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Pgc1β

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PGC1β is a protein that plays a key role in the regulation of cellular energy metabolism. It is involved in the control of mitochondrial biogenesis and function, as well as the expression of genes related to energy production and utilization.

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Lab products found in correlation

Pgc 1α, by Abcam (3 mentions) Tissue homogenizer, by Omni International (2 mentions) Westernbright ecl kit solution, by Advansta (2 mentions) Hrp conjugated secondary antibody, by Cell Signaling Technology (2 mentions) Gapdh, by Biosynth (2 mentions) Protease inhibitor, by Thermo Fisher Scientific (2 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Ne per nuclear and cytoplasmic extraction reagent, by Thermo Fisher Scientific (2 mentions) Rpn2106, by GE Healthcare (1 mentions) Phospho nf kb p65, by Cell Signaling Technology (1 mentions) Nf kb p65, by Cell Signaling Technology (1 mentions) Phospho akt, by Cell Signaling Technology (1 mentions) P38 mapk, by Cell Signaling Technology (1 mentions) Phospho p38 mapk, by Cell Signaling Technology (1 mentions) β actin, by Merck Group (1 mentions) Horse serum, by Thermo Fisher Scientific (1 mentions) Trypsin, by Thermo Fisher Scientific (1 mentions) Uqcrc2, by Abcam (1 mentions) Ndufb8, by Abcam (1 mentions) Xf palmitate bsa fao substrate, by Agilent Technologies (1 mentions)

9 protocols using pgc1β

1

Western Blot Analysis of Signaling Proteins

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Western blotting was performed following standard protocols. Primary antibodies reactive to phospho-NF-kB p65 (1:1000; Cell Signaling, 3033), NF-kB p65 (1:1000; Cell Signaling, 4764), phospho-AKT (1:500; Cell Signaling, 4060), AKT (1:1000; Cell Signaling, 4685), phospho-p38 MAPK (1:800; Cell Signaling, 4631), p38 MAPK (1:1000; Cell Signaling, 9212), PGC1β (1:1000; Abcam, 176328), and β-actin (1:20,000; Sigma, F3022) were used. Primary antibody binding was detected by enhanced chemoluminescence (GE Healthcare, RPN2106).
Rao S., Sigl V., Wimmer R.A., Novatchkova M., Jais A., Wagner G., Handschuh S., Uribesalgo I., Hagelkruys A., Kozieradzki I., Tortola L., Nitsch R., Cronin S.J., Orthofer M., Branstetter D., Canon J., Rossi J., D'Arcangelo M., Botling J., Micke P., Fleur L.L., Edlund K., Bergqvist M., Ekman S., Lendl T., Popper H., Takayanagi H., Kenner L., Hirsch F.R., Dougall W, & Penninger J.M. (2017). RANK rewires energy homeostasis in lung cancer cells and drives primary lung cancer. Genes & Development, 31(20), 2099-2112.
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2

Adipose Tissue Protein Expression

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20mg of brown and white adipose tissue was homogenized with a tissue homogenizer (Omni) in a small volume of RIPA buffer containing protease inhibitors. Lysates were cleared by spinning in a refrigerated centrifuge. Protein concentration was determined by performing a BCA Assay (Pierce, Rockford, IL). 1μg of each protein lysate was boiled under reducing conditions and subjected to SDS-PAGE. Protein was transferred onto PVDF membrane and then blocked in 5% nonfat milk in TBST (Tris-buffered saline with 0.5% Tween 20) for 1hr at room temperature, then blotted overnight at 4°C with the following antibodies at concentrations suggested by the manufacturer: UCP-1 (Abcam, Cambridge, MA ), PGC1-α (Abcam), PGC1-β (Abcam), and GAPDH (Fitzgerald, Acton, MA). Membranes were probed with HRP-conjugated secondary antibodies at a 1:5000 dilution for 1hr at room temperature (Cell Signaling, Danvers, MA), washed, submersed in WesternBright ECL kit solution (Advansta, Menlo Park, CA) and exposed to autoradiography film.
Chrysovergis K., Wang X., Kosak J., Lee S.H., Kim J.S., Foley J.F., Travlos G., Singh S., Baek S.J, & Eling T.E. (2014). NAG-1/GDF15 prevents obesity by increasing thermogenesis, lipolysis and oxidative metabolism. International journal of obesity (2005), 38(12), 1555-1564.
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3

Mitochondrial Protein Expression Analysis

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The Eu was purchased from National Institutes for Food and Drug Control (China). Dulbecco’s modified Eagle medium (DMEM), Nutrient Mixture F-12 (DMEM/F12) media, horse serum, trypsin and penicillin/streptomycin, were obtained from Life Technologies, GibcoBRL (Rockville,MD,USA). Antibodies against CPT-1C and Tomm20 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA,USA). Antibodies against ERRα and c-Myc were obtained from Cell Signaling Technology (Beverly, MA,USA) and an antibody against the HRAS, PGC-1β, NDUFB8, SDH-B, UQCRC2, ATP5A, COX5A, MCAD and PPARα were obtained from Abcam (Cambridge, MA, USA). 10058F4 were purchased from Sigma-Aldrich (St.Lois, MO, USA). XF Palmitate-BSA FAO Substrate was obtained from seahorse Bioscience (North Billerica, MA, USA).
Yan X., Zhang G., Bie F., Lv Y., Ma Y., Ma M., Wang Y., Hao X., Yuan N, & Jiang X. (2017). Eugenol inhibits oxidative phosphorylation and fatty acid oxidation via downregulation of c-Myc/PGC-1β/ERRα signaling pathway in MCF10A-ras cells. Scientific Reports, 7, 12920.
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4

Western Blot Analysis of Protein Biomarkers

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Protein extracts (40∼80 μg) were electrophoresed at 80 volts on 10% or 12% SDS/PAGE, and then transferred onto nitrocellulose filter membrane (PALL BioTrace, Port Washington, NY, USA). Immunoblotting was performed according to the manufacturer’s instructions using the following antibodies: brain natriuretic peptide (BNP, abcam), ANP32E (abcam), FGF1 (abcam), Smad3 (Cell Signalling Technology), Phospho-Smad3 (CST), GAPDH (Zhong Shan Golden Bridge ZSGB), PGC1α (abcam), PGC1β (abcam), FGFR1 (abcam), Mac-2(Santa Cruz Biotechnology), IL-1β (abcam, Catalogue: ab9722), and NF-κB p65 (abcam). A Bio-Rad Imager facility was used to detect the expression of proteins.
Li G., Shao Y., Guo H.C., Zhi Y., Qiao B., Ma K., Du J., Lai Y.Q, & Li Y. (2021). MicroRNA-27b-3p down-regulates FGF1 and aggravates pathological cardiac remodelling. Cardiovascular Research, 118(9), 2139-2151.
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5

Investigating Metabolic and Oxidative Stress Responses

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Purified CD4 T cells were treated with 5 μM KML001 or DPBS control for different times and nucleofected cells were harvested and lysed on ice in RIPA lysis buffer (Boston BioProducts Inc, Ashland, MA) in the presence of protease inhibitors (Thermo Fisher Scientific). The protein concentrations were measured by the Pierce BCA protein assay kit (Thermo Fisher Scientific). The proteins were separated by SDS-PAGE and transferred to polyvinylidene difluoride membranes. The membranes were blocked with 5% non-fat milk, 0.1% Tween-20 in Tris-buffered saline (TBS), and then incubated overnight with primary antibodies for Fasn, PGC1α, NRF1, Pepck, G-6-PD, hmgcs1, ERRα, GPAT, mtTFA, GPx, SOD1, γH2AX, PARP1, TRF2, β-Actin (Cell Signaling, Danvers, MA), PGC1β, HMGCR, DGAT1, PPARα, ACADL, ACADM (Abcam), p53 (Santa Cruz Biotechnology, Dallas, TX), and NEIL1, XRCC1 (Novus Biologicals). Appropriate horseradish peroxide-conjugated secondary antibodies (Cell Signaling) were then used, and the proteins were visualized with the Amersham ECL Prime Western Blotting Detection Reagent (GE Healthcare Bio-Sciences, Pittsburgh, PA). The protein bands were captured and quantitatively analyzed by Chemi DocTM MP Imaging System (Bio-Rad).
Schank M., Zhao J., Wang L., Li Z., Cao D., Nguyen L.N., Dang X., Khanal S., Nguyen L.N., Thakuri B.K., Ogbu S.C., Lu Z., Zhang J., Wu X.Y., Morrison Z.D., El Gazzar M., Ning S., Moorman J.P, & Yao Z.Q. (2020). Telomeric injury by KML001 in human T cells induces mitochondrial dysfunction through the p53-PGC-1α pathway. Cell Death & Disease, 11(12), 1030.
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6

Characterization of Cellular Bioenergetics

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All chemicals, including tosyl phenylalanyl chloromethyl ketone (TPCK)-treated trypsin, were from Thermo Scientific, unless otherwise stated. Acetonitrile (CH3CN, #15611400, Honeywell Riedel-de Haen), formic acid (FA, #10627431) and LC-MS grade water (#15651400, Honeywell Riedel-de Haen) were from Fisher Scientific. MC2494 was prepared as previously reported (17 (link)). Antibodies: PGC1α (#ab191838), PGC1β (#ab176328), and SOD2 (#ab13533) were from Abcam and GAPDH was from Santa Cruz (#sc-47724). Cell lines: U937 (#ACC5) human myeloid leukemia cells were purchased from DSMZ and MCF7 (#ICLCHTL95021) breast cancer cells from Cell Bank Interlab Cell Line Collection. U937 and MCF7 cells were propagated in RPMI (Euroclone #ECB9006) and DMEM (Euroclone #ECB7501), respectively, with 10% fetal bovine serum (FBS; Gibco #10270), 2 mM L-glutamine (Euroclone #ECB3000D), and antibiotics (100 U/mL penicillin, 100 μg/mL streptomycin, Euroclone #ECB3001D, and 250 ng/mL amphotericin-B; Euroclone #ECM0009). All cell lines were grown at 37°C with 5% CO2, and were then tested, authenticated, and used for 10–20 passages. Mycoplasma contamination was checked using EZ-PCR Mycoplasma Test Kit (Biological Industries #20-700-20).
Carafa V., Russo R., Della Torre L., Cuomo F., Dell'Aversana C., Sarno F., Sgueglia G., Di Donato M., Rotili D., Mai A., Nebbioso A., Cobellis G., Chambery A, & Altucci L. (2020). The Pan-Sirtuin Inhibitor MC2494 Regulates Mitochondrial Function in a Leukemia Cell Line. Frontiers in Oncology, 10, 820.
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7

Adipose Tissue Protein Expression

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20mg of brown and white adipose tissue was homogenized with a tissue homogenizer (Omni) in a small volume of RIPA buffer containing protease inhibitors. Lysates were cleared by spinning in a refrigerated centrifuge. Protein concentration was determined by performing a BCA Assay (Pierce, Rockford, IL). 1μg of each protein lysate was boiled under reducing conditions and subjected to SDS-PAGE. Protein was transferred onto PVDF membrane and then blocked in 5% nonfat milk in TBST (Tris-buffered saline with 0.5% Tween 20) for 1hr at room temperature, then blotted overnight at 4°C with the following antibodies at concentrations suggested by the manufacturer: UCP-1 (Abcam, Cambridge, MA ), PGC1-α (Abcam), PGC1-β (Abcam), and GAPDH (Fitzgerald, Acton, MA). Membranes were probed with HRP-conjugated secondary antibodies at a 1:5000 dilution for 1hr at room temperature (Cell Signaling, Danvers, MA), washed, submersed in WesternBright ECL kit solution (Advansta, Menlo Park, CA) and exposed to autoradiography film.
Chrysovergis K., Wang X., Kosak J., Lee S.H., Kim J.S., Foley J.F., Travlos G., Singh S., Baek S.J, & Eling T.E. (2014). NAG-1/GDF15 prevents obesity by increasing thermogenesis, lipolysis and oxidative metabolism. International journal of obesity (2005), 38(12), 1555-1564.
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8

Profiling Monocyte Protein Expression

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Cytoplasmic or nuclear proteins from human peripheral blood CD14+ monocytes were extracted separately using nuclear protein extraction kits (Pierce). Target proteins from cytoplasm were detected with primary antibodies to TRAP, DC‐STAMP, cathepsin K (1:1,000; all from Abcam), matrix metalloproteinase 9 (MMP‐9), TRAF6, phospho‐p38, p38, phospho‐ERK1/2, ERK1/2, phospho‐JNK, JNK, and β‐tubulin (1:1,000; all from Cell Signaling Technology). Target proteins from nuclei were detected with primary antibodies to NFATc1, c‐Fos, c‐Jun, and fibrillarin (1:1,000; all from Cell Signaling Technology), as well as PGC‐1β (1:1,000; Abcam). Protein bands were visualized using enhanced chemiluminescence (Millipore) plus Western blot detection reagents, followed by exposure to a scanning imager (G:BOX Gel & Blot Imaging Series; Syngene) 26.
Ma J., Jing J., Wang J., Mo Y., Li Q., Lin J., Chen L., Shao L., Miossec P, & Dai L. (2019). Activation of the Peroxisome Proliferator–Activated Receptor γ Coactivator 1β/NFATc1 Pathway in Circulating Osteoclast Precursors Associated With Bone Destruction in Rheumatoid Arthritis. Arthritis & Rheumatology (Hoboken, N.j.), 71(8), 1252-1264.
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9

Western Blot Analysis of Nuclear Proteins

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Nuclear proteins were extracted using NE-PER nuclear and cytoplasmic extraction reagents (Thermo Fisher Scientific) with protease inhibitors (Roche). Ten μg of proteins was separated onto precast 4% to 12% polyacrylamide gels (Sigma-Aldrich) and transferred to 0.2-μm PVDF membranes using a trans-blot turbo system (Bio-Rad). Blots were blocked at room temperature for 2 hours in blocking buffer [5% nonfat dry milk or 5% BSA in TBS/0.1% Tween 20 (Sigma-Aldrich)], then incubated overnight at 4°C with the appropriate primary antibody: PGC1α (1:1,000, Novus Biologicals; cat. #NBP1-04676, RRID:AB_1522118), PGC1β (1:10,000, Abcam; cat. #ab176328, RRID:AB_2893194), Histone H3 (1:2,000, Abcam; cat. #ab1791, RRID:AB_302613). Antibody binding was detected using anti-rabbit IgG-HRP (1:5,000, Sigma-Aldrich; cat. #A6154, RRID:AB_258284) for 1 hour at room temperature. The signals were revealed with ECL (Thermo Fisher Scientific) and acquired with an Odyssey Fc Imaging System. Histone H3 was used for normalization.
Ghilardi C., Moreira-Barbosa C., Brunelli L., Ostano P., Panini N., Lupi M., Anastasia A., Fiordaliso F., Salio M., Formenti L., Russo M., Arrigoni E., Chiaradonna F., Chiorino G., Draetta G., Marszalek J.R., Vellano C.P., Pastorelli R., Bani M., Decio A, & Giavazzi R. (2022). PGC1α/β Expression Predicts Therapeutic Response to Oxidative Phosphorylation Inhibition in Ovarian Cancer. Cancer Research, 82(7), 1423-1434.
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