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Ampk β1 2

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AMPK-β1/2 is a protein subunit of the AMP-activated protein kinase (AMPK) complex. AMPK is a cellular energy sensor that plays a crucial role in regulating cellular metabolism and energy homeostasis. The AMPK-β1/2 subunit is responsible for binding the AMPK catalytic subunits and regulatory subunits, thereby facilitating the assembly and activation of the AMPK complex.

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Lab products found in correlation

β actin, by Merck Group (2 mentions) Aicar, by Bio-Techne (1 mentions) Pan ampkα, by Cell Signaling Technology (1 mentions) Praptor ser792, by Cell Signaling Technology (1 mentions) Ampkγ1, by Abcam (1 mentions) Ptbc1d1 ser237, by Merck Group (1 mentions) P acc ser79, by Cell Signaling Technology (1 mentions) P ampk thr172, by Cell Signaling Technology (1 mentions) Tbc1d1, by Cell Signaling Technology (1 mentions) A769662, by Abcam (1 mentions) Epidermal growth factor (egf), by Merck Group (1 mentions) 5 amino 4 imidazole carboxamide riboside aicar, by LGC (1 mentions) Anti rabbit and anti mouse secondary antibody, by Cell Signaling Technology (1 mentions) Ampkα1 2, by Santa Cruz Biotechnology (1 mentions) Myc tag, by Cell Signaling Technology (1 mentions) Rnaimax, by Thermo Fisher Scientific (1 mentions) Anti actin antibody, by Merck Group (1 mentions) Phospho ampkα thr172, by Cell Signaling Technology (1 mentions) Phospho raptor s792, by Cell Signaling Technology (1 mentions) Ampkβ1, by Cell Signaling Technology (1 mentions)

4 protocols using ampk β1 2

1

AMPK Signaling Pathway Profiling

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Unless otherwise stated all reagents were from Sigma–Aldrich. Primary antibodies against pan-AMPK-α (#2532), AMPK-β1/2 (#4150), pAMPK Thr172 (#2531), AKT (#4685), ACC (#3876), pACC Ser79 (#3661), IRAP (#3808), rabbit FLAG (#2368), Raptor (#2280), pRaptor Ser792 (#2083) and TBC1D1 (#5929) were from Cell Signaling Technology. Other antibodies were pan-14-3-3 (sc-629) and GST (sc-138) from Santa Cruz Biotechnology Inc., GFP (118144600001; Roche), AMPK-γ1 (ab32508; Abcam), pTBC1D1 Ser237 (#07-2268; Merck-Millipore), β-actin (A1978) and mouse FLAG (F1804). Isoform-specific AMPK-α antibodies were described previously [34 (link)]. A769662 (Abcam) and AICAR (Tocris) were used.
Thomas E.C., Hook S.C., Gray A., Chadt A., Carling D., Al-Hasani H., Heesom K.J., Hardie D.G, & Tavaré J.M. (2018). Isoform-specific AMPK association with TBC1D1 is reduced by a mutation associated with severe obesity. Biochemical Journal, 475(18), 2969-2983.
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2

Phosphorylation-specific Antibody Assay

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Primary antibodies specific for phospho-AMPKα (172), phospho-HER2 (1221/1222, 1248, 1196), phospho-HER2/phospho-EGFR (1248/1173), phospho-HER3 (1289), phospho-EGFR (1068, 1046/1047S, 1045), phospho-ACC (79), phospho-AMPK substrates (LXRXXpS/pT), AMPKα, AMPKβ1/2, ACC, PARP, HER2, EGFR, HER3, LKB1 and Myc-tag were purchased from Cell Signaling Technology. Anti-rabbit and anti-mouse secondary antibodies were also purchased from Cell Signaling Technology. Anti-phospho-EGFR (1142) was purchased from ECM Biosciences. Anti-Actin antibody was purchased from Sigma-Aldrich. 5-Amino-4-imidazole carboxamide riboside (AICAR), a pharmacological activator of AMPK (Toronto Research Chemicals, Toronto, ON) was dissolved in DMSO to make a 500mM stock concentration. EGF (Sigma) was prepared in a stock concentration of 100μg/ml in purified water. Compound C, an inhibitor of AMPK, was obtained from Calbiochem, and G418, used for selection of transfected cells, was obtained from Invitrogen. Control, AMPKα1/2, AMPKβ1, and AMPKβ2 siRNAs were purchased from Santa Cruz Biotechnology. All siRNA transfections were conducted in triplicate over a period of 72 hours using RNAiMax (Invitrogen) according to the manufacturer's recommendation.
Jhaveri T.Z., Woo J., Shang X., Park B.H, & Gabrielson E. (2015). AMP-activated kinase (AMPK) regulates activity of HER2 and EGFR in breast cancer. Oncotarget, 6(17), 14754-14765.
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3

AMPK Pathway Protein Detection

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ACC (catalog no.: 3676), phospho-ACC1 (Ser79; catalog no.: 3661), AMPKα (catalog no.: 2532), phospho-AMPKα (Thr172; catalog no.: 2535), AMPKβ1 (catalog no.: 4178), Raptor (catalog no.: 2280), phospho-Raptor (S792; catalog no.: 2083), AMPKβ1 (catalog no.: 12063), AMPKβ2 (catalog no.: 4148), and AMPKβ1/2 (catalog no.: 4150) were obtained from Cell Signaling Technology. Antibody against AMPKγ1 (catalog no.: ab32508) was purchased from Abcam, and tubulin (catalog no.: T6074), FLAG (catalog no.: F7425), and β-actin (catalog no.: A2228) were purchased from MilliporeSigma. HTRF assay for detection of phospho-ACC was obtained from Cisbio (catalog no.: 64ACCPET).
Sanders M.J., Ratinaud Y., Neopane K., Bonhoure N., Day E.A., Ciclet O., Lassueur S., Naranjo Pinta M., Deak M., Brinon B., Christen S., Steinberg G.R., Barron D, & Sakamoto K. (2022). Natural (dihydro)phenanthrene plant compounds are direct activators of AMPK through its allosteric drug and metabolite–binding site. The Journal of Biological Chemistry, 298(5), 101852.
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4

Immunofluorescence Staining of AMPK Subunits

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Cells were grown to 70% confluency on poly-lysine coated cover slips. After treatment, immunofluorescent staining was performed following published procedures [5] (link). Antibody dilutions were: AMPK-α1 and AMPK-α2 (1:1000; Upstate 07-350; 07-363; or 1:2000; Bethyl A300-507A, A300-508A), AMPK-β1/2 (1:400; Cell Signaling ♯4150), AMPK-γ1 (1:200; Cell Signaling ♯4187), AMPK-γ2 (1:200; Cell Signaling ♯2536), HuR (1:2000; sc-5261), G3BP1 (1:1000; BD Biosciences ♯611126). In brief, cells were fixed with 3.7% formaldehyde in phosphate-buffered saline (PBS), permeabilized with 0.1% Triton X-100 in PBS/2 mg/BSA and blocked for 1 h in PBS containing 5% fetal bovine serum (FBS), 0.05% Tween-20 and 1 mM NaN3. Samples were incubated overnight with primary antibodies diluted in PBS/FBS/Tween. Purified Alexa Fluor® 488, Alexa Fluor® 647 and Cy3 fluorescently-labeled secondary antibodies were added the following day for 2 h [6] (link). Nuclei were visualized with 1 µg/ml 4׳,6 Diamidino-2-phenylindole dihydrochloride (DAPI).
Mahboubi H., Barisé R, & Stochaj U. (2015). Data in support of 5′AMP-activated protein kinase alpha regulates stress granule biogenesis. Data in Brief, 4, 54-59.
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