Phospho acc

Manufactured by Cell Signaling Technology

Sourced in United States, United Kingdom

Phospho-ACC is a laboratory product that detects the phosphorylation of acetyl-CoA carboxylase (ACC), a key enzyme involved in fatty acid metabolism. This product is designed to help researchers study the regulation of ACC activity in various biological systems.

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Lab products found in correlation

Close spelling variants of this product

Phospho-ACC (Ser79) (39 citations) Anti-phospho-ACC (20 citations) Anti-phospho-ACC Ser79 (15 citations) Anti-phospho-ACC-S79 (4 citations) Phospho-ACC (S79) (3 citations) Phospho-ACC (Serine 79) (2 citations) Phospho-Acetyl-CoA carboxylase (p-ACC) (2 citations) Anti-phospho acetyl-CoA carboxylase (p-ACC) (2 citations)

53 protocols using phospho acc

Adipocyte Differentiation Assay

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d-glucose, 3-isobutyl-1-methylxanthine, dexamethasone, insulin, ethanol (EtOH), Oil-Red O, propylene glycol, dimethyl sulfoxide (DMSO), pioglitazone, metformin, dorsomorphin (an AMPK inhibitor, AI), and all other chemicals were of analytical grade and obtained from Sigma-Aldrich Inc. (St. Louis, MO, USA). Normal glucose (100 mg/dL) Dulbecco’s Modified Eagle Medium (DMEM; Catalog No.: 12320), penicillin/streptomycin (P/S), fetal bovine serum (FBS), horse serum, and trypsin-EDTA were bought from Invitrogen (Carlsbad, CA, USA). Rosiglitazone used in this experiment did not contain any inactive ingredients, and the pure compound was kindly provided by GlaxoSmithKline, Ltd. (Taipei, Taiwan). Phospho-AMPK, AMPK, phospho-ACC, ACC, phospho-AKT, AKT, PPARγ, C/EBPα, C/EBPβ, and GAPDH antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA).

Hsieh C.T., Chang F.R., Tsai Y.H., Wu Y.C, & Hsieh T.J. (2018). 2-Bromo-4′-methoxychalcone and 2-Iodo-4′-methoxychalcone Prevent Progression of Hyperglycemia and Obesity via 5′-Adenosine-Monophosphate-Activated Protein Kinase in Diet-Induced Obese Mice. International Journal of Molecular Sciences, 19(9), 2763.

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Immunoblot Analysis of Cell Signaling Pathways

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CD8⁺ T cells were lysed with RIPA buffer (50mM Tris-HCl, pH 8.0, 1% NP-40, 0.5% Sodium deoxycholate, and 150mM NaCl) with addition of Halt Protease & Phosphatase Inhibitor Cocktail (Thermo Scientific, Waltham, Massachusetts, U.S.A.) and protein concentration was determined using the Bio-Rad protein assay (Hercules, California, U.S.A.). Proteins were resuspended in LDS Sample Buffer, loaded (15 μg/lane) onto NuPAGE Bis-Tris gels, and transferred to polyvinylidene difluoride (PVDF) membranes using iBlot2 system (LifeTechnologies, Grand Island, New York, U.S.A.). Proteins were detected using rabbit anti-S6K, phospho-S6K, AMPKα, phospho-AMPKα, Acetyl-CoA carboxylase (ACC), phospho-ACC, cyclin E, cdk2, phospho-Rb (Ser780), tubulin, and anti-rabbit IgG HRP-linked antibody from Cell Signaling Technology (Danvers, Massachusetts, U.S.A.). Primary antibodies were used at a 1:1,000 dilution whereas secondary antibody was used at a 1:50,000 dilution. PVDF membranes were developed using ECL Western Blotting Substrate (Thermo Scientific, Waltham, Massachusetts, U.S.A.) and exposed to Amersham hyperfilm ECL (GE Healthcare Life Sciences, Piscataway, New Jersey, U.S.A.)

Lee P.H., Yamada T., Park C.S., Shen Y., Puppi M, & Lacorazza H.D. (2015). G0S2 modulates homeostatic proliferation of naïve CD8+ T cells and inhibits oxidative phosphorylation in mitochondria. Immunology and cell biology, 93(7), 605-615.

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Immunoblotting and Antibody Detection

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Immunoblotting was performed as described previously (Jacobs et al., 2008). Primary antibodies were followed by mouse- or rabbit-conjugated horseradish peroxidase (HRP). HRP-conjugated antibodies (anti-mouse or anti-rabbit IgG HRP conjugate, Promega) were detected by enhanced chemiluminescence detection (Thermofisher). This included the following antibodies: ACC (3662, Cell Signaling), phospho-ACC (3661, Cell Signaling), pan-AMPKα (2532, Cell Signaling), phospho-AMPKα (2535, Cell Signaling), 4EBP1 (9644, Cell Signaling), phospho-4EBP1 (2855, Cell Signaling), c-myc (9402, Cell Signaling), phospho-mTOR (5536, Cell Signaling), Activated Notch (ab8925, Abcam), RAPTOR (2280, Cell Signaling), phospho-RAPTOR (2083, Cell Signaling), S6 (2217, Cell Signaling), phospho-p70 S6K (9204, Cell Signaling), p70 S6K (2708, Cell Signaling), phosphor-TSC2 (5584, Cell Signaling), TSC2 (3612, Cell Signaling). Alternatively, primary antibodies were followed by fluorescently labeled anti-mouse or rabbit antibodies (LiCor) and imaged using the Odyssey infrared imaging system (LiCor). This included the following antibodies: Glut1 (ab652, Abcam), hexokinase 2 (2867, Cell Signaling), hexokinase 1 (ab104835, Abcam), cytochrome C (556433, BD Biosciences), β-actin (A5441, Sigma), phospho-S6 (4858, Cell Signaling). Western blots were quantified using ImageJ software.

Kishton R.J., Barnes C.E., Nichols A.G., Cohen S., Gerriets V.A., Siska P.J., Macintyre A.N., Goraksha-Hicks P., de Cubas A.A., Liu T., Warmoes M.O., Abel E.D., Yeoh A.E., Gershon T.R., Rathmell W.K., Richards K.L., Locasale J.W, & Rathmell J.C. (2016). AMPK is essential to balance glycolysis and mitochondrial metabolism to control T-ALL cell stress and survival. Cell metabolism, 23(4), 649-662.

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Elaiophylin Biosynthesis from Deep-Sea Streptomyces

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Elaiophylin was prepared from the deep-sea-derived Streptomyces sp. SCSIO 1934 [44 (link)]. Rapamycin, torin1, bafilomycin A1, compound C, STO-609, and rosiglitazone were purchased from Selleck. Insulin, IBMX, and dexamethasone were purchased from Sigma-Aldrich. Antibodies recognizing phospho-AMPKα (#2535), AMPKα (# 2532), phospho-ACC (#3661), ACC (#3662), phospho-AKT (#4060), phospho-S6K (#9204), S6K (#9202), 4EBP1 (#9644), phospho-ERK (#4370), phospho-c-JUN (#3270), and LC3B (#3868) were procured from Cell Signaling. Other antibodies included anti-AKT (Proteintech, 60203-2-Ig), anti-GST (Proteintech, 10000-0-AP), anti-TXNIP (Abcam, ab188865), and anti-ACTIN (Genescript, A00702).

Bao R., Meng Y., Zhang H., Yang C., Li W., Zhang C., Zhang J., Sun R., Li Z., Jiang W., Zhang C., Zhang C., Yuan H.X, & Dang Y. (2021). Elaiophylin reduces body weight and lowers glucose levels in obese mice by activating AMPK. Cell Death & Disease, 12(11), 972.

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Western Blot Analysis of Protein Signaling

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Protein lysates were prepared in either RIPA or CHAPS buffer and quantified using the BCA Protein Assay (Thermo Scientific). Protein was separated on 4%–20% SDS-PAGE gels, transferred to PVDF membranes, and probed with antibodies against CPS1 (ab3682), β-actin (ab8227), ASS1 (clone 2B10, ab124465), cyclophilin B (clone EPR12703(B), ab178397), total AMPK (Cell Signaling, #2603), phospho-AMPK (Cell signaling, #2531), total ACC (Cell Signaling, #3662), phospho-ACC (Cell Signaling, #11818), LKB1 (Cell Signaling, #3050), γH2AX (Cell Signaling, #9718), total CAD (Cell Signaling, #11933), phospho-CAD (Cell Signaling, #12662), NOS3 (BD, 610298), phospho-S6 (Cell Signaling, #2211), phospho-4E-BP1 (Cell Signaling, #2855), CREB1 (Santa Cruz, sc-186X), FOXA1 (ab23738), TEAD4 (ab58310).

Kim J., Hu Z., Cai L., Li K., Choi E., Faubert B., Bezwada D., Rodriguez-Canales J., Villalobos P., Lin Y.F., Ni M., Huffman K.E., Girard L., Byers L.A., Unsal-Kacmaz K., Peña C.G., Heymach J.V., Wauters E., Vansteenkiste J., Castrillon D.H., Chen B.P., Wistuba I., Lambrechts D., Xu J., Minna J.D, & DeBerardinis R.J. (2017). CPS1 maintains pyrimidine pools and DNA synthesis in KRAS/LKB1-mutant lung cancer cells. Nature, 546(7656), 168-172.

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Western Blot Analysis of Hepatic Proteins

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Liver lysates were made by homogenizing pulverized, freeze-clamped liver tissue in lysis buffer (50 mM Tris-HCl pH 7.4, 1 mM EGTA, 150 mM NaCl, 5 mM MgCl_2, 1% NP-40) with added cOmplete protease inhibitor tablet (Roche) per 50 mL buffer. SDS samples were prepared by adding 2x SDS sample buffer (20% glycerol, 120 mM Tris-HCl pH 6.8, 4% SDS, 0.02% bromophenol blue, 4% β-mercaptoethanol) to one volume of lysate and boiling at 95°C for 5 min. Samples were run on Novex 4–12% Tris-Glycine gels (Invitrogen) and gels were transferred to PVDF membranes by semi-dry transfer. Antibodies used to determine protein expression were: PEPCK-C H-300 (Santa Cruz Biotechnology, Cat. No.: sc-32879), PCB H-2 (Santa Cruz Biotechnology, Cat. No.: sc-271493), Phospho-CREB Ser133 87G3 (Cell Signaling, Cat. No.: 9198), CREB 48H2 (Cell Signaling, Cat. No.: 9197), Phospho-AMPKα Thr172 40H9 (Cell Signaling, Cat. No.: 2535), AMPKα (Cell Signaling, Cat. No.: 2532), GAPDH D16H11 XP^® (Cell Signaling, Cat. No.: 5174), G6Pase (Santa Cruz Biotechnology, Cat. No.: sc-27198), Phospho-ACC (Cell Signaling, Cat. No.: 3661), and ACC (Cell Signaling, Cat. No.: 3676)

Madiraju A.K., Qiu Y., Perry R.J., Rahimi Y., Zhang X.M., Zhang D., Camporez J.P., Cline G.W., Butrico G.M., Kemp B.E., Casals G., Steinberg G.R., Vatner D.F., Petersen K.F, & Shulman G.I. (2018). Metformin Inhibits Gluconeogenesis by a Redox-Dependent Mechanism In Vivo. Nature medicine, 24(9), 1384-1394.

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Immunoblotting: Protein Detection and Quantification

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Immunoblots were performed using standard protocols. Cells were lysed directly in Laemmli loading buffer, proteins were resolved by SDS-PAGE and transferred to PVDF membranes (EMD Millipore, Billerica, MA). Membranes were blocked in Tris buffered saline (TBS) containing 0.1% Tween and 5% non-fat powdered milk (TBS-T, 5% milk). Primary antibody incubation was performed in TBS-T, 5% milk overnight at 4°C. Proteins were visualized using a species-specific HRP-conjugated secondary antibody and the ECL Plus chemifluorescent detection system (Thermo Fisher Scientific Inc., Waltham, MA) on a STORM scanner ((GE Healthcare, Piscataway, NJ). Signal was quantified using ImageJ software (http://rsb.info.nih.gov/ij/). The following primary antibodies were used: ACC (#3676, Cell Signaling), phospho-ACC (Ser79, #3661, Cell Signaling), AMPKα (#2603, Cell Signaling), phospho-AMPKα (Thr172, #2535, Cell Signaling), DDIT3/CHOP (#2895, Cell Signaling), HER2 (#4290, Cell Signaling), HER3 (#4754, Cell Signaling), α-tubulin (MCA78G, AbD Serotec, Oxford, UK). Secondary antibodies: goat-anti-rabbit-HRP (#7074, Cell Signaling), horse-anti-mouse-HRP (#7076, Cell Signaling), goat-anti-rat-HRP (sc-2303), Santa Cruz Biotechnology, Inc., Santa Cruz, CA).

Baumann J., Kokabee M., Wong J., Balasubramaniyam R., Sun Y, & Conklin D.S. (2018). Global metabolite profiling analysis of lipotoxicity in HER2/neu-positive breast cancer cells. Oncotarget, 9(43), 27133-27150.

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PDAC Cell Protein Quantification

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For the assessment of STAT3, AMPK, or ACC level and phosphorylation status, total cell protein extracts were obtained from human and mouse PDAC cell lines, as well as from mouse tissue samples, via lysis and sonication in RIPA buffer (Cell Signaling, 9806). Total cell lysates were resolved by a 7.5% SDS-PAGE and probed with phospho-STAT3 (Cell Signaling, 9145S), phospho-AMPK (Cell Signaling, 2535S), phospho-ACC (Cell Signaling, 11818T), total STAT3 (Cell Signaling, 9139S) total AMPK (Cell Signaling, 5832S), and total ACC (Cell Signaling, 3676T). To determine equal loading, control membranes were probed with β-actin (Abgent, AM1829B). Quantitation of protein relative amounts of triplicate experiments were analyzed by ImageJ software as a ratio of each phospho protein band relative to the correspondent lane total protein band, followed by ratio to the lane’s loading control.

Messaggio F., Mendonsa A.M., Castellanos J., Nagathihalli N.S., Gorden L., Merchant N.B, & VanSaun M.N. (2017). Adiponectin receptor agonists inhibit leptin induced pSTAT3 and in vivo pancreatic tumor growth. Oncotarget, 8(49), 85378-85391.

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Protein Expression Analysis in Gastrocnemius Muscle

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Gastrocnemius muscle samples were lysed in a buffer containing 20mM Tris-Hcl, 150mM NaCl, 1mM EDTA, 1mM EGTA, 0.5% NP-40, 2.5mM sodium pyrophosphate, 1mM sodium orthovanadate, and protease inhibitor cocktail. Protein lysates were isolated by centrifuging samples at 12,000 rpm at 4°C for 15 minutes. Protein concentration was determined and 20 μg of protein was loaded per well. The protein bands were transferred to a PVDF membrane and immunoblotted using the following antibodies: SOD1 (Santa Cruz Biotechnology), 5′ Adenosine monophosphate-activated protein kinase alpha (AMPKα), phospho AMPKα, Acetyl-CoA carboxylase (ACC), and phospho ACC (Cell Signaling Technology). The bands were visualized using infrared (IR 700 or 800)-conjugated secondary antibodies using the Odyssey Imaging System (LI-COR Biosciences).

Natarajan G., Perriotte-Olson C., Bhinderwala F., Powers R., Desouza C.V., Talmon G.A., Yuhang J., Zimmerman M.C., Kabanov A.V, & Saraswathi V. (2017). Nanoformulated Copper/Zinc Superoxide Dismutase Exerts Differential Effects on Glucose versus Lipid Homeostasis Depending on the Diet Composition Possibly via Altered AMPK Signaling. Translational research : the journal of laboratory and clinical medicine, 188, 10-26.

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Western Blot Analysis of Protein Expression

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Total protein from
HAECs was obtained using radioimmunoprecipitation assay (RIPA) buffer
(Beyotime, China). Total proteins (20 μg) were electrically
separated on a sodium dodecyl sulfate polyacrylamide gel and then
transferred to a polyvinylidene difluoride (PVDF) membrane. The membrane
was then blocked with 5% slim milk for 1 h. The PVDF membranes were
incubated with a specific primary antibody diluted in block buffer
at 4 °C overnight. The following primary antibodies were used:
NOX4 (1:2000, Cat#GTX12206, Gene Tax), TxNIP (1:2000, Cat#ab188865,
Abcam), NLRP3 (1:2000, Cat#19771-1-AP, Protein Tech), p10 (1:2000,
Cat#sc-514, Santa Cruz Biotechnology), phospho-AMPKα (1:2000,
Cat #50081, Cell Signaling), AMPKα (1: 2000, Cat#2532, Cell
Signaling), phospho-ACC (1:2000 Cat# 3661, Cell Signaling), ACC (1:2000,
Cat#3662, Cell Signaling), and β-actin (1:5000, Cat#sc-130656,
Santa Cruz Biotechnology). After three washes, the membrane was incubated
with horseradish peroxidase (HRP)-conjugated secondary antibodies.
The protein signals were detected using an enhanced chemiluminescence
reagent, and the protein band was analyzed using Quantity One software
(Bio-Rad, USA).

Li W., Zhang D., Yuan W., Wang C., Huang Q, & Luo J. (2020). Humanin Ameliorates Free Fatty Acid-Induced Endothelial Inflammation by Suppressing the NLRP3 Inflammasome. ACS Omega, 5(35), 22039-22045.

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