Summary of Study ST003198
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 PR001988. The data can be accessed directly via it's Project DOI: 10.21228/M8RJ07 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 | ST003198 |
Study Title | UQ/RQ panel on human tissue |
Study Summary | The extraction of nonpolar metabolites from the tissues of human for analysis by LCMS to measure levels of Ubiquinone and Rhodoquinone. |
Institute | UMass Chan Medical School |
Department | Program in Molecular Medicine |
Laboratory | Spinelli Lab |
Last Name | Jerome |
First Name | Madison |
Address | 55 N Lake Ave, Worcester, MA 01655 |
madison.jerome@umassmed.edu | |
Phone | (508) 856-8989 ext. 68148 |
Submit Date | 2024-05-01 |
Raw Data Available | Yes |
Raw Data File Type(s) | raw(Thermo) |
Analysis Type Detail | LC-MS |
Release Date | 2025-02-04 |
Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
Project ID: | PR001988 |
Project DOI: | doi: 10.21228/M8RJ07 |
Project Title: | Rhodoquinone is an Electron Carrier in the Mammalian Electron Transport Chain |
Project Summary: | Ubiquinone (UQ), the only known electron carrier in the mammalian electron transport chain (ETC), delivers electrons to both oxygen (O2) and fumarate as terminal electron acceptors. As fumarate has a lower reduction potential than UQ, fumarate reduction is only thermodynamically favorable when ubiquinol, the reduced form of UQ, accumulates. Paradoxically, some tissues reduce fumarate without ubiquinol buildup, suggesting another mechanism enables fumarate reduction in mammals. Here, we identify rhodoquinone (RQ), a novel mammalian electron carrier that directs electrons to fumarate, instead of O2, as the favored terminal electron acceptor. RQ, which is undetectable in cultured mammalian cells, is enriched in tissues that catalyze fumarate reduction. RQ and UQ-directed ETC circuits support distinct programs of mitochondrial function. Through expression of a bacterial enzyme that converts UQ into RQ and development a novel RQ analog, we demonstrate that reprogramming the mammalian ETC from the UQ to RQ circuit renders cells highly resistant to hypoxia exposure. Thus, we establish RQ as a fundamental component of the mammalian ETC and unveil reprogramming the ETC to the RQ-circuit as a tractable strategy to treat hypoxia-related diseases. |
Institute: | UMass Chan Medical School |
Department: | Program in Molecular Medicine |
Laboratory: | Spinelli Lab |
Last Name: | UMass Chan |
First Name: | Spinelli Lab |
Address: | 55 Lake Avenue North, Worcester, Massachusetts, 01605, USA |
Email: | spinellilab@gmail.com |
Phone: | (508) 856-8989 ext. 68148 |
Subject:
Subject ID: | SU003317 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Species Group: | Mammals |
Factors:
Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)
mb_sample_id | local_sample_id | Sample source | Sample type |
---|---|---|---|
SA348075 | Mito AP01 | Adipose Pannus | Mitochondria |
SA348076 | Mito AP02 | Adipose Pannus | Mitochondria |
SA348077 | Mito AP03 | Adipose Pannus | Mitochondria |
SA348078 | Mito AP05 | Adipose Pannus | Mitochondria |
SA348079 | Mito AP04 | Adipose Pannus | Mitochondria |
SA348080 | WholeTissue AP01 | Adipose Pannus | Whole Tissue |
SA348081 | WholeTissue AP03 | Adipose Pannus | Whole Tissue |
SA348082 | WholeTissue AP02 | Adipose Pannus | Whole Tissue |
SA348083 | WholeTissue AP05 | Adipose Pannus | Whole Tissue |
SA348084 | WholeTissue AP04 | Adipose Pannus | Whole Tissue |
SA348085 | Mito B04 | Brain | Mitochondria |
SA348086 | Mito B03 | Brain | Mitochondria |
SA348087 | Mito B02 | Brain | Mitochondria |
SA348088 | Mito B01 | Brain | Mitochondria |
SA348089 | Mito B05 | Brain | Mitochondria |
SA348090 | WholeTissue B02 | Brain | Whole Tissue |
SA348091 | WholeTissue B04 | Brain | Whole Tissue |
SA348092 | WholeTissue B03 | Brain | Whole Tissue |
SA348093 | WholeTissue B01 | Brain | Whole Tissue |
SA348094 | WholeTissue B05 | Brain | Whole Tissue |
SA348095 | Mito K01 | Kidney | Mitochondria |
SA348096 | Mito K02 | Kidney | Mitochondria |
SA348097 | Mito K03 | Kidney | Mitochondria |
SA348098 | Mito K05 | Kidney | Mitochondria |
SA348099 | Mito K04 | Kidney | Mitochondria |
SA348100 | WholeTissue K05 | Kidney | Whole Tissue |
SA348101 | WholeTissue K04 | Kidney | Whole Tissue |
SA348102 | WholeTissue K03 | Kidney | Whole Tissue |
SA348103 | WholeTissue K01 | Kidney | Whole Tissue |
SA348104 | WholeTissue K02 | Kidney | Whole Tissue |
SA348105 | Mito M01 | Muscle | Mitochondria |
SA348106 | Mito M05 | Muscle | Mitochondria |
SA348107 | Mito M02 | Muscle | Mitochondria |
SA348108 | Mito M03 | Muscle | Mitochondria |
SA348109 | Mito M04 | Muscle | Mitochondria |
SA348110 | WholeTissue M05 | Muscle | Whole Tissue |
SA348111 | WholeTissue M04 | Muscle | Whole Tissue |
SA348112 | WholeTissue M02 | Muscle | Whole Tissue |
SA348113 | WholeTissue M01 | Muscle | Whole Tissue |
SA348114 | WholeTissue M03 | Muscle | Whole Tissue |
SA348115 | CoQ10 5µM | Standard | CoQ10 Standard |
SA348116 | CoQ10 10µM | Standard | CoQ10 Standard |
SA348117 | CoQ10 1µM | Standard | CoQ10 Standard |
SA348118 | CoQ10 0.1µM | Standard | CoQ10 Standard |
SA348119 | CoQ10 0.001µM | Standard | CoQ10 Standard |
SA348120 | CoQ10 0.01µM | Standard | CoQ10 Standard |
SA348121 | CoQ10 0.25µM | Standard | CoQ10 Standard |
SA348122 | CoQ10 0.5µM | Standard | CoQ10 Standard |
Showing results 1 to 48 of 48 |
Collection:
Collection ID: | CO003310 |
Collection Summary: | Tissues were harvested, flash frozen in liquid nitrogen, and stored at -80°C until samples were ready for processing. |
Sample Type: | Brain, Kidney, Muscle, Adipose Pannus |
Treatment:
Treatment ID: | TR003326 |
Treatment Summary: | Untreated |
Sample Preparation:
Sampleprep ID: | SP003324 |
Sampleprep Summary: | UQRQ isolation from tissues: Tissues were flash frozen in liquid nitrogen and powderized using a mortar and pestle. Approximately 10 mg of tissue powder was transferred into an Eppendorf tube. The tissue powder was re-suspended in 500 µL of 99.9% LCMS-grade methanol (Fisher) 0.1% HCl and then vortexed for 15 minutes at 4°C. 500 µL LCMS-grade hexane (Honeywell) was added to the lysate and again vortexed for 15 minutes at 4°C. Samples were then centrifuged at 16,000 x g for 10 minutes at 4°C with the top (hexane) layer containing UQ and RQ, the middle layer containing the acidified methanol, and the bottom layer containing the protein. The top layer was transferred into a new Eppendorf tube, dried down in a Refrigerated CentriVap Benchtop Vacuum Concentrator connected to a CentriVap-105 Cold Trap (Labconco), and stored at -80°C until they were re-suspended for LC-MS analysis. The acidified methanol layer was discarded, and the bottom protein layer was saved for protein quantification. The protein layer from the metabolite isolation was re-suspended in 1 mL of RIPA buffer (150 mM NaCl, 50 mM Tris HCl pH 7.5, 0.1% SDS, 1% Triton-X 100 (Sigma), 0.5% deoxycholate (Sigma), cOmplete EDTA-free protease inhibitor (Sigma)). The samples were vortexed for 10 minutes at 4°C and then centrifuged at 16,000 x g for 10 minutes at 4°C. Protein concentrations for each sample were calculated using the Pierce BCA Protein Assay Kit (Life Technologies). The protein concentrations were used to calculate the re-suspension volume to normalize all samples for LC-MS analysis to equal (20 µg/µL ) concentration. Isolation of mitochondria from tissues: Purified mitochondria pellets were subsequently vortexed in 500 µL of 99.9% LCMS-grade methanol (Fisher) 0.1% HCl for 10 minutes at 4°C. 500 µL of 100% LCMS-grade hexane (Honeywell) was added to each tube and vortexed for an additional 10 minutes at 4°C. Samples were then centrifuged for 10 minutes at 21,300 x g at 4°C. The top (hexane) layer of the sample (containing UQ and RQ) was transferred to a new labeled 1.5 mL Eppendorf tube and dried in a Refrigerated CentriVap Benchtop Vacuum Concentrator connected to a CentriVap-105 Cold Trap (Labconco). Samples were stored in -80°C freezer prior to resuspension and running on LC-MS. The bottom layer was discarded and the pellet was saved for protein quantification. Protein was isolated as described above. Protein concentrations were calculated using the Pierce BCA Protein Assay Kit (Life Technologies). The protein concentrations were used to calculate the re-suspension volume to normalize all samples for LC-MS analysis to equal (20 µg/uL) concentration. Isolation of UQ/RQ from purified mitochondria: Purified mitochondria pellets were subsequently vortexed in 500 µL of 99.9% LCMS-grade methanol (Fisher) 0.1% HCl for 10 minutes at 4°C. 500 µL of 100% LCMS-grade hexane (Honeywell) was added to each tube and vortexed for an additional 10 minutes at 4°C. Samples were then centrifuged for 10 minutes at 21,300 x g at 4°C. The top (hexane) layer of the sample (containing UQ and RQ) was transferred to a new labeled 1.5 mL Eppendorf tube and dried in a Refrigerated CentriVap Benchtop Vacuum Concentrator connected to a CentriVap-105 Cold Trap (Labconco). Samples were stored in -80°C freezer prior to resuspension and running on LC-MS. The bottom layer was discarded and the pellet was saved for protein quantification. Protein was isolated as described above. Protein concentrations were calculated using the Pierce BCA Protein Assay Kit (Life Technologies). The protein concentrations were used to calculate the re-suspension volume to normalize all samples for LC-MS analysis to equal (20 µg/uL) concentration. |
Combined analysis:
Analysis ID | AN005248 |
---|---|
Analysis type | MS |
Chromatography type | Reversed phase |
Chromatography system | Thermo Vanquish |
Column | Phenomenex Luna PFP(2) (100 x 2mm,3µm) |
MS Type | ESI |
MS instrument type | Orbitrap |
MS instrument name | Thermo Q Exactive Plus Orbitrap |
Ion Mode | POSITIVE |
Units | peak area |
Chromatography:
Chromatography ID: | CH003973 |
Instrument Name: | Thermo Vanquish |
Column Name: | Phenomenex Luna PFP(2) (100 x 2mm,3µm) |
Column Temperature: | 25 |
Flow Gradient: | 0 - 3 min, 30% A; 3 - 3.25 min, 30% - 2% A; 3.25 - 5min, 2% A; 5 - 6min, 2% - 1% A; 6 - 8.75min, 1% A; 8.75 - 9min, 1% - 30% A; 9 - 10min, 30% A |
Flow Rate: | 0.5mL/min |
Solvent A: | 100% water; 0.1% formic acid |
Solvent B: | 100% acetonitrile; 0.1% formic acid |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS004981 |
Analysis ID: | AN005248 |
Instrument Name: | Thermo Q Exactive Plus Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | The mass spectrometer was operated in full scan, positive-ion mode, with the spray voltage set to 3.0 kV, the heated capillary at 275°C, and the HESI probe at 350°C. The sheath gas flow was 40 units, the auxiliary gas flow was 15 units, and the sweep gas flow was 1 unit. MS data was collected in a range of m/z = 200 –1000. The resolution set at 17,500, the AGC target at 3x106, and the maximum injection time at 250 msec. Data acquired by Thermo Fisher's Xcalibur software and analyzed by their Tracefinder software. |
Ion Mode: | POSITIVE |