple Quad 5500+
Ion Mode POSITIVE NEGATIVE
Units Peak intensity Peak intensity

Chromatography:

Chromatography ID:CH003941
Instrument Name:Shimadzu 20AD
Column Name:Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um)
Column Temperature:45
Flow Gradient:0.01 min 80% B, 20 min 20% B, 20.5 min 80% B, 24 min 80% B.
Flow Rate:0.15 mL/min
Solvent A:100% water; 20 mM ammonium carbonate
Solvent B:100% acetonitrile
Chromatography Type:HILIC

MS:

MS ID:MS004942
Analysis ID:AN005209
Instrument Name:ABI Sciex Triple Quad 5500+
Instrument Type:QTRAP
MS Type:ESI
MS Comments:The raw data were extracted with the software Analyst v1.7.2 and OS v1.7. This is a continuation of study ST002927 where a different set precusror/product ions were chosen for MS.
Ion Mode:POSITIVE
  
MS ID:MS004943
Analysis ID:AN005210
Instrument Name:ABI Sciex Triple Quad 5500+
Instrument Type:QTRAP
MS Type:ESI
MS Comments:The raw data were extracted with the software Analyst v1.7.2 and OS v1.7. This is a continuation of study ST002927 where a different set precursor/product ions were chosen for MS.
Ion Mode:NEGATIVE
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