Product: Phospho-Histone H2A.X (Tyr143) Antibody
Catalog: AF8482
Description: Rabbit polyclonal antibody to Phospho-Histone H2A.X (Tyr143)
Application: WB IF/ICC
Reactivity: Human, Mouse, Rat
Prediction: Bovine, Sheep, Dog
Mol.Wt.: 15kDa; 15kD(Calculated).
Uniprot: P16104
RRID: AB_2840536

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 100ul $350 In stock
 200ul $450 In stock

Lead Time: Same day delivery

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Product Info

Source:
Rabbit
Application:
WB 1:1000-3000, IF/ICC 1:200
*The optimal dilutions should be determined by the end user.
*Tips:

WB: For western blot detection of denatured protein samples. IHC: For immunohistochemical detection of paraffin sections (IHC-p) or frozen sections (IHC-f) of tissue samples. IF/ICC: For immunofluorescence detection of cell samples. ELISA(peptide): For ELISA detection of antigenic peptide.

Reactivity:
Human,Mouse,Rat
Prediction:
Bovine(100%), Sheep(100%), Dog(100%)
Clonality:
Polyclonal
Specificity:
Phospho-Histone H2A.X (Tyr143) Antibody detects endogenous levels of Histone H2A.X only when phosphorylated at Tyr143.
RRID:
AB_2840536
Cite Format: Affinity Biosciences Cat# AF8482, RRID:AB_2840536.
Conjugate:
Unconjugated.
Purification:
The antibody is from purified rabbit serum by affinity purification via sequential chromatography on phospho-peptide and non-phospho-peptide affinity columns.
Storage:
Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol. Store at -20 °C. Stable for 12 months from date of receipt.
Alias:

Fold/Unfold

AW228881; H2A histone family member X; H2A.FX; H2A.X; H2a/x; H2AFX; H2AX; H2AX histone; H2AX_HUMAN; Hist5.2ax; Histone 2A; Histone 2AX; Histone H2A.X; Histone H2AX; RGD1566119;

Immunogens

Immunogen:
Uniprot:
Gene(ID):
Sequence:
MSGRGKTGGKARAKAKSRSSRAGLQFPVGRVHRLLRKGHYAERVGAGAPVYLAAVLEYLTAEILELAGNAARDNKKTRIIPRHLQLAIRNDEELNKLLGGVTIAQGGVLPNIQAVLLPKKTSATVGPKAPSGGKKATQASQEY

Predictions

Predictions:

Score>80(red) has high confidence and is suggested to be used for WB detection. *The prediction model is mainly based on the alignment of immunogen sequences, the results are for reference only, not as the basis of quality assurance.

Species
Results
Score
Bovine
100
Sheep
100
Dog
100
Pig
0
Horse
0
Xenopus
0
Zebrafish
0
Chicken
0
Rabbit
0
Model Confidence:
High(score>80) Medium(80>score>50) Low(score<50) No confidence

PTMs - P16104 As Substrate

Site PTM Type Enzyme
S2 Phosphorylation
K6 Acetylation
K10 Acetylation
K14 Ubiquitination
K16 Ubiquitination
S19 Phosphorylation
K119 Acetylation
K119 Ubiquitination
K120 Ubiquitination
T121 Phosphorylation
S122 Phosphorylation
T124 Phosphorylation
K134 Methylation
K134 Sumoylation
K134 Ubiquitination
T137 Phosphorylation
S140 Phosphorylation Q13315 (ATM) , P45984 (MAPK9) , P45983 (MAPK8)
Y143 Phosphorylation Q9UIG0 (BAZ1B)

Research Backgrounds

Function:

Variant histone H2A which replaces conventional H2A in a subset of nucleosomes. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling. Required for checkpoint-mediated arrest of cell cycle progression in response to low doses of ionizing radiation and for efficient repair of DNA double strand breaks (DSBs) specifically when modified by C-terminal phosphorylation.

PTMs:

Phosphorylated on Ser-140 (to form gamma-H2AX or H2AX139ph) in response to DNA double strand breaks (DSBs) generated by exogenous genotoxic agents and by stalled replication forks, and may also occur during meiotic recombination events and immunoglobulin class switching in lymphocytes. Phosphorylation can extend up to several thousand nucleosomes from the actual site of the DSB and may mark the surrounding chromatin for recruitment of proteins required for DNA damage signaling and repair. Widespread phosphorylation may also serve to amplify the damage signal or aid repair of persistent lesions. Phosphorylation of Ser-140 (H2AX139ph) in response to ionizing radiation is mediated by both ATM and PRKDC while defects in DNA replication induce Ser-140 phosphorylation (H2AX139ph) subsequent to activation of ATR and PRKDC. Dephosphorylation of Ser-140 by PP2A is required for DNA DSB repair. In meiosis, Ser-140 phosphorylation (H2AX139ph) may occur at synaptonemal complexes during leptotene as an ATM-dependent response to the formation of programmed DSBs by SPO11. Ser-140 phosphorylation (H2AX139ph) may subsequently occurs at unsynapsed regions of both autosomes and the XY bivalent during zygotene, downstream of ATR and BRCA1 activation. Ser-140 phosphorylation (H2AX139ph) may also be required for transcriptional repression of unsynapsed chromatin and meiotic sex chromosome inactivation (MSCI), whereby the X and Y chromosomes condense in pachytene to form the heterochromatic XY-body. During immunoglobulin class switch recombination in lymphocytes, Ser-140 phosphorylation (H2AX139ph) may occur at sites of DNA-recombination subsequent to activation of the activation-induced cytidine deaminase AICDA. Phosphorylation at Tyr-143 (H2AXY142ph) by BAZ1B/WSTF determines the relative recruitment of either DNA repair or pro-apoptotic factors. Phosphorylation at Tyr-143 (H2AXY142ph) favors the recruitment of APBB1/FE65 and pro-apoptosis factors such as MAPK8/JNK1, triggering apoptosis. In contrast, dephosphorylation of Tyr-143 by EYA proteins (EYA1, EYA2, EYA3 or EYA4) favors the recruitment of MDC1-containing DNA repair complexes to the tail of phosphorylated Ser-140 (H2AX139ph).

Monoubiquitination of Lys-120 (H2AXK119ub) by RING1 and RNF2/RING2 complex gives a specific tag for epigenetic transcriptional repression (By similarity). Following DNA double-strand breaks (DSBs), it is ubiquitinated through 'Lys-63' linkage of ubiquitin moieties by the E2 ligase UBE2N and the E3 ligases RNF8 and RNF168, leading to the recruitment of repair proteins to sites of DNA damage. Ubiquitination at Lys-14 and Lys-16 (H2AK13Ub and H2AK15Ub, respectively) in response to DNA damage is initiated by RNF168 that mediates monoubiquitination at these 2 sites, and 'Lys-63'-linked ubiquitin are then conjugated to monoubiquitin; RNF8 is able to extend 'Lys-63'-linked ubiquitin chains in vitro. H2AK119Ub and ionizing radiation-induced 'Lys-63'-linked ubiquitination (H2AK13Ub and H2AK15Ub) are distinct events.

Acetylation at Lys-37 increases in S and G2 phases. This modification has been proposed to play a role in DNA double-strand break repair (By similarity).

Subcellular Location:

Nucleus. Chromosome.

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionSubcellular location
Subunit Structure:

The nucleosome is a histone octamer containing two molecules each of H2A, H2B, H3 and H4 assembled in one H3-H4 heterotetramer and two H2A-H2B heterodimers. The octamer wraps approximately 147 bp of DNA (Probable). Interacts with numerous proteins required for DNA damage signaling and repair when phosphorylated on Ser-140. These include MDC1, TP53BP1, BRCA1 and the MRN complex, composed of MRE11, RAD50, and NBN. Interaction with the MRN complex is mediated at least in part by NBN. Also interacts with DHX9/NDHII when phosphorylated on Ser-140 and MCPH1 when phosphorylated at Ser-140 or Tyr-143. Interacts with ARRB2; the interaction is detected in the nucleus upon OR1D2 stimulation. Interacts with WRAP53/TCAB1.

(Microbial infection) Interacts with Epstein-Barr virus protein EBNA6.

Family&Domains:

The [ST]-Q motif constitutes a recognition sequence for kinases from the PI3/PI4-kinase family.

Belongs to the histone H2A family.

Research Fields

· Cellular Processes > Cell growth and death > Necroptosis.   (View pathway)

· Human Diseases > Substance dependence > Alcoholism.

· Human Diseases > Immune diseases > Systemic lupus erythematosus.

References

1). Shikonin alleviates the biotoxicity produced by pneumococcal pneumolysin. LIFE SCIENCES, 2017 (PubMed: 28385613) [IF=6.1]

Application: IF/ICC    Species: mouse    Sample:

Figure 3. Shikonin inhibits PLY-induced nuclear DNA damage. (A) The solvent or shikonin solution does not lead to DSBs in cells. (B) Nuclear γH2AX foci in cells exposed in vitro to rPLY for 6 hours. (C) Histogram of DSBs foci, ** indicates p <0.01 compared with the shikonin-free group.

2). Chidamide-Induced Accumulation of Reactive Oxygen Species Increases Lenalidomide Sensitivity Against Multiple Myeloma Cells. OncoTargets and Therapy, 2021 (PubMed: 34262292) [IF=4.0]

Application: IF/ICC    Species: Human    Sample: ARP1 cells

Figure 6 Combination of chidamide (CHI) and lenalidomide (Len) increase ROS-related DNA damage in MM cells. (A and B) DNA damage analysis by γ-H2AX foci immunofluorescence. Representative images of γ-H2AX (Red) and nuclei (Blue) in ARP-1 and RPMI-8226 cells treated with single agents or combination for 24 hours. Scale bars represent 20 μm. (C and D) after treated with 1 μM chidamide and/ or 4 μM lenalidomide for 24 hours, expression of γ-H2AX was determined using Western blotting.

Application: WB    Species: Human    Sample: ARP1 cells

Figure 6 Combination of chidamide (CHI) and lenalidomide (Len) increase ROS-related DNA damage in MM cells. (A and B) DNA damage analysis by γ-H2AX foci immunofluorescence. Representative images of γ-H2AX (Red) and nuclei (Blue) in ARP-1 and RPMI-8226 cells treated with single agents or combination for 24 hours. Scale bars represent 20 μm. (C and D) after treated with 1 μM chidamide and/ or 4 μM lenalidomide for 24 hours, expression of γ-H2AX was determined using Western blotting.

3). Betulin attenuates pneumolysin‐induced cell injury and DNA damage. Journal of Applied Microbiology, 2021 (PubMed: 32621771) [IF=4.0]

Application: IF/ICC    Species: human    Sample: A549 cells

Fig 4. |Betulin inhibits PLY-induced DNA damage. The A549 cells were exposed in vitro to PLY with or without betulin for 6 h. (A) The images were selected from immunofluorescence assays and were captured by confocal microscopy.

4). Chidamide, a subtype-selective histone deacetylase inhibitor, enhances Bortezomib effects in multiple myeloma therapy. Journal of Cancer, 2023 (PubMed: 34539893) [IF=3.9]

Application: WB    Species: Human    Sample: MM cells

Figure 4 A combination of Chidamide and Bortezomib increases production of ROS dependent DNA damage and the changes of cell apoptosis and cycle pathway in MM cells. (A) ARP-1 cells were pretreated with or without NAC (15.0 mmol/L) for 2 hours at 37°C and then incubated with CHI (1.0 µmol/L) and/or BTZ (5.0 nmol/L) for 24 hours, then ROS generation was detected. (B) ARP-1 cells were pretreated with or without 15 mmol/L NAC and then treated with Chidamide or Bortezomib alone or in combination, and cell viabilities were evaluated using CCK-8 assays. (C) The expression of γ-H2AX in ARP-1 cells treated with CHI (1.0 µmol/L) and/or BTZ (5.0 nmol/L) were determined by Western blot. (D) Representative images of γ-H2AX (Red) and nuclei (Blue) in ARP-1 cells treated with single agent or combination for 24 hours by immunofluorescence assay. Scale bars represent 20 µm. (E, F) Western blot analysis of the expressions of cleaved caspase3, cleaved caspase8, cleaved PARP-1 and HDAC1 in XG1 (E) and ARP-1 (F) cells after 48 hours treatment with single agent or in combination. Error bars indicate mean ± SD. **p < 0.01.

Application: IF/ICC    Species: Human    Sample: MM cells

Figure 4 A combination of Chidamide and Bortezomib increases production of ROS dependent DNA damage and the changes of cell apoptosis and cycle pathway in MM cells. (A) ARP-1 cells were pretreated with or without NAC (15.0 mmol/L) for 2 hours at 37°C and then incubated with CHI (1.0 µmol/L) and/or BTZ (5.0 nmol/L) for 24 hours, then ROS generation was detected. (B) ARP-1 cells were pretreated with or without 15 mmol/L NAC and then treated with Chidamide or Bortezomib alone or in combination, and cell viabilities were evaluated using CCK-8 assays. (C) The expression of γ-H2AX in ARP-1 cells treated with CHI (1.0 µmol/L) and/or BTZ (5.0 nmol/L) were determined by Western blot. (D) Representative images of γ-H2AX (Red) and nuclei (Blue) in ARP-1 cells treated with single agent or combination for 24 hours by immunofluorescence assay. Scale bars represent 20 µm. (E, F) Western blot analysis of the expressions of cleaved caspase3, cleaved caspase8, cleaved PARP-1 and HDAC1 in XG1 (E) and ARP-1 (F) cells after 48 hours treatment with single agent or in combination. Error bars indicate mean ± SD. **p < 0.01.

5). Morin Moderates the Biotoxicity of Pneumococcal Pneumolysin by Weakening the Oligomers' Formation. CHEMICAL & PHARMACEUTICAL BULLETIN, 2017 (PubMed: 28566646) [IF=1.7]

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