Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

SIN3A (D9D6) Rabbit mAb #8056

No. Size Price
8056S 100 µl ( 10 western blots ) ¥3,100.00 现货查询 购买询价
8056 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Monkey, Endogenous 145 Rabbit IgG
IP 1:50
IF-IC 1:200
ChIP 1:50

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, IF-IC=Immunofluorescence (Immunocytochemistry), ChIP=Chromatin IP,

Specificity / Sensitivity

SIN3A (D9D6) Rabbit mAb recognizes endogenous levels of total SIN3A protein. Based on protein sequence, this antibody is not predicted to cross-react with SIN3B protein.

SIN3A (D9D6) Rabbit mAb兔单抗能够检测内源性SIN3A总蛋白水平。基于蛋白序列,该抗体不能与SIN3B蛋白发生交叉反应。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu530 of human SIN3A protein.

通过人工合成人源SIN3A蛋白Leu530位点周围相应的多肽片段去免疫动物从而制备出此单克隆抗体。

IF-IC

IF-IC

Confocal immunofluorescent analysis of 293 cells using SIN3A (D9D6) Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red).

使用SIN3A (D9D6) Rabbit mAb兔单抗 (绿色),共聚焦免疫荧光分析293细胞。DY-554 phalloidin标记微丝蛋白(红色)。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 MCF7 cells grown in phenol red free medium and charcoal stripped FBS for 4 days then treated with β-estradiol (10 nM) for 1 hr and either 10 μl of SIN3A (D9D6) 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 ESR1 Promoter Primers #9673, SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human MyoD Exon 1 Primers #4490, 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,来自在无酚红培养基和葡聚糖处理胎牛血清中培养4天之后处理了β-estradiol (10 nM) 1小时的4 x 106 MCF7细胞的交联染色质以及10µl SIN3A (D9D6) Rabbit mAb或2 µl Normal Rabbit IgG #2729进行染色质免疫沉淀实验。使用SimpleChIP® Human ESR1 Promoter Primers #9673、SimpleChIP® Human GAPDH Exon 1 Primers #5516、SimpleChIP® Human MyoD Exon 1 Primers #4490和SimpleChIP® Human α Satellite Repeat Primers #4486,浓缩的DNA通过real-time PCR定量。在每个样品中免疫沉淀DNA的数量被当做一个相对于总input chromatin的数量的信号,这相当于一。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using SIN3A (D9D6) Rabbit mAb.

使用SIN3A (D9D6) Rabbit mAb兔单抗,免疫印迹(Western blot)分析不同细胞中SIN3A蛋白水平。

Background

SIN3 was originally identified as a negative regulator of transcription in budding yeast (1,2). Since then, three isoforms of the SIN3 proteins have been identified in mammalian cells, as products of two different genes, SIN3A and SIN3B (3,4). Both SIN3A and SIN3B are nuclear proteins that function as scaffolding subunits for the multi-subunit SIN3 transcriptional repressor complex, containing SIN3A or SIN3B, HDAC1, HDAC2, SDS3, RBBP4/RBAP48, RBBP7/RBAP46, SAP30, and SAP18 (3,4). SIN3 proteins contain four paired amphipathic alpha-helix (PAH) motifs that function in the recruitment of the SIN3 complex to target genes by binding a multitude of DNA-binding transcriptional repressor proteins, including Mad1, p53, E2F4, HCF-1, AML1, Elk-1, NRSF, CTCF, ERα, and MeCP2 (3,4). In addition, SIN3 proteins contain an HDAC interaction domain (HID), which mediates binding of HDAC1 and HDAC2 via the SDS3 bridging protein, and a highly conserved region (HCR) at the carboxy terminus, which contributes to repressor protein binding (3,4). RBBP4 and RBBP7 proteins also bind to SDS3 and contribute to nucleosome binding of the complex. The SIN3 complex functions to repress transcription, in part, by deacetylating histones at target gene promoters (3,4). In addition, recent studies have shown that SIN3 is recruited to the coding regions of repressed and active genes, where it deacetylates histones and suppresses spurious transcription by RNA polymerase II (3,5). In addition to histone deacetylase activity, the SIN3 complex associates with histone methyltransferase (ESET), histone demethylase (JARID1A/RBP2), ATP-dependent chromatin remodeling (SWI/SNF), methylcytosine dioxygenase (TET1), and O-GlcNAc transferase (OGT) activities, all of which appear to contribute to the regulation of target genes (5-9). The SIN3 complex is critical for proper regulation of embryonic development, cell growth and proliferation, apoptosis, DNA replication, DNA repair, and DNA methylation (imprinting and X-chromosome inactivation) (3,4).

SIN3 起初被鉴定在出芽酵母中作为转录的负性调节因子(1,2)。自从那时起,SIN3蛋白的三个亚基在哺乳动物中被鉴定,如同两个不同基因产物一样SIN3A和SIN3B (3,4)。SIN3A和SIN3B都是细胞核蛋白,它们的功能是作为多亚单位SIN3转录抑制复合物的支架亚单位,该复合物包含SIN3A或SIN3B、HDAC1、HDAC2、SDS3、RBBP4/RBAP48、RBBP7/RBAP46、SAP30, and SAP18 (3,4)。SIN3蛋白包含四个paired amphipathic alpha-helix (PAH)结构域,该结构域通过结合大量DNA-结合转录抑制蛋白使SIN3复合物招募到靶基因上 ,而这些转录抑制蛋白包含Mad1、 p53、E2F4、HCF-1、AML1、Elk-1、NRSF、CTCF、ERα和MeCP2 (3,4)。因此,SIN3蛋白包含一个HDAC interaction domain (HID),该结构域通过SDS3桥接蛋白介导HDAC1和HDAC2的结合,以及在羧基端含有一个highly conserved region (HCR),该区域有助于抑制性蛋白质结合(3,4)。RBBP4和RBBP7蛋白也结合到SDS3,并且有助于核小体复合物的结合。SIN3复合物的功能去抑制转录,在部分程度上时通过靶基因启动子上的去乙酰化的组蛋白(3,4)。因此,最近研究证明SIN3被招募到抑制性和活性基因的编码区域,在该区域它能使组蛋白去乙酰化以及通过RNA polymerase II抑制假转录(3,5)。除了组蛋白去乙酰化酶活性之外,SIN3复合物与组蛋白甲基化转移酶(ESET)、组蛋白去甲基酶 (JARID1A/RBP2)、ATP依赖的染色质重塑(SWI/SNF)、甲基胞嘧啶双加氧酶(TET1)和O-GlcNAc transferase (OGT) 有关联,这些蛋白都有助于靶基因的调节(5-9)。SIN3复合物对于胚胎发育、细胞生长和增殖、凋亡、DNA复制、DNA修复和DNA甲基化(印记和x染色体失活)起着关键作用(3,4)。

  1. Sternberg, P.W. et al. (1987) Cell 48, 567-77.
  2. Nasmyth, K. et al. (1987) Cell 48, 579-87.
  3. Grzenda, A. et al. Biochim Biophys Acta 1789, 443-50.
  4. McDonel, P. et al. (2009) Int J Biochem Cell Biol 41, 108-16.
  5. van Oevelen, C. et al. (2008) Mol Cell 32, 359-70.
  6. Yang, L. et al. (2003) Biochem J 369, 651-7.
  7. Sif, S. et al. (2001) Genes Dev 15, 603-18.
  8. Williams, K. et al. (2011) Nature 473, 343-8.
  9. Yang, X. et al. (2002) Cell 110, 69-80.

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Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.

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