Summary of Study ST004097

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 PR002513. The data can be accessed directly via it's Project DOI: 10.21228/M8T536 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.

Study ID	ST004097
Study Title	ODC1 overexpression increased the extracellular putrescine concentration in both WT and SLC45A4 KO cells
Study Summary	We found that the expression of SLC45A4 has a strong positive correlation with the cellular level of γ-aminobutyric acid (GABA). Using mass spectrometry and the stable isotope tracing approach, we demonstrated that SLC45A4 promotes GABA de novo synthesis through the Arginine/Ornithine/Putrescine (AOP) pathway. SLC45A4 functions as a putrescine transporter localized to the peroxisome membrane to facilitate GABA production. Taken together, our results revealed a new biochemical mechanism where SLC45A4 controls GABA production. Overexpression of the human ornithine decarboxylase (hODC1) significantly increased the level of GABA in both SLC45A4 WT and KO cells. However, the ODC1 overexpression did not fully rescue the deficiency in GABA production in SLC45A4 KO cells, suggesting SLC45A4 is required for efficient putrescine oxidation. Meanwhile, ODC1 overexpression increased the extracellular putrescine concentration in both WT and SLC45A4 KO cells.
Institute	Rutgers University
Department	Department of Medicine
Last Name	Su
First Name	Xiaoyang
Address	7118 Clinical Academic Building, 125 Paterson Street, New Brunswick, NJ, 08901
Email	xs137@rwjms.rutgers.edu
Phone	17322355447
Submit Date	2025-06-24
Raw Data Available	Yes
Raw Data File Type(s)	mzXML
Analysis Type Detail	LC-MS
Release Date	2025-08-07
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002513
Project DOI:	doi: 10.21228/M8T536
Project Title:	SLC45A4 encodes a peroxisomal putrescine transporter that promotes GABA de novo synthesis
Project Type:	Cellular polar metabolomics
Project Summary:	Solute carriers (SLC) are membrane proteins that facilitate the transportation of ions and metabolites across either the plasma membrane or the membrane of intracellular organelles. With more than 450 human genes annotated as SLCs, many of them are still orphan transporters without known biochemical functions. We developed a metabolomic-transcriptomic association analysis, and we found that the expression of SLC45A4 has a strong positive correlation with the cellular level of γ-aminobutyric acid (GABA). Using mass spectrometry and the stable isotope tracing approach, we demonstrated that SLC45A4 promotes GABA de novo synthesis through the Arginine/Ornithine/Putrescine (AOP) pathway. SLC45A4 functions as a putrescine transporter localized to the peroxisome membrane to facilitate GABA production. Taken together, our results revealed a new biochemical mechanism where SLC45A4 controls GABA production.
Institute:	Rutgers University
Department:	Department of Medicine
Last Name:	Su
First Name:	Xiaoyang
Address:	7118 Clinical Academic Building
Email:	xs137@rwjms.rutgers.edu
Phone:	7322355447
Funding Source:	R01 GM149664

Subject:

Subject ID:	SU004243
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Cell Strain Details:	A549, H1299

Factors:

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

mb_sample_id	local_sample_id	Sample source	Cell line	Genotype	Overexpression
SA474275	mediaA549-4F1_ctlGFP_2	Media	A549	SLC45A4-KO	Ctrl
SA474276	mediaA549-2E2_ctlGFP_2	Media	A549	SLC45A4-KO	Ctrl
SA474277	mediaA549-2E2_ctlGFP_1	Media	A549	SLC45A4-KO	Ctrl
SA474278	mediaA549-4F1_ctlGFP_1	Media	A549	SLC45A4-KO	Ctrl
SA474279	mediaA549-4F1_hODC1_1	Media	A549	SLC45A4-KO	human-ODC1
SA474280	mediaA549-4F1_hODC1_2	Media	A549	SLC45A4-KO	human-ODC1
SA474281	mediaA549-2E2_hODC1_1	Media	A549	SLC45A4-KO	human-ODC1
SA474282	mediaA549-2E2_hODC1_2	Media	A549	SLC45A4-KO	human-ODC1
SA474283	mediaA549-WT_ctlGFP_1	Media	A549	WT	Ctrl
SA474284	mediaA549-WT_ctlGFP_2	Media	A549	WT	Ctrl
SA474285	mediaA549-WT_hODC1_1	Media	A549	WT	human-ODC1
SA474286	mediaA549-WT_hODC1_2	Media	A549	WT	human-ODC1
SA474287	mediaH1299-6C1_ctlGFP_1	Media	H1299	SLC45A4-KO	Ctrl
SA474288	mediaH1299-6C1_ctlGFP_2	Media	H1299	SLC45A4-KO	Ctrl
SA474289	mediaH1299-3G2_ctlGFP_1	Media	H1299	SLC45A4-KO	Ctrl
SA474290	mediaH1299-3G2_ctlGFP_2	Media	H1299	SLC45A4-KO	Ctrl
SA474291	mediaH1299-3G2_hODC1_2	Media	H1299	SLC45A4-KO	human-ODC1
SA474292	mediaH1299-6C1_hODC1_1	Media	H1299	SLC45A4-KO	human-ODC1
SA474293	mediaH1299-6C1_hODC1_2	Media	H1299	SLC45A4-KO	human-ODC1
SA474294	mediaH1299-3G2_hODC1_1	Media	H1299	SLC45A4-KO	human-ODC1
SA474295	mediaH1299-WT_ctlGFP_1	Media	H1299	WT	Ctrl
SA474296	mediaH1299-WT_ctlGFP_2	Media	H1299	WT	Ctrl
SA474297	mediaH1299-WT_hODC1_1	Media	H1299	WT	human-ODC1
SA474298	mediaH1299-WT_hODC1_2	Media	H1299	WT	human-ODC1

Showing results 1 to 24 of 24

Showing results 1 to 24 of 24

Collection:

Collection ID:	CO004236
Collection Summary:	0.5×10^6 cells in the culture were washed twice with PBS and extracted with 0.5 mL ice-cold 40:40:20 (methanol:acetonitrile:water) solution with 0.5% formic acid. The cells were scraped off the plate followed by incubation on ice for 10 min, and neutralized by the addition of 25 μL 15% (m/v) NH4HCO3 solution. The extracts were transferred to a 1.5 ml tube. For conditioned media metabolite extraction, 5 µL of culture media was mixed with 500 µl of ice-cold 40:40:20 (methanol:acetonitrile:water) solution with 0.5% formic acid and neutralized by the addition of 25 μL 15% (m/v) NH4HCO3 solution. The whole cell or media extracts were then centrifuged at 4°C and 16,000 × g for 10 min and transferred to a clean tube and stored at -80°C until analysis.
Sample Type:	Cultured cells

Treatment:

Treatment ID:	TR004252
Treatment Summary:	For transient overexpression of ODC1, cells were seeded in new 12-well plates at the density of 2×105 / mL. When the cells reached about 70% confluence, cells were transfected with Lipofectamine 3000 according to manufacturer’s instructions (Invitrogen, L3000015). 1 μg expression vector plasmid DNA was first mixed with 2 μL P3000 reagent in 50 μL Opti-MEM reduced serum medium (Gibco, 31985070), then mixed with 3 μL Lipofectamine 3000 in another 50 μL Opti-MEM. The mixture was incubated at room temperature for 15 min and added directly to the cell culture. 24h later, the media were replaced with the one containing stable isotope tracers if necessary.

Treatment ID:

TR004252

Treatment Summary:

For transient overexpression of ODC1, cells were seeded in new 12-well plates at the density of 2×105 / mL. When the cells reached about 70% confluence, cells were transfected with Lipofectamine 3000 according to manufacturer’s instructions (Invitrogen, L3000015). 1 μg expression vector plasmid DNA was first mixed with 2 μL P3000 reagent in 50 μL Opti-MEM reduced serum medium (Gibco, 31985070), then mixed with 3 μL Lipofectamine 3000 in another 50 μL Opti-MEM. The mixture was incubated at room temperature for 15 min and added directly to the cell culture. 24h later, the media were replaced with the one containing stable isotope tracers if necessary.

Sample Preparation:

Sampleprep ID:	SP004249
Sampleprep Summary:	For conditioned media metabolite extraction, 5 µL of culture media was mixed with 500 µL of ice-cold 40:40:20 (methanol:acetonitrile:water) solution with 0.5% formic acid and neutralized by the addition of 25 μL 15% (m/v) NH4HCO3 solution

Chromatography:

Chromatography ID:	CH005160
Instrument Name:	Thermo Vanquish
Column Name:	Waters XBridge BEH Amide (150 mm × 2.1 mm, 2.5 µm)
Column Temperature:	25°C
Flow Gradient:	0 min, 100% B; 3 min, 100% B; 3.2 min, 90% B; 6.2 min, 90% B; 6.5 min, 80% B; 10.5 min, 80% B; 10.7 min, 70% B; 13.5 min, 70% B; 13.7 min, 45% B; 16 min, 45% B; 16.5 min, 100% B; and 22 min, 100% B
Flow Rate:	0.3 mL/min
Solvent A:	95% Water/5% Acetonitrile; 20 mM acetic acid; 40 mM ammonium hydroxide (pH 9.4)
Solvent B:	80% Acetonitrile/20% Water; 20 mM acetic acid; 40 mM ammonium hydroxide (pH 9.4)
Chromatography Type:	HILIC

Analysis:

Analysis ID:	AN006792
Laboratory Name:	Metabolomics Shared Resource of Rutgers Cancer Institute
Analysis Type:	MS
Detector Type:	Orbitrap
Data Format:	mzXML
Chromatography ID:	CH005160
Num Factors:	8
Num Metabolites:	19
Units:	Ion counts