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.
| Study ID | ST002850 |
| Study Title | Bap1 Promotes Osteoclast Function by Metabolic Reprogramming |
| Study Type | Untargeted Metabolomics |
| Study 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 |
| kevin.cho@wustl.edu | |
| Phone | 314-935-8813 |
| Submit Date | 2023-08-26 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2023-09-11 |
| Release Version | 1 |
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 |
|---|---|---|
| SA308592 | Pos_KO_4 | Knockout |
| SA308593 | Pos_KO_5 | Knockout |
| SA308594 | Neg_KO_1 | Knockout |
| SA308595 | Pos_KO_3 | Knockout |
| SA308596 | Neg_KO_2 | Knockout |
| SA308597 | Pos_KO_2 | Knockout |
| SA308598 | Neg_KO_5 | Knockout |
| SA308599 | Neg_KO_4 | Knockout |
| SA308600 | Neg_KO_3 | Knockout |
| SA308601 | Pos_KO_1 | Knockout |
| SA308602 | Neg_WT_5 | Wild-type |
| SA308603 | Neg_WT_4 | Wild-type |
| SA308604 | Neg_WT_1 | Wild-type |
| SA308605 | Pos_WT_3 | Wild-type |
| SA308606 | Pos_WT_2 | Wild-type |
| SA308607 | Pos_WT_4 | Wild-type |
| SA308608 | Pos_WT_5 | Wild-type |
| SA308609 | Neg_WT_2 | Wild-type |
| SA308610 | Pos_WT_1 | Wild-type |
| SA308611 | Neg_WT_3 | Wild-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 |
