Summary of Study ST002850

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 PR001784. The data can be accessed directly via it's Project DOI: 10.21228/M83T50 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.

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Study IDST002850
Study TitleBap1 Promotes Osteoclast Function by Metabolic Reprogramming
Study TypeUntargeted Metabolomics
Study SummaryTreatment of osteoporosis most commonly diminishes osteoclast number which suppresses bone formation thus compromising fracture prevention. Bone formation is not suppressed, however, when bone degradation is reduced by retarding osteoclast functional resorptive capacity, rather than differentiation. We find deletion of deubiquitinase, BRCA1-associated protein 1 (Bap1), in myeloid cells (Bap1∆LysM), arrests osteoclast function but not formation. Bap1∆LysM osteoclasts fail to organize their cytoskeleton which is essential for bone degradation. Consequently, bone mass increases in the mutant mice. We find the deubiquitinase activity of Bap1 regulates osteoclast function by metabolic reprogramming. Bap1 deficient osteoclast lineage cells upregulate the cystine transporter, Slc7a11, by enhanced H2Aub occupancy of its promoter. SLC7A11 regulates cellular ROS levels and redirects the mitochondrial metabolites away from the TCA cycle, both of which are necessary for osteoclast function. Thus in osteoclasts, Bap1 appears to regulate epigenetic-metabolic axis and is a potential target to reduce bone degradation while maintaining osteogenesis in osteoporotic patients.
Institute
Washington University in St. Louis
DepartmentPathology and Immunology, Medicine, Chemistry
LaboratoryTeitelbaum and Patti Laboratories
Last NameCho
First NameKevin
Address1 Brookings Drive, Campus Box 1134, St. Louis, MO, 63130, USA
Emailkevin.cho@wustl.edu
Phone314-935-8813
Submit Date2023-08-26
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailLC-MS
Release Date2023-09-11
Release Version1
Kevin Cho Kevin Cho
https://dx.doi.org/10.21228/M83T50
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR001784
Project DOI:doi: 10.21228/M83T50
Project Title:Bap1 Promotes Osteoclast Function by Metabolic Reprogramming
Project Type:Untargeted Metabolomics
Project Summary:Treatment of osteoporosis most commonly diminishes osteoclast number which suppresses bone formation thus compromising fracture prevention. Bone formation is not suppressed, however, when bone degradation is reduced by retarding osteoclast functional resorptive capacity, rather than differentiation. We find deletion of deubiquitinase, BRCA1-associated protein 1 (Bap1), in myeloid cells (Bap1∆LysM), arrests osteoclast function but not formation. Bap1∆LysM osteoclasts fail to organize their cytoskeleton which is essential for bone degradation. Consequently, bone mass increases in the mutant mice. We find the deubiquitinase activity of Bap1 regulates osteoclast function by metabolic reprogramming. Bap1 deficient osteoclast lineage cells upregulate the cystine transporter, Slc7a11, by enhanced H2Aub occupancy of its promoter. SLC7A11 regulates cellular ROS levels and redirects the mitochondrial metabolites away from the TCA cycle, both of which are necessary for osteoclast function. Thus in osteoclasts, Bap1 appears to regulate epigenetic-metabolic axis and is a potential target to reduce bone degradation while maintaining osteogenesis in osteoporotic patients.
Institute:Washington University in St. Louis
Department:Pathology and Immunology, Medicine, Chemistry
Laboratory:Teitelbaum and Patti Laboratories
Last Name:Cho
First Name:Kevin
Address:1 Brookings Drive, Campus Box 1134, St. Louis, MO, 63130, USA
Email:kevin.cho@wustl.edu
Phone:314-935-8813

Subject:

Subject ID:SU002962
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606

Factors:

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

mb_sample_id local_sample_id Genotype
SA308592Pos_KO_4Knockout
SA308593Pos_KO_5Knockout
SA308594Neg_KO_1Knockout
SA308595Pos_KO_3Knockout
SA308596Neg_KO_2Knockout
SA308597Pos_KO_2Knockout
SA308598Neg_KO_5Knockout
SA308599Neg_KO_4Knockout
SA308600Neg_KO_3Knockout
SA308601Pos_KO_1Knockout
SA308602Neg_WT_5Wild-type
SA308603Neg_WT_4Wild-type
SA308604Neg_WT_1Wild-type
SA308605Pos_WT_3Wild-type
SA308606Pos_WT_2Wild-type
SA308607Pos_WT_4Wild-type
SA308608Pos_WT_5Wild-type
SA308609Neg_WT_2Wild-type
SA308610Pos_WT_1Wild-type
SA308611Neg_WT_3Wild-type
Showing results 1 to 20 of 20

Collection:

Collection ID:CO002955
Collection Summary:Primary mus musculus cells
Sample Type:Osteoclast

Treatment:

Treatment ID:TR002971
Treatment Summary:All in vitro experiments were performed at least 3 times. Primary bone marrow macrophages (BMMs) were prepared as described. Marrow was extracted from femora and tibiae of 6- to 8-week-old mice with α minimum essential medium (α-MEM) and cultured in α-MEM containing 10% inactivated fetal bovine serum, 100 IU/mL penicillin, and 100 μg/mL streptomycin (α-10 medium) with 1:10 of mMCSF producing cell line, CMG 14-12 condition media on petri-plastic dishes. Cells were incubated at 37°C in 5% CO2 for 3 days and then washed with phosphate-buffered saline (PBS) and lifted with 1X trypsin/EDTA in PBS. A total of 1.2 × 104 BMMs were cultured in 500 μL α-MEM containing 10% heat-inactivated fetal bovine serum with glutathione-S transferase–RANKL and 30 ng/mL of mouse recombinant macrophage colony-stimulating factor (M-CSF) in 48-well tissue culture plates, some containing sterile bovine bone slices.

Sample Preparation:

Sampleprep ID:SP002968
Sampleprep Summary:Cells were quenched with cold LC/MS-grade methanol, then scraped and transferred to Eppendorf tubes. Samples were dried in a SpeedVac. The samples were then reconstituted in 1 mL of cold methanol:acetonitrile:water (2:2:1) and subjected to three cycles of vortexing, freezing in liquid nitrogen, and 10 min of sonication at 25 °C. Samples were stored at −20 °C overnight and then centrifuged for 10 min at 14,000×g and 4 °C. Supernatants were transferred to new tubes and dried by a SpeedVac. The protein abundance of each sample was determined by using BCA. A quantity of 1 μl of acetonitrile:water (2:1) per each 2.5 μg of protein was used. Samples were subjected to two cycles of vortexing and 10 min of sonication at 25 °C. Next, samples were centrifuged for 10 min at 14,000×g and 4 °C, transferred supernatant to LC vials, and stored at −80 °C until MS analysis

Combined analysis:

Analysis ID AN004668 AN004669
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Thermo Vanquish Flex UHPLC Systems Thermo Vanquish Flex UHPLC Systems
Column HILICON iHILIC-(P) Classic (100 x 2.1mm,5um) HILICON iHILIC-(P) Classic (100 x 2.1mm,5um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Orbitrap ID-X Tribrid Thermo Orbitrap ID-X Tribrid
Ion Mode NEGATIVE POSITIVE
Units peak area peak area

Chromatography:

Chromatography ID:CH003514
Instrument Name:Thermo Vanquish Flex UHPLC Systems
Column Name:HILICON iHILIC-(P) Classic (100 x 2.1mm,5um)
Column Temperature:45
Flow Gradient:0–1 min: 90% B, 1–12 min: 90-35% B, 12–12.5 min: 35-25% B, 12.5–14.5 min: 25% B
Flow Rate:250 uL/min
Solvent A:20 mM ammonium bicarbonate, 0.1% ammonium hydroxideand 2.5 μM medronic acid in water:acetonitrile (95:5)
Solvent B:acetonitrile:water (95:5)
Chromatography Type:HILIC

MS:

MS ID:MS004415
Analysis ID:AN004668
Instrument Name:Thermo Orbitrap ID-X Tribrid
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Data were collected with the following MS source settings: spray voltage, -2.8 kV; sheath gas, 50; auxiliary gas, 10; sweep gas, 1; ion transfer tube temperature, 300°C; vaporizer temperature, 200°C; mass range, 67 – 1000 Da; resolution, 120,000; maximum injection time, 200 ms; isolation window, 1.5 Da. XCMS and Skyline software were used for data processing
Ion Mode:NEGATIVE
  
MS ID:MS004416
Analysis ID:AN004669
Instrument Name:Thermo Orbitrap ID-X Tribrid
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Data were collected with the following MS source settings: spray voltage, 3.5 kV; sheath gas, 50; auxiliary gas, 10; sweep gas, 1; ion transfer tube temperature, 300°C; vaporizer temperature, 200°C; mass range, 67 – 1000 Da; resolution, 120,000; maximum injection time, 200 ms; isolation window, 1.5 Da. XCMS and Skyline software were used for data processing
Ion Mode:POSITIVE
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