Summary of Study ST003306

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 PR002056. The data can be accessed directly via it's Project DOI: 10.21228/M8VG0G 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 IDST003306
Study TitleASCT2 is a major contributor to serine uptake in cancer cells
Study SummaryThe non-essential amino acid serine is a critical nutrient for cancer cells due to its diverse biosynthetic functions. While some tumors can synthesize serine de novo, others are auxotrophic and therefore reliant on serine uptake. Importantly, despite several transporters being known to be capable of transporting serine, the transporter(s) that mediate serine uptake in cancer cells are not known. Here, we characterize the amino acid transporter ASCT2 (SLC1A5) as a major contributor to serine uptake in cancer cells. ASCT2 is well-known as a glutamine transporter in cancer, and our work demonstrates that serine and glutamine compete for uptake through ASCT2. We further show that ASCT2-mediated serine uptake is essential for purine nucleotide biosynthesis and that ERα promotes serine uptake by directly activating SLC1A5 transcription. Together, our work defines an additional important role for ASCT2 as a serine transporter in cancer and evaluates ASCT2 as a potential therapeutic target.
Institute
University of Illinois Chicago
DepartmentPhysiology and Biophysics
LaboratoryColoff Lab
Last NameConger
First NameKelly
Address909 S Wolcott Ave, Chicago, IL, 60612
Emailkconge2@uic.edu
Phone2314320406
Submit Date2024-06-17
Raw Data AvailableYes
Raw Data File Type(s)d
Analysis Type DetailGC-MS
Release Date2024-07-15
Release Version1
Kelly Conger Kelly Conger
https://dx.doi.org/10.21228/M8VG0G
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR002056
Project DOI:doi: 10.21228/M8VG0G
Project Title:ASCT2 is a major contributor to serine uptake in cancer cells
Project Type:GCMS quantitative analysis
Project Summary:Polar metabolite abundance studies from MCF7 human ER+ breast cancer cell line with or without CRISPR-Cas9 knockout of ASCT2. Studies used RPMI media with either complete serine levels (285uM), low serine levels (50uM), or no glutamine.
Institute:University of Illinois Chicago
Department:Physiology and Biophysics
Laboratory:Coloff Lab
Last Name:Conger
First Name:Kelly
Address:909 S Wolcott Ave, Chicago, IL, 60612
Email:kconge2@uic.edu
Phone:2314320406

Subject:

Subject ID:SU003427
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606
Gender:Female
Cell Strain Details:MCF7
Cell Primary Immortalized:Immortalized

Factors:

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

mb_sample_id local_sample_id Sample source Genotype Treatment
SA358636SLC1A5-3,2Human breast cancer cells ASCT2-KO 285uM Serine
SA358637SLC1A5-3,3Human breast cancer cells ASCT2-KO 285uM Serine
SA358638SLC1A5-1,1Human breast cancer cells ASCT2-KO 285uM Serine
SA358639SLC1A5-1,2Human breast cancer cells ASCT2-KO 285uM Serine
SA358640SLC1A5-1,3Human breast cancer cells ASCT2-KO 285uM Serine
SA358641SLC1A5-3,1Human breast cancer cells ASCT2-KO 285uM Serine
SA358642sgSLC1A5-1 IM 1Human breast cancer cells ASCT2-KO 50uM Serine
SA358643sgSLC1A5-1 IM 2Human breast cancer cells ASCT2-KO 50uM Serine
SA358644sgSLC1A5-1 IM 3Human breast cancer cells ASCT2-KO 50uM Serine
SA358645sgSLC1A5-3 IM 1Human breast cancer cells ASCT2-KO 50uM Serine
SA358646sgSLC1A5-3 IM 2Human breast cancer cells ASCT2-KO 50uM Serine
SA358647sgSLC1A5-3 IM 3Human breast cancer cells ASCT2-KO 50uM Serine
SA358651Luc Puro 3Human breast cancer cells Ctrl 285uM Serine
SA358652Luc Puro 1Human breast cancer cells Ctrl 285uM Serine
SA358653Luc Puro 2Human breast cancer cells Ctrl 285uM Serine
SA358654sgLuc IM 2Human breast cancer cells Ctrl 50uM Serine
SA358655sgLuc IM 3Human breast cancer cells Ctrl 50uM Serine
SA358656sgLuc IM 1Human breast cancer cells Ctrl 50uM Serine
SA358648sgLuc -Gln 60 3Human breast cancer cells Ctrl (-Gln)1hr
SA358649sgLuc -Gln 60 2Human breast cancer cells Ctrl (-Gln)1hr
SA358650sgLuc -Gln 60 1Human breast cancer cells Ctrl (-Gln)1hr
Showing results 1 to 21 of 21

Collection:

Collection ID:CO003420
Collection Summary:Cells were washed with PBS and fed media containing either complete serine (285uM) or low serine (50uM) for six hours. Cells were washed with saline and harvested in cold GCMS-grade methanol with norvaline diluted in water. Polar metabolites were extracted with the addition of chloroform. Samples were dried under constant air flow for 2 hours.
Collection Protocol Filename:CL_PM2024.pdf
Sample Type:Cultured cells
Storage Conditions:-80℃

Treatment:

Treatment ID:TR003436
Treatment Summary:Cells are either control (sgLuc) or ASCT2-KO (sgSLC1A5). All groups were plated in triplicate in a six well plate 48hours prior to treatment. Cells were treated with either complete media, or media containing low serine for six hours, or media lacking glutamine (-Q) for one hour prior to harvesting.
Treatment Doseduration:1-6 hours
Cell Growth Container:Corning 6-well plates
Cell Media:RPMI w/o Glucose, Sodium Pyruvate, Amino acids
Cell Pct Confluence:80%

Sample Preparation:

Sampleprep ID:SP003434
Sampleprep Summary:Samples were harvested in cold GCMS-grade methanol with norvaline diluted in water. Polar metabolites were separated with chloroform and air dried for two hours. Samples were then rehydrated in 15uL MOX reagent and heated at 37C for 90 minutes. Then 20uL of TBDMS was added to each sample and heated at 60C for 60 minutes.
Sampleprep Protocol Filename:CL_PM2024.pdf

Combined analysis:

Analysis ID AN005417
Analysis type MS
Chromatography type GC
Chromatography system Agilent 7890B
Column Agilent HP5-MS (30m x 0.25mm, 0.25 um)
MS Type EI
MS instrument type Single quadrupole
MS instrument name Agilent 5977B
Ion Mode POSITIVE
Units Abundance

Chromatography:

Chromatography ID:CH004107
Chromatography Summary:All samples were analyzed by GC/MS using a HP-5MS Ultra Inert GC column (19091S-433UI, Agilent Technologies) installed in an Agilent 7890B gas chromatograph coupled to an Agilent 5779B mass spectrometer. Helium was used as the carrier gas. One microliter was injected (split inlet) at 280 degrees C. After injection, the GC oven was held at 60 degrees C for 1 minute before ramping to 320 degrees C at 10C/min and held for 9 minutes at the maximum temperature.
Instrument Name:Agilent 7890B
Column Name:Agilent HP5-MS (30m x 0.25mm, 0.25 um)
Column Temperature:60-320
Flow Gradient:N/A
Flow Rate:1.5mL/min
Solvent A:N/A
Solvent B:N/A
Chromatography Type:GC

MS:

MS ID:MS005143
Analysis ID:AN005417
Instrument Name:Agilent 5977B
Instrument Type:Single quadrupole
MS Type:EI
MS Comments:The MS system operated under electron impact ionization mode at 70 eV and the MS source and quadrupole were held at 230 degrees C and 150 degrees C respectively. Peak areas were determined using MassHunter software.
Ion Mode:POSITIVE
Analysis Protocol File:CL_GCMS2024.pdf
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