Summary of Study ST001909

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 PR001203. The data can be accessed directly via it's Project DOI: 10.21228/M86Q68 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 IDST001909
Study TitleATF3 regulation of serine metabolism
Study Typestable isotope tracing
Study SummaryATF3 is a common stress sensor, and its expression can be induced by serine deprivation. The goal of this project is to determine whether ATF3 regulates serine biosynthesis. ATF3-wildtype and –knockout cells are cultured in complete or serine-free medium supplemented with 13C-6-glucose for 24 h for stable isotope tracing. The results show that ATF3 appears to promote serine biosynthesis.
Institute
Augusta University
Last NameYan
First NameChunhong
Address1410 Laney Walker Blvd, Augusta, GA, 30912, USA
Emailcyan@augusta.edu
Phone7067210099
Submit Date2021-08-11
Raw Data AvailableYes
Raw Data File Type(s)d
Analysis Type DetailGC-MS
Release Date2021-10-02
Release Version1
Chunhong Yan Chunhong Yan
https://dx.doi.org/10.21228/M86Q68
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001203
Project DOI:doi: 10.21228/M86Q68
Project Title:ATF3 regulation of serine metabolism
Project Type:Isotope Tracing, GC-MS
Project Summary:ATF3 is a common stress sensor, and its expression can be induced by serine deprivation. The goal of this project is to determine whether ATF3 regulates serine biosynthesis. ATF3-wildtype and –knockout cells are cultured in complete or serine-free medium supplemented with 13C-6-glucose for 24 h for stable isotope tracing. The results show that ATF3 appears to promote serine biosynthesis.
Institute:Augusta University
Department:Georgia Cancer Center
Last Name:Yan
First Name:Chunhong
Address:1410 Laney Walker Blvd, Augusta, GA, 30912, USA
Email:cyan@augusta.edu
Phone:7067210099
Funding Source:NIH/NCI R01CA240933, R01CA190429, R01CA236890

Subject:

Subject ID:SU001987
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606
Genotype Strain:ATF3(+/+) vs ATF3(-/-)
Cell Strain Details:HCT116 cells engineered to knockout ATF3 expression via rAAV-mediated homologous recombination

Factors:

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

mb_sample_id local_sample_id Genotype_Media supplement
SA176874Ding_C-3_015KO-complete
SA176875Ding_C-4_016KO-complete
SA176876Ding_C-6_018KO-complete
SA176877Ding_C-2_014KO-complete
SA176878Ding_C-5_017KO-complete
SA176879Ding_C-1_013KO-complete
SA176880Ding_D-2_020KO-serine(-)
SA176881Ding_D-1_019KO-serine(-)
SA176882Ding_D-3_021KO-serine(-)
SA176883Ding_D-4_022KO-serine(-)
SA176884Ding_D-5_023KO-serine(-)
SA176885Ding_D-6_024KO-serine(-)
SA176886Ding_A-3_003WT-complete
SA176887Ding_A-2_002WT-complete
SA176888Ding_A-4_004WT-complete
SA176889Ding_A-5_005WT-complete
SA176890Ding_A-6_006WT-complete
SA176891Ding_A-1_001WT-complete
SA176892Ding_B-4_010WT-serine(-)
SA176893Ding_B-3_009WT-serine(-)
SA176894Ding_B-5_011WT-serine(-)
SA176895Ding_B-6_012WT-serine(-)
SA176896Ding_B-2_008WT-serine(-)
SA176897Ding_B-1_007WT-serine(-)
Showing results 1 to 24 of 24

Collection:

Collection ID:CO001980
Collection Summary:Cells were counted, washed with cold PBS and then flash-frozen in liquid nitrogen
Sample Type:Cultured cells

Treatment:

Treatment ID:TR001999
Treatment Summary:HCT116 cells (~50% confluence) were washed with PBS, and cultured in HEPES buffered Krebs-Ringer solution supplemented with 25 mM [U-13C]-D-glucose, 10% dialysed FBS, 2×MEM amino acids, 2× Vitamin Solution and 4 mM L-glutamine, with or without 0.4 mM serine and glycine, for 24 h. Cells were washed with PBS and metabolites extracted using 1 mL of acetonitrile:isopropanol:water (3:3:2, v/v/v) mixture.

Sample Preparation:

Sampleprep ID:SP001993
Sampleprep Summary:The extraction solvent was degassed and pre-chilled at −20°C. Samples were homogenized using Geno/Grinder 2010 (SPEX SamplePrep) at 1500 rpm for 30s, then shaken at 4°C for 5 min and centrifuged for 2 minutes at 14,000 rcf. 450 μL supernatant was transferred to a new tube and dried down using Centrivap cold trap concentrator (Labconco). 10 μL of methoxyamine hydrochloride in pyridine (40 mg/mL) was added to dried sample and shaken at 30°C for 1.5 hours for methoximation. 90 μL of N-tert-Butyldimethylsilyl-N-methyltrifluoroacetamide (MTBSTFA, Sigma-Aldrich) was used for tertbutylsilylation. C8–C30 fatty acid methyl esters (FAMEs) were added to MTBSTFA as internal standards for retention time correction. Samples were shaken at 37°C for 30 min for TMS or shaken at 80°C for 30 min for TBS and then ready for injection.

Combined analysis:

Analysis ID AN003106
Analysis type MS
Chromatography type GC
Chromatography system Agilent 7890A
Column Restek Rtx-5Sil (30m x 0.25mm,0.25um)
MS Type EI
MS instrument type GC-TOF
MS instrument name Agilent 7890A
Ion Mode UNSPECIFIED
Units enrichment percentage

Chromatography:

Chromatography ID:CH002293
Chromatography Summary:Gas chromatography-Quadrupole-time of flight mass spectrometry (GC-Q-TOF MS) was used for Stable Isotope enrichment. Rtx-5Sil MS column (30m length, 0.25 mm i.d, 0.25 μM 95% dimethyl 5% diphenyl polysiloxane film) with an additional 10 m guard column was installed on Agilent 7890 GC (Agilent Technologies). 99.9999% pure Helium gas was used as a mobile phase with a flow rate of 1mL/min. GC temperature was held at 50°C for 1 min, ramped at 20°C/min to 330°C and then held for 5 min. Electron ionization at -70eV was employed on a Agilent 7250 Quadrupole time of flight mass spectrometer (Agilent Technologies) with ion source temperature at 250°C and detector voltage at 1850V. Mass spectra were acquired at an acquisition rate of 5 spectra/s, with a scan range of 85-900 Da.
Instrument Name:Agilent 7890A
Column Name:Restek Rtx-5Sil (30m x 0.25mm,0.25um)
Chromatography Type:GC

MS:

MS ID:MS002888
Analysis ID:AN003106
Instrument Name:Agilent 7890A
Instrument Type:GC-TOF
MS Type:EI
MS Comments:Agilent MassHunter Quantitative Analysis software B.10.00 (Agilent Technologies) was used for post data processing.
Ion Mode:UNSPECIFIED
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