| 货号 | 80312S |
| 同种亚型 | Mouse IgG1 |
| 反应种属 | Human, Mouse, Rat, Monkey |
| 应用 | WB,IHC,IF,F |
| 目标/特异性 | Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb recognizes endogenous levels of Histone H2A.X protein only when phosphorylated at Ser139. |
| 使用方法 | Western Blotting (1:1000) Immunohistochemistry (Paraffin) (1:200) Immunofluorescence (Immunocytochemistry) (1:100) Flow Cytometry (1:100) |
| 供应商 | CST |
| 灵敏度 | Endogenous |
| 背景 | Histone H2A.X is a variant histone that represents approximately 10% of the total H2A histone proteins in normal human fibroblasts (1). H2A.X is required for checkpoint-mediated cell cycle arrest and DNA repair following double-stranded DNA breaks (1). DNA damage, caused by ionizing radiation, UV-light, or radiomimetic agents, results in rapid phosphorylation of H2A.X at Ser139 by PI3K-like kinases, including ATM, ATR, and DNA-PK (2,3). Within minutes following DNA damage, H2A.X is phosphorylated at Ser139 at sites of DNA damage (4). This very early event in the DNA-damage response is required for recruitment of a multitude of DNA-damage response proteins, including MDC1, NBS1, RAD50, MRE11, 53BP1, and BRCA1 (1). In addition to its role in DNA-damage repair, H2A.X is required for DNA fragmentation during apoptosis and is phosphorylated by various kinases in response to apoptotic signals. H2A.X is phosphorylated at Ser139 by DNA-PK in response to cell death receptor activation, c-Jun N-terminal Kinase (JNK1) in response to UV-A irradiation, and p38 MAPK in response to serum starvation (5-8). H2A.X is constitutively phosphorylated on Tyr142 in undamaged cells by WSTF (Williams-Beuren syndrome transcription factor) (9,10). Upon DNA damage, and concurrent with phosphorylation of Ser139, Tyr142 is dephosphorylated at sites of DNA damage by recruited EYA1 and EYA3 phosphatases (9). While phosphorylation at Ser139 facilitates the recruitment of DNA repair proteins and apoptotic proteins to sites of DNA damage, phosphorylation at Tyr142 appears to determine which set of proteins are recruited. Phosphorylation of H2A.X at Tyr142 inhibits the recruitment of DNA repair proteins and promotes binding of pro-apoptotic factors such as JNK1 (9). Mouse embryonic fibroblasts expressing only mutant H2A.X Y142F, which favors recruitment of DNA repair proteins over apoptotic proteins, show a reduced apoptotic response to ionizing radiation (9). Thus, it appears that the balance of H2A.X Tyr142 phosphorylation and dephosphorylation provides a switch mechanism to determine cell fate after DNA damage. |
| 存放说明 | -20C |
| 计算分子量 | 15 |
| 参考文献 | 1 . Yuan, J. et al. (2010) FEBS Lett 584, 3717-24. 2 . Rogakou, E.P. et al. (1998) J Biol Chem 273, 5858-68. 3 . Burma, S. et al. (2001) J Biol Chem 276, 42462-7. 4 . Rogakou, E.P. et al. (1999) J Cell Biol 146, 905-16. 5 . Mukherjee, B. et al. (2006) DNA Repair (Amst) 5, 575-90. 6 . Solier, S. et al. (2009) Mol Cell Biol 29, 68-82. 7 . Lu, C. et al. (2006) Mol Cell 23, 121-32. 8 . Lu, C. et al. (2008) FEBS Lett 582, 2703-8. 9 . Cook, P.J. et al. (2009) Nature 458, 591-6. 10 . Xiao, A. et al. (2009) Nature 457, 57-62. |
Western blot analysis of extracts from HeLa cells, untreated (-), UV-treated (+), or UV-treated and subsequently treated with calf intestinal phosphatase (CIP) plus λ-phosphatase (+), using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb (upper) or Histone H2A (D6O3A) Rabbit mAb (lower). | |
Immunohistochemical analysis of paraffin-embedded human appendix using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb. | |
Immunohistochemical analysis of paraffin-embedded human breast carcinoma, untreated (left) or λ-phosphatase-treated (right), using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb | |
Immunohistochemical analysis of paraffin-embedded OVCAR8 cell pellets, untreated (left) or UV-treated (right), using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb. | |
Immunohistochemical analysis of paraffin-embedded human lymph node using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb. | |
Immunohistochemical analysis of paraffin-embedded human endometrioid adenocarcinoma using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb. | |
Confocal immunofluorescent analysis of HeLa cells, untreated (left), treated with UV (100 mJ/cm2, 2 hr recovery; center), or treated with UV and post-processed with λ-phosphatase (right), using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red). | |
Flow cytometric analysis of HeLa cells, untreated (blue) or treated with UV (100 mJ, 2 hr recovery; green), using Phospho-Histone H2A.X (Ser139) (D7T2V) Mouse mAb (solid lines) or concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control #5415 (dashed lines). Anti-mouse IgG (H+L), F(ab)2 Fragment (Alexa Fluor® 488 Conjugate) #4408 was used as a secondary antibody. |
