Summary of Study ST002422
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 PR001559. The data can be accessed directly via it's Project DOI: 10.21228/M85X3W 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 | ST002422 |
| Study Title | UBXD8 lipidomics from whole cells (Part 2) |
| Study Summary | The intimate association between the endoplasmic reticulum (ER) and mitochondrial membranes at ER-mitochondria contact sites (ERMCS) serves as a platform for several critical cellular processes, in particular lipid synthesis. Enzymes involved in lipid biosynthesis are enriched at contacts and membrane lipid composition at contacts is distinct relative to surrounding membranes. How contacts are remodeled and the subsequent biological consequences of altered contacts such as perturbed lipid metabolism remains poorly understood. Here we investigate if the ER-tethered ubiquitin-X domain adaptor 8 (UBXD8) regulates the lipids found in mitochondria-associated membranes (MAM). LC-MS/MS lipidomics found significant changes in distinct lipid species in the MAM fraction of UBXD8 knockout cells. Our results suggest that lipids in MAM are regulated by UBXD8. |
| Institute | University of Arizona |
| Department | Immunobiology |
| Laboratory | Purdy Lab |
| Last Name | Purdy |
| First Name | John |
| Address | PO Box 245221, Tucson, Arizona, 85724, USA |
| purdylab@gmail.com | |
| Phone | 520-626-4371 |
| Submit Date | 2023-01-01 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzXML |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-01-16 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Combined analysis:
| Analysis ID | AN003944 | AN003945 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Thermo Vanquish | Thermo Vanquish |
| Column | Waters ACQUITY UPLC CSH C18 (150 x 2.1mm,1.7um) | Waters ACQUITY UPLC CSH C18 (150 x 2.1mm,1.7um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | peak area | peak area |
MS:
| MS ID: | MS003680 |
| Analysis ID: | AN003944 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Samples were run in a semi-random order where WT or UBXD8 KO samples were interspersed with blank samples. Lipids were ionized using a heated electrospray ionization (HESI) source and nitrogen gas and measured using a Q-Exactive Plus mass spectrometer operating at a MS1 resolution of either 70,000 or 140,000 and a MS2 resolution of 35,000. MS1 Spectra were collected over a mass range of 200 to 1,600 m/z with an automatic gain control (AGC) setting of 1e6 and transient times of 250 ms (70,000 resolution) or 520 ms (140,000 resolution). MS2 spectra were collected using a transient time of 120 ms and an AGC setting of 1e5. Each sample was analyzed using negative and positive ion modes. The mass analyzer was calibrated weekly. SPLASH LIPIDOMIX mass spectrometry standards (Avanti Polar Lipids) were used in determining extraction efficiencies and lipid quantitation. Lipids were identified and quantified using MAVEN, and El-MAVEN (Elucidata). UHPLC retention time, MS1 peaks, and MS2 fragments were used to identify lipids. Lipids were included if they were observed in 3-6 samples in both UBXD8 KO and WT cells. Missing values in a sample were not imputed. The following lipid classes were included in the analysis: cholesteryl esters (CE), diacylglycerol (DG), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylinositol (PI), phosphatidylserine (PS), and triacylglycerol (TG). Guidelines from the Lipidomic Standards Initiative were followed for lipid species identification and quantification, including consideration of isotopic patterns resulting from naturally occurring 13C atoms and isomeric overlap. The following MS2 information was used to confirm each lipid species: PC fragment of 184.073 (positive mode) and tail identification using formic adduct (negative mode); PE fragment of 196.038 or the tail plus 197.046 (negative mode) and neutral loss (NL) of 141.019 (positive mode); PG fragment of 152.996 plus the identification of the FA tails (negative mode) and NL 189.04 of [M+NH4]+ adduct (positive mode); PI fragment of 241.012 (negative) and NL 277.056 of [M+NH4]+ adduct (positive mode); PS NL of 87.032 (negative); DG and TG by NL of FA tails (positive mode); and CE fragment of 369.352 or neutral loss of 368.35 (positive). |
| Ion Mode: | POSITIVE |
| MS ID: | MS003681 |
| Analysis ID: | AN003945 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Samples were run in a semi-random order where WT or UBXD8 KO samples were interspersed with blank samples. Lipids were ionized using a heated electrospray ionization (HESI) source and nitrogen gas and measured using a Q-Exactive Plus mass spectrometer operating at a MS1 resolution of either 70,000 or 140,000 and a MS2 resolution of 35,000. MS1 Spectra were collected over a mass range of 200 to 1,600 m/z with an automatic gain control (AGC) setting of 1e6 and transient times of 250 ms (70,000 resolution) or 520 ms (140,000 resolution). MS2 spectra were collected using a transient time of 120 ms and an AGC setting of 1e5. Each sample was analyzed using negative and positive ion modes. The mass analyzer was calibrated weekly. SPLASH LIPIDOMIX mass spectrometry standards (Avanti Polar Lipids) were used in determining extraction efficiencies and lipid quantitation. Lipids were identified and quantified using MAVEN, and El-MAVEN (Elucidata). UHPLC retention time, MS1 peaks, and MS2 fragments were used to identify lipids. Lipids were included if they were observed in 3-6 samples in both UBXD8 KO and WT cells. Missing values in a sample were not imputed. The following lipid classes were included in the analysis: cholesteryl esters (CE), diacylglycerol (DG), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylinositol (PI), phosphatidylserine (PS), and triacylglycerol (TG). Guidelines from the Lipidomic Standards Initiative were followed for lipid species identification and quantification, including consideration of isotopic patterns resulting from naturally occurring 13C atoms and isomeric overlap. The following MS2 information was used to confirm each lipid species: PC fragment of 184.073 (positive mode) and tail identification using formic adduct (negative mode); PE fragment of 196.038 or the tail plus 197.046 (negative mode) and neutral loss (NL) of 141.019 (positive mode); PG fragment of 152.996 plus the identification of the FA tails (negative mode) and NL 189.04 of [M+NH4]+ adduct (positive mode); PI fragment of 241.012 (negative) and NL 277.056 of [M+NH4]+ adduct (positive mode); PS NL of 87.032 (negative); DG and TG by NL of FA tails (positive mode); and CE fragment of 369.352 or neutral loss of 368.35 (positive). |
| Ion Mode: | NEGATIVE |
