Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

BRD4 (E2A7X) Rabbit mAb #13440

No. Size Price
13440S 100 µl ( 10 western blots ) ¥3,100.00 现货查询 购买询价
13440 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human, Endogenous 200 Rabbit IgG
IP 1:50
ChIP 1:50

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, ChIP=Chromatin IP,

Homology

Species predicted to react based on 100% sequence homology: Bovine, Dog, Pig,

Specificity / Sensitivity

BRD4 (E2A7X) Rabbit mAb recognizes endogenous levels of total BRD4 protein. This antibody specifically recognizes the BRD4 long isoform (UniProt #O60885-1) and does not recognize other BRD4 isoforms.

BRD4 (E2A7X) Rabbit mAb 兔单抗能够检测内源性BRD4总蛋白水平。该抗体能特异性识别BRD4长亚型(UniProt#O60885-1),不识别其他BRD4亚型。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu1129 of human BRD4 protein.

该单克隆抗体是采用合成的与人源BRD4蛋白Leu129残基周围序列相对应的肽段免疫动物而生产的。

Western Blotting

Western Blotting

Western blot analysis of extracts from RL-7, 293T, and Jurkat cells using BRD4 (E2A7X) Rabbit mAb.Western blot方法检测RL-7, 293T,以及Jurkat细胞系的提取物,使用的抗体为BRD4 (E2A7X) Rabbit mAb。

IP

IP

Immunoprecipitation of BRD4 from RL-7 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or BRD4 (E2A7X) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using BRD4 (E2A7X) Rabbit mAb.从RL-7细胞提取物中免疫沉淀BRD4,使用的抗体为Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) 或BRD4 (E2A7X) Rabbit mAb (lane 3)。Lane 1为10% input。 使用BRD4 (E2A7X) Rabbit mAb进行Western blot检测。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 MV-4-11 cells and either 10 µl of BRD4 (E2A7X) Rabbit mAb or 2 µl of Normal Rabbit IgG #2729, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human Bcl-2 Promoter Primers #12924, human c-Myc intron 1 primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.使用SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003 对4 x 106 MV-4-11细胞中交联过的染色质进行染色质免疫沉淀,所用抗体为10 µl BRD4 (E2A7X) Rabbit mAb 或2 µl Normal Rabbit IgG #2729,富集到的DNA经过实时PCR定量,所使用产品为SimpleChIP® Human Bcl-2 Promoter Primers #12924, human c-Myc intron 1 primers以及SimpleChIP® Human α Satellite Repeat Primers #4486。每个样品中免疫沉淀得到的DNA量由与input chromatin( 相当于1)的相对信号值来表示。

Background

Bromodomain-containing protein 4 (BRD4) is a member of the bromodomains and extra terminal (BET) family of proteins, which also includes BRD2, BRD3, and BRDT (1-3). BET family proteins contain two tandem bromodomains and an extra terminal (ET) domain, and bind acetyl lysine residues (3). BRD4 is a chromatin-binding protein with a preference for Lys14 on histone H3 as well as Lys5 and Lys12 on histone H4 (4). BRD4 chromatin binding occurs throughout the cell cycle, including condensed mitotic chromosomes, when the majority of genes are silenced (5). BRD4 association with chromatin during mitosis is thought to be an important part of the bookmarking mechanism to accelerate re-activation of the silenced genes upon exit from mitosis (2,6). BRD4 has been shown to facilitate transcription by recruiting the positive transcription elongation factor b (pTEFb) complex that phosphorylates Ser2 of the heptapeptide repeat of the carboxy-terminal domain of RNA polymerase II, promoting transcription elongation (3,7,8). In addition, BRD4 has been found to be part of the super elongation complex and the polymerase associated factor complex (PAFc) in MLL-fusion derived leukemia cell lines, demonstrating a role for BRD4 in the regulation of transcription elongation (9). Research studies have shown that BRD4 (and BET family proteins) may be promising therapeutic targets for various Myc-driven cancers, such as Burkitt’s lymphoma and certain acute myeloid leukemias (1,10,11). Investigators have found molecular inhibition of BET proteins to be effective in inducing apoptosis in various MLL-fusion driven leukemic cell lines by competing BRD3 and BRD4 from chromatin, leading to reduced expression of Bcl-2, Myc, and CDK6 (9). BET inhibition has also been shown to have antitumor activities against nuclear protein in testis (NUT) midline carcinoma cell lines and xenografts in mice where BRD4 is found to be a frequent translocation partner of the NUT protein (12). In addition, BRD4 regulates the expression of some inflammatory genes, and inhibition of BRD4 (and BET family proteins) chromatin binding causes reduced expression of a subset of inflammatory genes in macrophages, leading to protection against endotoxic shock and sepsis (13).

Bromodomain-containing protein 4(BRD4)是Bromodomains and extrateminal (BET)家族成员之一,该家族还包含了BRD2,BRD3和BRDT等(1-3)。BET蛋白家族包含了两个串联的溴区结构域和一个额外的尾部结构域,可以结合乙酰化的赖氨酸残基(3)。BRD4是染色质结合蛋白,倾向于结合组蛋白H3的Lys14残基和组蛋白H4的Lys5及Lys12位点。BRD4结合染色质通常发生于整个细胞周期,其中包括当主要基因处于沉默时,结合凝集的有丝分裂的染色体。BRD4结合有丝分裂时期的染色体,被认为是书签机制的一个重要组成部分,在细胞退出有丝分裂时加速激活沉默基因。BRD4招募正转录延伸因子b(pTEFb)复合物促进转录进行,pTEFb磷酸化 RNA聚合酶II的C端结构域的七肽重复区域的Ser2,从而转录的延伸(3,7,8)。此外,已经发现在MLL融合的白血病细胞株中,BRD4是超延伸复合物和聚合酶相关因子复合物(PAFc)的一部分,这也证实BRD4调控转录延伸的作用。调查研究表明,BRD4(和BET 家族蛋白)有可能成为有前途的治疗各种由Myc驱动的各种癌症的药物,如Burkitt淋巴瘤和某些急性髓细胞性白血病(1,10,11)。研究者发现,BET蛋白能够与BRD3和BRD4竞争性结合染色质,降低Bcl-2,Myc和CDK的表达,减少细胞凋亡。对BET蛋白进行抑制,可以有效引起各种MLL融合的白血病细胞的凋亡(9)。在睾丸(NUT)中线癌细胞线和异种移植小鼠中,BRD4被证实是核蛋白NUT频繁易位的伴侣蛋白,抑制BRD4活性具有针对核蛋白的抗肿瘤活性(12)。此外,BRD4也能调节一些炎症基因的表达。抑制BRD4(和BET家族蛋白)结合染色质的能力,能够减少一系列炎症基因在巨噬细胞中的表达,防止内毒性休克和败血症产生(13)。

  1. Belkina, A.C. and Denis, G.V. (2012) Nat Rev Cancer 12, 465-77.
  2. Voigt, P. and Reinberg, D. (2011) Genome Biol 12, 133.
  3. Wu, S.Y. and Chiang, C.M. (2007) J Biol Chem 282, 13141-5.
  4. Dey, A. et al. (2003) Proc Natl Acad Sci U S A 100, 8758-63.
  5. Dey, A. et al. (2009) Mol Biol Cell 20, 4899-909.
  6. Zhao, R. et al. (2011) Nat Cell Biol 13, 1295-304.
  7. Jang, M.K. et al. (2005) Mol Cell 19, 523-34.
  8. Yang, Z. et al. (2005) Mol Cell 19, 535-45.
  9. Dawson, M.A. et al. (2011) Nature 478, 529-33.
  10. Muller, S. et al. (2011) Expert Rev Mol Med 13, e29.
  11. Mertz, J.A. et al. (2011) Proc Natl Acad Sci U S A 108, 16669-74.
  12. Filippakopoulos, P. et al. (2010) Nature 468, 1067-73.
  13. Nicodeme, E. et al. (2010) Nature 468, 1119-23.

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