FIH-1/HIF-1AN Antibody

Western Blot: FIH-1/HIF-1AN Antibody [NB100-428] - Analysis of astrocytes lysate from Rat brain using FIH-1/HIF-1AN antibody at 1:500 dilution with ECL detection. The antibody detected a major band at ~40 kDa position representing FIH-1/HIF-1AN protein.

Western Blot: FIH-1/HIF-1AN Antibody [NB100-428] - Representative picture of western blot in histopathologically unchanged tissue (N) and primary cancerous tissue (C) from patients with CRC. Immunodetection of bands was performed with Rp anti- PHD1 (Catalog # NB100-310), - PHD2 (Catalog # NB100-137), - PHD3 (Catalog # NB100-139) and -FIH-1/HIF-1AN Ab (Catalog # NB100-139), followed by incubation with goat anti-rabbit HRP-conjugated Ab. The membrane was stripped and incubated with Rp anti-GAPDH Ab, followed by incubation with goat anti-rabbit HRP-conjugated Ab. Image collected and cropped by CiteAb from the following publication (//bmccancer.biomedcentral.com/articles/10.1186/1471-2407-13-526), licensed under a CC-BY licence.

Immunohistochemistry: FIH-1/HIF-1AN Antibody [NB100-428] - Detection of FIH-1 in rat brain tissue.

Western Blot: FIH-1/HIF-1AN Antibody [NB100-428] - Rat VSMC (vascular smoot muscle) cells treated (siFIH) or not (CTL) with a specific siRNA against FIH-1. Western blot image submitted by a verified customer review.

Knockdown Validated: FIH-1/HIF-1AN Antibody [NB100-428] - (C) Cytoplasmic FIH-1 (lower band) and MMP14 expression levels in both aerobic and hypoxic conditions in the 3 cell lines. (E) siRNA targeting FIH-1 was used to assess whether FIH-1 had any effect on CAIX expression in MCF-7 cells cultured in 0.5% O2 conditions. cont, control. m, transfection reagent alone. scr, scrambled siRNA. pool, mixture of 4 siRNAs targeting FIH-1. 21, single siRNA targeting FIH-1. (F) FIH-1 and CAIX expression levels in aerobic MCF-7 cells treated with either siRNA against FIH-1 alone, or both 400 uM CoCl2 and siRNA together for 24, 48 and 72h. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/28476026/) licensed under a CC-BY licence.

• Reactivity: Hu, Mu, Rt
• Applications: WB, EM, ICC/IF, IHC, IHC-Fr, IHC-P, IP, KD, KO
• Clonality: Polyclonal
• Host: Rabbit
• Conjugate: Unconjugated
• Concentration: 1.0 mg/ml

FIH-1/HIF-1AN Antibody Summary

• Immunogen: A synthetic peptide made to the C-terminal region of human Factor Inhibiting HIF-1 (between residues 300 and the C-terminus). [UniProt# Q9NWT6]
• Localization: Nuclear
• Isotype: IgG
• Clonality: Polyclonal
• Host: Rabbit
• Gene: HIF1AN
• Purity: Immunogen affinity purified

Applications/Dilutions

• Dilutions:
  • Electron Microscopy
  • Immunocytochemistry/Immunofluorescence
  • Immunohistochemistry 1:100
  • Immunohistochemistry-Frozen 1:50-1:200
  • Immunohistochemistry-Paraffin 1:50-1:200
  • Immunoprecipitation 1:10-1:500
  • Knockdown Validated
  • Knockout Validated
  • Western Blot 1:1000-1:5000
• Application Notes: Knockout data from PMID 31937588.
• Theoretical MW: 40.3 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• Control:

  FIH-1/HIF-1AN Overexpression Lysate
  HIF-1 alpha Knockout CoCl2-treated/untreated HeLa Cell Lysate
  HIF-1 alpha Knockout Hypoxic-treated/untreated HeLa Cell Lysate
  NIH 3T3 CoCl2 treated/Untreated Cell Lysate
  L929 CoCl2 treated/Untreated Cell Lysate
  Raw 264.7 CoCl2 treated/Untreated Cell Lysate
  PC12 CoCl2 treated/Untreated Cell Lysate
  PC12 Hypoxic / Normoxic Cell Lysate
  Raw 264.7 Hypoxic / Normoxic Cell Lysate
  L929 Hypoxic / Normoxic Cell Lysate
  NIH 3T3 Hypoxic / Normoxic Cell Lysate

• Control Peptide:

  FIH-1/HIF-1AN Peptide (NB100-428PEP)

Reactivity Notes

Human, rat and mouse (PMID 19720742).

Packaging, Storage & Formulations

• Storage: Store at 4C. Do not freeze.
• Buffer: Tris-Citrate/Phosphate (pH 7.0 - 8.0)
• Preservative: 0.1% Sodium Azide
• Concentration: 1.0 mg/ml
• Purity: Immunogen affinity purified

Alternate Names for FIH-1/HIF-1AN Antibody

• DKFZp762F1811
• EC 1.14.11.16
• factor inhibiting HIF1
• Factor inhibiting HIF-1
• FIH1
• FIH-1
• FIH1FIH-1
• FLJ20615
• FLJ22027
• HIF1AN
• HIF-1AN
• hypoxia inducible factor 1, alpha subunit inhibitor
• hypoxia-inducible factor 1-alpha inhibitor
• Hypoxia-inducible factor asparagine hydroxylase
• Peptide-aspartate beta-dioxygenase

Background

Factor inhibiting HIF1 (FIH1) is an asparaginyl hydroxylase enzyme that regulates HIF's transcriptional activity by hydroxylating HIF-1 alpha at 'Asp-803' in the CAD (C-terminal transactivation domain). Hydroxylation via PHDs followed by proteosome degradation is another mechanism that regulates HIF1, however, FIH1 is able to exercise the control even under severe hypoxic conditions, when PHD enzymes fail to do so. Localized mainly in cytoplasm, FIH1 functions as oxygen sensor and FIH1 interaction with NOTCH1 results in its nuclear localization whereas FIH1-ABPA3 interaction leads to its perinuclear localization. During normoxic conditions, FIH1 hydroxylates a conserved asparaginyl residue within CTAD of HIF-1 alpha/HIF-2 alpha, leading to steric clash that prevents the recruitment of the co-activators p300 and CBP. On the other hand, under hypoxic conditions, FIH1 is inactive, which results in activation of HIF-alpha signaling. FIH1 is involved in transcriptional repression through interaction with HIF1A, VHL and HDAC and hydroxylates specific Asn residues within ARD (ankyrin repeat domains) of NFKB1, NFKBIA, NOTCH1, ASB4, PPP1R12A as well as other ARD-containing proteins. FIH1 also hydroxylates Asp and His residues within ARDs of ANK1 and TNKS2, respectively. FIH1 has the ability to negatively regulate NOTCH1 activity thus accelerating myogenic differentiation, and positively regulates ASB4 activity which promotes vascular differentiation. FIH1-Mint3 binding affects FIH1's ability to modify HIF-1 alpha in an oxygen-independent manner and FIH1-Bax interaction plays inhibitory role in apoptosis.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for FIH-1/HIF-1AN Antibody (NB100-428)(26)

Publications using NB100-428

• E G, Sun B, Liu B et al. Enhanced BPGM/2,3-DPG pathway activity suppresses glycolysis in hypoxic astrocytes via FIH-1 and TET2 Brain research bulletin 2022-11-08 [PMID: 36334804] (WB, Human)
  
  Details:
  Dilution used in WB 1:1000
• Li ZL, Wang B, Lv LL et al. FIH-1-modulated HIF-1 alpha C-TAD promotes acute kidney injury to chronic kidney disease progression via regulating KLF5 signaling Acta pharmacologica Sinica 2021-03-03 [PMID: 33658705]
• Chung Y, Saitoh Y, Hayashi T et al. Mint3 is dispensable for pancreatic and kidney functions in mice Biochemistry and biophysics reports 2020-12-01 [PMID: 33319072] (WB)
• Mader J, Huber J, Bonn F et al. Oxygen-dependent asparagine hydroxylation of the ubiquitin-associated (UBA) domain in Cezanne regulates ubiquitin binding J. Biol. Chem. 2020-01-14 [PMID: 31937588] (WB, KO, Human)
• Docherty C K, Nilsen M et al. Influence of 2-Methoxyestradiol and Sex on Hypoxia-Induced Pulmonary Hypertension and Hypoxia-Inducible Factor-1-alpha. J Am Heart Assoc 2019-05-03 [PMID: 30819028] (WB, Human)
• Pillai MR, Mihi B, Ishiwata K et al. Myc-induced nuclear antigen constrains a latent intestinal epithelial cell-intrinsic anthelmintic pathway PLoS ONE 2019-02-26 [PMID: 30807587] (WB, Mouse)
• Bullen JW, Tchernyshyov I, Holewinski RJ et al. Protein kinase A-dependent phosphorylation stimulates the transcriptional activity of hypoxia-inducible factor 1 Sci Signal 2016-05-31 [PMID: 27245613] (WB, Human)
• Nakaoka HJ, Hara T, Yoshino S et al. NECAB3 Promotes Activation of Hypoxia-inducible factor-1 during Normoxia and Enhances Tumourigenicity of Cancer Cells Sci Rep 2016-03-07 [PMID: 26948053] (IP, Human)
• Kiriakidis S, Henze AT, Kruszynska-Ziaja I et al. Factor-inhibiting HIF-1 (FIH-1) is required for human vascular endothelial cell survival FASEB J. 2015-04-02 [PMID: 25837583] (WB, Human)
• Karttunen S, Duffield M, Scrimgeour Nr et al. Oxygen-dependent hydroxylation by Factor Inhibiting HIF (FIH) regulates the TRPV3 ion channel. J. Cell. Sci. 2014-11-20 [PMID: 25413349] (WB)
• Rawluszko AA, Bujnicka KE, Horbacka K et al. Expression and DNA methylation levels of prolyl hydroxylases PHD1, PHD2, PHD3 and asparaginyl hydroxylase FIH in colorectal cancer. BMC Cancer. 2013-11-06 [PMID: 24195777] (WB, Human)
• Janke K, Brockmeier U, Kuhlmann K et al. Factor Inhibiting HIF-1 (FIH-1) modulates protein interactions of Apoptosis-Stimulating p53 binding Protein 2 (ASPP2). J Cell Sci 2013-04-19 [PMID: 23606740] (WB, Human)
• Nishida C, Kusubata K, Tashiro Y et al. MT1-MMP plays a critical role in hematopoiesis by regulating HIF-mediated chemokine/cytokine gene transcription within niche cells Blood 2012-06-07 [PMID: 22544701]
• Lim CS, Kiriakidis S, Paleolog EM, Davies AH. Increased activation of the hypoxia-inducible factor pathway in varicose veins. J Vasc Surg;55(5):1427-1439e1. 2012-05-01 [PMID: 22277691] (WB, Human)
• Wilkins SE, Karttunen S, Hampton-Smith RJ et al. Factor inhibiting HIF (FIH) recognises distinct molecular features within hypoxia inducible factor (HIF)-[alpha] versus ankyrin repeat substrates. The Journal of Biological Chemistry. 2012-01-23 [PMID: 22270367]
• Melvin A, Mudie S, Rocha S. The chromatin remodeler ISWI regulates the cellular response to hypoxia: role of FIH. Mol Biol Cell 2011-11-01 [PMID: 21900490] (Human)
• Singleton RS, Trudgian DC, Fischer R et al. Quantitative mass spectrometry reveals the dynamics of Factor Inhibiting HIF (FIH)-catalysed hydroxylation. J Biol Chem. 2011-07-30 [PMID: 21808058]
• Schodel J, Bohr D, Klanke B et al. Factor inhibiting HIF limits the expression of hypoxia-inducible genes in podocytes and distal tubular cells. Kidney Int. 2010-01-01 [PMID: 20720525]
• Luo T, Zhang X, Nicholson WL et al. Molecular characterization of antibody epitopes of Ehrlichia chaffeensis ankyrin protein 200 and tandem repeat protein 47 and evaluation of synthetic immunodeterminants for serodiagnosis of human monocytotropic ehrlichiosis. Clin Vaccine Immunol 2010-01-01 [PMID: 19955322]
• Minamishima YA, Moslehi J, Padera RF et al. A feedback loop involving the Phd3 prolyl hydroxylase tunes the mammalian hypoxic response in vivo. Mol Cell Biol 2009-11-01 [PMID: 19720742] (WB, Mouse)
• Chan DA, Kawahara TL, Sutphin PD et al. Tumor vasculature is regulated by PHD2-mediated angiogenesis and bone marrow-derived cell recruitment. Cancer Cell 2009-06-02 [PMID: 19477431]
• Xu X, Vaithiyalingam S, Glick GG et al. The basic cleft of RPA70N binds multiple checkpoint proteins, including RAD9, to regulate ATR signaling. Mol Cell Biol 2008-12-01 [PMID: 18936170]
• Zheng X, Linke S, Dias JM et al. Interaction with factor inhibiting HIF-1 defines an additional mode of cross-coupling between the Notch and hypoxia signaling pathways. Proc Natl Acad Sci U S A;105(9):3368-73. 2008-03-04 [PMID: 18299578] (WB)
• Li, J et al. Protein kinase C-mediated modulation of FIH-1 expression by the homeodomain protein CDP/Cut/Cux. Mol Cell Biol;27(20):7345-53. 2007-10-01 [PMID: 17682059]
• Ferguson III, JE, Wu, Y et al. ASB4 is a hydroxylation substrate of FIH and promotes vascular differentiation via an oxygen-dependent mechanism. Mol Cell Biol;27(18):6407-19. 2007-09-01 [PMID: 17636018]
• Khan Z, Michalopoulos GK, Stolz DB. Peroxisomal localization of hypoxia-inducible factors and hypoxia-inducible factor regulatory hydroxylases in primary rat hepatocytes exposed to hypoxia-reoxygenation. Am J Pathol 2006-10-01 [PMID: 17003483] (ICC/IF, EM, WB, IHC-P, Rat)

Reviews for FIH-1/HIF-1AN Antibody (NB100-428) (3)

Average Rating: 4.3

(Based on 3 reviews)

We have 3 reviews tested in 2 species: Human, Rat.

reviewed by: Verified Customer WB Rat 06/05/2020

Summary

• Application: Western Blot
• Sample Tested: Vascular Smooth Muscle Cells,Whole cell lysate
• Species: Rat
• Lot: A3

reviewed by: Prabir Chakraborty WB Human 11/12/2017

Summary

• Application: Western Blot
• Sample Tested: Cell lysates from prostate cancer cell lines
• Species: Human
• Lot: A2

Comments

• Comments: Sample: Whole cell lysates of prostate cancer cell lines (20ug)
  Blocking: 5% nonfat dry milk reconstituted in TBS-T for 1 hour at room temperature
  Primary Antibody diluted (1:500) in 5% nonfat dry milk reconstituted in TBS-T overnight at 4deg C
  Secondary Antibody: Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution in TBS-T

reviewed by: cynthia morimoto WB Human 12/12/2014

Summary

• Application: Western Blot
• Sample Tested: See PMID 22277691
• Species: Human

Bioinformatics

• Gene Symbol: HIF1AN
• Entrez:
  • 55662(Human)
  • 29560(Rat)
• Uniprot:
  • Q9NWT6(Human)