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Hif 1β

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HIF-1β is a laboratory product offered by Cell Signaling Technology. It is a subunit of the hypoxia-inducible factor 1 (HIF-1) transcription factor complex, which plays a crucial role in cellular responses to low oxygen conditions.

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Lab products found in correlation

Hif 1α, by BD (4 mentions) β actin, by Cell Signaling Technology (3 mentions) Hif 1α, by Cell Signaling Technology (3 mentions) β actin, by Merck Group (2 mentions) Stat3, by Cell Signaling Technology (2 mentions) Bcl2 associated x (bax), by Cell Signaling Technology (2 mentions) Hrp conjugated secondary antibody, by Agilent Technologies (1 mentions) Ecl detection system, by Cytiva (1 mentions) Horseradish peroxidase conjugated secondary antibody, by Cell Signaling Technology (1 mentions) Abe70, by Merck Group (1 mentions) Hif 2α, by Santa Cruz Biotechnology (1 mentions) Poly adp ribose polymerase parp, by Cell Signaling Technology (1 mentions) Caspase 3, by Cell Signaling Technology (1 mentions) Sin3a, by Santa Cruz Biotechnology (1 mentions) Hif 2α, by Thermo Fisher Scientific (1 mentions) Hdac1, by Merck Group (1 mentions) Cleaved parp, by Cell Signaling Technology (1 mentions) Cleaved caspase 3, by Cell Signaling Technology (1 mentions) Ab191562, by Abcam (1 mentions) Y705 pstat3, by Cell Signaling Technology (1 mentions)

Spelling variants of this product

Anti-HIF-1β (5 citations) Rabbit monoclonal anti-HIF-1β (2 citations) Anti-HIF-1β/ARNT (2 citations) HIF-1β/ARNT antibody (2 citations)

12 protocols using hif 1β

1

Western Blot Analysis of Cellular Proteins

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Whole cell extracts were prepared in RIPA lysis buffer and processed for Western blotting as published previously [16 (link)]. Proteins were separated on 10% reducing SDS gels and blotted onto PVDF membranes. Unspecific binding sites were blocked with 5% skimmed milk in TBST. Rabbit polyclonal antibodies against MTH1 (Novus Biologicals, Littleton, CO, USA) and HIF-1β (Cell Signaling, Cambridge, UK) as well as monoclonal mouse antibodies against β-actin (Sigma), were used overnight at 4 °C. HRP-conjugated secondary antibodies (Dako, Hamburg, Germany) were used for 1 h at room temperature.
Pompsch M., Vogel J., Classen F., Kranz P., Iliakis G., Riffkin H., Brockmeier U, & Metzen E. (2018). The presumed MTH1-inhibitor TH588 sensitizes colorectal carcinoma cells to ionizing radiation in hypoxia. BMC Cancer, 18, 1190.
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2

Protein Expression Analysis by Western Blot

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Equal amount of control and SAHA-treated cell lysates were mixed with Laemmli's buffer [20 (link)], boiled, and separated on SDS–polyacrylamide gels. Proteins were transferred onto PVDF membranes and probed with polyclonal PKCε, HIF1β, CDK6, and RhoC antibodies (Cell Signaling, MA, USA) followed by anti-rabbit/mouse HRP-conjugated secondary antibodies. Tubulin was used as a loading control. Membranes were visualized using ECL detection system (Amersham Life Science, NJ, USA).
Datta J., Islam M., Dutta S., Roy S., Pan Q, & Teknos T.N. (2016). Suberoylanilide hydroxamic acid inhibits growth of head and neck cancer cell lines by reactivation of tumor suppressor microRNAs. Oral oncology, 56, 32-39.
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3

Lysate Preparation and Immunoblotting

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The cells were lysed in the RIPA buffer (50 mM Tris-HCl (pH 8), 150 mM NaCl, 1% (v/v) NP40, 0.25% (w/v) Na deoxycholate, 0.1% (w/v) SDS, 10 mM NaF, 2 mM Na3VO4 and 1 tablet/10 mL cOmplete, Mini, EDTA-free protease inhibitors (Roche)), centrifuged for 10 mins at 13,000 rpm at 4 °C, collecting the supernatant for analysis. SDS-PAGE and immunoblotting were performed using standard protocols. The following primary antibodies were used for immunoblotting: HIF-1α (610958, BD Biosciences, Workingham, UK), HIF-2α (sc-13596, Santa Cruz, Dallas, TX, USA), HIF-1β (3718, Cell Signalling, Leiden, Holland), β-Actin (3700, Cell Signaling, Leiden, Holland/60009-1-Ig, Proteintech, Manchester, UK), PBRM1 (ABE70, Millipore, Feltham, UK), YTHDF2 (24744-1-AP, Proteintech, Manchester, UK/80014, Cell Signaling, Leiden, Holland), GFP (2956, Cell Signaling), BRG1 (Santa Cruz, Dallas, TX, USA, sc-17796). Following incubation with a horseradish peroxidase-conjugated secondary antibody (Cell Signalling Technology, Leiden, Holland), chemiluminescence (Pierce/ThermoFisher, Paisley, UK) was detected. All the figures are representative of a minimum of three independent experiments.
Shmakova A., Frost M., Batie M., Kenneth N.S, & Rocha S. (2021). PBRM1 Cooperates with YTHDF2 to Control HIF-1α Protein Translation. Cells, 10(6), 1425.
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4

Comprehensive Protein Extraction and Analysis

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Cells were lysed in RIPA buffer, 50 mM Tris–HCl (pH 8), 150 mM NaCl, 1% (v/v) NP40, 0.5% (v/v) Na-deoxycholate, 0.1% (v/v) SDS, and 1 tablet/10 ml Complete, Mini, EDTA-free protease inhibitors (Roche). SDS–PAGE and immunoblots were carried out using standard protocols.
Antibodies were used as follows: SINHCAF [1 (link)], HIF-2α (PA1-16510, Thermo Scientific; 7096, Cell Signaling), β-actin (3700, Cell Signaling), HIF-1α (610958, BD Biosciences), HIF-1β (3718, Cell Signaling), PHD2 (Bethyl A300-322A; 4835, Cell Signaling); p52 (05-361, Merck/Millipore), E2F1 (3742, Cell Signaling), SP1 (07-645, Merck/Millipore), SP3 (sc-644, Santa Cruz Biotech), Sin3A (sc-994, Santa Cruz Biotech; 8056, Cell Signaling); HDAC1 (17-10199; Merck/Millipore), AcH3 (acetylated histone H3; 06-599, Merck/Millipore), p62/STQM (610832, BD Biosciences), poly-ADP ribose polymerase (PARP) (9532, Cell Signaling), cleaved PARP (9541, Cell Signaling), caspase-3 (9662, Cell Signaling), cleaved caspase-3 (9661, Cell Signaling), and LC3B (3868, Cell Signaling).
Biddlestone J., Batie M., Bandarra D., Munoz I, & Rocha S. (2018). SINHCAF/FAM60A and SIN3A specifically repress HIF-2α expression. Biochemical Journal, 475(12), 2073-2090.
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5

Elucidating PRMT3 and HIF-1 Signaling

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RKO, LoVo, HEK293T, and HUVEC cell lines were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). Antibodies against PRMT3 (Abcam; ab191562), GAPDH (Abcam; ab9485), H4R3me2a (Abcam; ab194683), histone 4 (Abclone; A1131), VEGFA (Abclone; A12303), HIF1α (cell signaling technology; #36169); HIF1β (cell signaling technology; #3718), flag-tag (cell signaling technology; #14793), stat3 (cell signaling technology; #9139), Y705-p-stat3 (cell signaling technology; #9145), β-actin (cell signaling technology; #3700) were used in the current study. SGC707 (HY-19715) was obtained from MedChemExpress company. All cell lines were authenticated by STR profiling and there was no mycoplasma contamination.
Zhang X., Wang K., Feng X., Wang J., Chu Y., Jia C., He Q, & Chen C. (2021). PRMT3 promotes tumorigenesis by methylating and stabilizing HIF1α in colorectal cancer. Cell Death & Disease, 12(11), 1066.
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6

Fibroblast Protein Expression Analysis

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Fibroblasts were lysed using 2 x Laemmli SDS sample buffer or urea buffer (8 M Urea, 1 M Thiourea, 0.5% CHAPS, 50 mM DTT, and 24 mM Spermine). Western blotting of cellular lysates was performed for β-actin (1:100.000, Sigma-Aldrich, Poole, UK), LOXL2 (1:1000, R&D Systems, Abingdon, UK), HIF1α (1:1000, BD Biosciences, Wokingham, UK), FIH (1:200, mouse monoclonal 162 C) (Wang et al., 2018 (link)), β-tubulin (1:1000, Cell Signaling Technology, London, UK), HIF1 β (1:1000, Cell Signaling Technology), p-Smad2/3 (1:1000, Cell Signaling Technology), p-ERK (1:1000, Cell Signaling Technology), active β-catenin (1:1000, Cell Signaling Technology). Immunodetected proteins were identified using the enhanced chemiluminescence system (Clarity Western Blotting ECL Substrate, Bio-Rad Laboratories Ltd, Watford, UK) or Odyssey imaging system (LI-COR), and evaluated by ImageJ 1.42q software (National Institutes of Health).
Brereton C.J., Yao L., Davies E.R., Zhou Y., Vukmirovic M., Bell J.A., Wang S., Ridley R.A., Dean L.S., Andriotis O.G., Conforti F., Brewitz L., Mohammed S., Wallis T., Tavassoli A., Ewing R.M., Alzetani A., Marshall B.G., Fletcher S.V., Thurner P.J., Fabre A., Kaminski N., Richeldi L., Bhaskar A., Schofield C.J., Loxham M., Davies D.E., Wang Y, & Jones M.G. (2022). Pseudohypoxic HIF pathway activation dysregulates collagen structure-function in human lung fibrosis. eLife, 11, e69348.
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7

Comparative Analysis of Pancreatic and Cholangiocarcinoma Cell Lines

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The human pancreatic cancer cell lines PANC-1 and CFPAC-1 were cultured in Dulbecco's modified Eagle's medium and the cholangiocarcinoma cell lines HuCCT-1 and SCK were cultured in RPMI-1640. PANC-1 and CFPAC-1 cells were purchased from the ATCC (Manassas, VA, USA). HuCCT-1 and SCK cells were procured from the Health Science Research Resources Bank (Osaka, Japan) and Dr. Dae-Ghon Kim of Chonbuk National University Medical School and Hospital (Jeonju, Korea), respectively. All cell lines were maintained in a humidified incubator at 37°C with 5% CO2. Antibodies against S6K, phospho-S6K, S6, phospho-S6, 4EBP1, phospho-4EBP1, cleaved caspase-3, CHOP, Bax, Bim, BCl-2, cyclin B1, HIF-1β, CD44, SPARC, vimentin, and GAPDH were obtained from Cell Signaling Technology. CD-31 and VEGF were purchased from Abcam (Cambridge, MA, USA). HIF-1α, VEFGR2/Flk-1, and MMP-2 were obtained from Santa Cruz Biotechnology.
Bang S., Jang S.I., Lee S.Y., Baek Y.Y., Yun J., Oh S.J., Lee C.W., Jo E.A., Na K., Yang S., Lee D.H, & Lee D.K. (2015). Molecular Mechanism of Local Drug Delivery with Paclitaxel-Eluting Membranes in Biliary and Pancreatic Cancer: New Application for an Old Drug. Gastroenterology Research and Practice, 2015, 568981.
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8

Hypoxia Signaling Pathway Regulation

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Brusatol was purchased from Tauto Biotech (Shanghai, China); cycloheximide (CHX) and MG132 were purchased from Calbiochem; 2,2'-bipyridiyl was purchased from Sigma-Aldrich; and dimethyloxalylglycine (DMOG) was purchased from Santa Cruz Biotechnology. Primary antibodies against the following proteins were used: HIF-1α (R&D Systems), HIF-1β (Cell Signaling Technology), PHD1 (R&D Systems), PHD2 (Cell Signaling Technology), PHD3 (Novus Biologicals), and c-Myc (Cell Signaling Technology). Secondary antibodies used for immunoblotting include horseradish peroxidase (HRP)-conjugated anti-mouse (Cell Signaling Technology), anti-rabbit (Cell Signaling Technology), and anti-goat (Upstate).
Oh E.T., Kim C.W., Kim H.G., Lee J.S, & Park H.J. (2017). Brusatol-Mediated Inhibition of c-Myc Increases HIF-1α Degradation and Causes Cell Death in Colorectal Cancer under Hypoxia. Theranostics, 7(14), 3415-3431.
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9

Comprehensive Protein Analysis from Cell Lysates

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A total of 1 × 106 cells were lysed for each sample, as described previously.25 (link), 62 (link) Equal amounts of protein were separated on 7.5–13% SDS-PAGE depending on the molecular size of the proteins of interest, blotted onto a nitrocellulose membrane (Amersham Biociences, Little Chalfon, UK) and blocked with 5% non-fat dry milk in PBS/Tween (0.05% Tween-20 in PBS). Antibodies directed against the following proteins were used: anti-c-Flip was made by our own laboratory;63 (link) anti-Bad, -Bak, -Bax, -Bcl-xL, -Bcl-w, -Bid, -Bim, -HIF-1α, -HIF-1β, -Puma, -VEGF and -XIAP were purchased from Cell Signaling Technology (Danvers, MA, USA); anti-Bcl-2, -Mcl-1, -p21 and -p27 were purchased from Santa Cruz Biotechnology (Heidelberg, Germany); anti-Tax from NIH (Hybridoma, Bethesda, MD, USA; cat. no. 1312); anti-tubulin and -actin were purchased from Sigma-Aldrich (Munich, Germany).
Mühleisen A., Giaisi M., Köhler R., Krammer P.H, & Li-Weber M. (2014). Tax contributes apoptosis resistance to HTLV-1-infected T cells via suppression of Bid and Bim expression. Cell Death & Disease, 5(12), e1575-.
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10

Investigating Cell Responses to Hypoxia and Chemotherapies

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Cells were exposed to normoxia or hypoxia, and/or subjected to treatment with cisplatin or docetaxel for 24h. Cells were lysed for 20 min in ice-cold RIPA lysis buffer supplemented with 1 mM PMSF and a cocktail of protease inhibitors. Blotting was performed with antibodies against UCP2 (Clone # sc-6526, Santa Cruz, Dallas, USA), PPARγ (Cat. # 2443, Cell Signaling Technology, Boston, USA), DEC1 (Santa Cruz), HIF1α (Cat. # 14179, Cell Signaling), HIF-1β (Cat. # 3414, Cell Signaling), Stat3 (Cat. # 9139, Cell Signaling), phospho-Stat3 (Tyr705, Cat. # 9145, Cell Signaling), and ABCG2 (Cat. # 4477, Cell Signaling). Goat anti-rabbit and goat anti-mouse immunoglobulin horseradish peroxidase-linked F(ab)2 fragments from Millipore (Billerica, MA, USA) were used as secondary antibodies.
Wang M., Li G., Yang Z., Wang L., Zhang L., Wang T., Zhang Y., Zhang S., Han Y, & Jia L. (2016). Uncoupling protein 2 downregulation by hypoxia through repression of peroxisome proliferator-activated receptor γ promotes chemoresistance of non-small cell lung cancer. Oncotarget, 8(5), 8083-8094.
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