Key Spec Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, M||IP, WB||M||Culture Supernatant||Monoclonal Antibody|
|Description||Anti-HDAC1 Antibody, clone 2E10|
|Presentation||100μl of culture supernatant containing 0.05% sodium azide. Frozen at -20°C.|
|Application||Detect HDAC1 with Anti-HDAC1 Antibody, clone 2E10 (Mouse Monoclonal Antibody), that has been shown to work in IP & WB.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||2 years at -20°C from date of shipment.|
|Material Size||100 µL|
|Reference overview||Application||Pub Med ID|
|Deacetylase inhibitors repress STAT5-mediated transcription by interfering with bromodomain and extra-terminal (BET) protein function.|
Pinz, S; Unser, S; Buob, D; Fischer, P; Jobst, B; Rascle, A
Nucleic acids research 43 3524-45 2015
Signal transducer and activator of transcription STAT5 is essential for the regulation of proliferation and survival genes. Its activity is tightly regulated through cytokine signaling and is often upregulated in cancer. We showed previously that the deacetylase inhibitor trichostatin A (TSA) inhibits STAT5-mediated transcription by preventing recruitment of the transcriptional machinery at a step following STAT5 binding to DNA. The mechanism and factors involved in this inhibition remain unknown. We now show that deacetylase inhibitors do not target STAT5 acetylation, as we initially hypothesized. Instead, they induce a rapid increase in global histone acetylation apparently resulting in the delocalization of the bromodomain and extra-terminal (BET) protein Brd2 and of the Brd2-associated factor TBP to hyperacetylated chromatin. Treatment with the BET inhibitor (+)-JQ1 inhibited expression of STAT5 target genes, supporting a role of BET proteins in the regulation of STAT5 activity. Accordingly, chromatin immunoprecipitation demonstrated that Brd2 is associated with the transcriptionally active STAT5 target gene Cis and is displaced upon TSA treatment. Our data therefore indicate that Brd2 is required for the proper recruitment of the transcriptional machinery at STAT5 target genes and that deacetylase inhibitors suppress STAT5-mediated transcription by interfering with Brd2 function.
|NFκB1 is a suppressor of neutrophil-driven hepatocellular carcinoma.|
Wilson, CL; Jurk, D; Fullard, N; Banks, P; Page, A; Luli, S; Elsharkawy, AM; Gieling, RG; Chakraborty, JB; Fox, C; Richardson, C; Callaghan, K; Blair, GE; Fox, N; Lagnado, A; Passos, JF; Moore, AJ; Smith, GR; Tiniakos, DG; Mann, J; Oakley, F; Mann, DA
Nature communications 6 6818 2015
Hepatocellular carcinoma (HCC) develops on the background of chronic hepatitis. Leukocytes found within the HCC microenvironment are implicated as regulators of tumour growth. We show that diethylnitrosamine (DEN)-induced murine HCC is attenuated by antibody-mediated depletion of hepatic neutrophils, the latter stimulating hepatocellular ROS and telomere DNA damage. We additionally report a previously unappreciated tumour suppressor function for hepatocellular nfkb1 operating via p50:p50 dimers and the co-repressor HDAC1. These anti-inflammatory proteins combine to transcriptionally repress hepatic expression of a S100A8/9, CXCL1 and CXCL2 neutrophil chemokine network. Loss of nfkb1 promotes ageing-associated chronic liver disease (CLD), characterized by steatosis, neutrophillia, fibrosis, hepatocyte telomere damage and HCC. Nfkb1(S340A/S340A)mice carrying a mutation designed to selectively disrupt p50:p50:HDAC1 complexes are more susceptible to HCC; by contrast, mice lacking S100A9 express reduced neutrophil chemokines and are protected from HCC. Inhibiting neutrophil accumulation in CLD or targeting their tumour-promoting activities may offer therapeutic opportunities in HCC.
|Regulation of CRADD-caspase 2 cascade by histone deacetylase 1 in gastric cancer.|
Shen, Q; Tang, W; Sun, J; Feng, L; Jin, H; Wang, X
American journal of translational research 6 538-47 2014
CRADD, also referred as RAIDD, is an adaptor protein that could interact with both caspase 2 and RIP that can promote apoptosis once activated. HDAC inhibitors are promising anti-cancer agents by inducing apoptosis of various cancer cells. In this study, we found that CRADD was induced by TSA (trichostatin A) to activate caspase 2-dependent apoptosis. CRADD was downregulated in gastric cancer and the restoration of its expression suppressed the viability of gastric cancer cells. HDAC1 was responsible for its downregulation in gastric cancer since HDAC1 siRNA upregulated CRADD expression and HDAC1 directly bound to the promoter of CRADD. Therefore, the high expression of HDAC1 can downregulate CRADD to confer gastric cancer cells the resistance to caspase 2-dependent apoptosis. HDAC inhibitors, potential anti-cancer drugs under investigation, can promote caspase 2-dependent apoptosis by inducing the expression of CRADD.
|Lysine deacetylase inhibition promotes relaxation of arterial tone and C-terminal acetylation of HSPB6 (Hsp20) in vascular smooth muscle cells.|
Chen, A; Karolczak-Bayatti, M; Sweeney, M; Treumann, A; Morrissey, K; Ulrich, SM; Europe-Finner, GN; Taggart, MJ
Physiological reports 1 e00127 2013
There is increasing interest in establishing the roles that lysine acetylation of non nuclear proteins may exert in modulating cell function. Lysine deacetylase 8 (KDAC8), for example, has been suggested to interact with α-actin and control the differentiation of smooth muscle cells. However, a direct role of smooth muscle non nuclear protein acetylation in regulating tone is unresolved. We sought to define the actions of two separate KDAC inhibitors on arterial tone and identify filament-interacting protein targets of acetylation and association with KDAC8. Compound 2 (a specific KDAC8 inhibitor) or Trichostatin A (TSA, a broad-spectrum KDAC inhibitor) inhibited rat arterial contractions induced by phenylephrine (PE) or high potassium solution. In contrast to the predominantly nuclear localization of KDAC1 and KDAC2, KDAC8 was positioned in extranuclear areas of native vascular smooth muscle cells. Several filament-associated proteins identified as putative acetylation targets colocalized with KDAC8 by immunoprecipitation (IP): cortactin, α-actin, tropomyosin, HSPB1 (Hsp27) and HSPB6 (Hsp20). Use of anti-acetylated lysine antibodies showed that KDAC inhibition increased acetylation of each protein. A custom-made antibody targeting the C-terminal acetylated lysine of human HSPB6 identified this as a novel target of acetylation that was increased by KDAC inhibition. HSPB6 phosphorylation, a known vasodilatory modification, was concomitantly increased. Interrogation of publicly available mass spectrometry data identified 50 other proteins with an acetylated C-terminal lysine. These novel data, in alliance with other recent studies, alert us to the importance of elucidating the mechanistic links between changes in myofilament-associated protein acetylation, in conjunction with other posttranslational modifications, and the regulation of arterial tone.
|Class I lysine deacetylases facilitate glucocorticoid-induced transcription.|
Kadiyala, V; Patrick, NM; Mathieu, W; Jaime-Frias, R; Pookhao, N; An, L; Smith, CL
The Journal of biological chemistry 288 28900-12 2013
Nuclear receptors use lysine acetyltransferases and lysine deacetylases (KDACs) in regulating transcription through histone acetylation. Lysine acetyltransferases interact with steroid receptors upon binding of an agonist and are recruited to target genes. KDACs have been shown to interact with steroid receptors upon binding to an antagonist. We have shown previously that KDAC inhibitors (KDACis) potently repress the mouse mammary tumor virus promoter through transcriptional mechanisms and impair the ability of the glucocorticoid receptor (GR) to activate it, suggesting that KDACs can play a positive role in GR transactivation. In the current study, we extended this analysis to the entire GR transcriptome and found that the KDACi valproic acid impairs the ability of agonist-bound GR to activate about 50% of its target genes. This inhibition is largely due to impaired transcription rather than defective GR processing and was also observed using a structurally distinct KDACi. Depletion of KDAC1 expression mimicked the effects of KDACi in over half of the genes found to be impaired in GR transactivation. Simultaneous depletion of KDACs 1 and 2 caused full or partial impairment of several more GR target genes. Altogether we found that Class I KDAC activity facilitates GR-mediated activation at a sizable fraction of GR-activated target genes and that KDAC1 alone or in coordination with KDAC2 is required for efficient GR transactivation at many of these target genes. Finally, our work demonstrates that KDACi exposure has a significant impact on GR signaling and thus has ramifications for the clinical use of these drugs.
|Efficient genetic manipulation of 1321N1 astrocytoma cells using lentiviral gene transfer.|
Anja Keim,Isabelle Müller,Gerald Thiel
Journal of neuroscience methods 206 2012
1321N1 astrocytoma cells are frequently used to analyze stimulus-induced intracellular signaling. These experiments require genetic manipulation of the cells and several chemical and physical methods have been employed in the past. Recently, microporation has been suggested as the best method to transfect 1321N1 astrocytoma cells. Here, we demonstrate that lentiviral gene transfer into 1321N1 cells is highly efficient, cheap and non-toxic. In addition, lentiviral gene transfer efficiently facilitates stable expression of small hairpin RNAs. Finally, lentiviral gene transfer can be used to implant promoter/luciferase reporter genes into the chromatin of the cells, allowing promoter studies using templates that are embedded into the nucleosomal structure of the chromatin.
|Breast cancer cells induce stromal fibroblasts to secrete ADAMTS1 for cancer invasion through an epigenetic change.|
Tyan, SW; Hsu, CH; Peng, KL; Chen, CC; Kuo, WH; Lee, EY; Shew, JY; Chang, KJ; Juan, LJ; Lee, WH
PloS one 7 e35128 2012
Microenvironment plays an important role in cancer development. We have reported that the cancer-associated stromal cells exhibit phenotypic and functional changes compared to stromal cells neighboring to normal tissues. However, the molecular mechanisms as well as the maintenance of these changes remain elusive. Here we showed that through co-culture with breast cancer cells for at least three to four passages, breast normal tissue-associated fibroblasts (NAFs) gained persistent activity for promoting cancer cell invasion, partly via up-regulating ADAM metallopeptidase with thrombospondin type 1 motif, 1 (ADAMTS1). Furthermore, we demonstrated that the DNA methylation pattern in the ADAMTS1 promoter has no alteration. Instead, the loss of EZH2 binding to the ADAMTS1 promoter and the resulting decrease of promoter-associated histone H3K27 methylation may account for the up-regulation of ADAMTS1. Importantly, the lack of EZH2 binding and the H3K27 methylation on the ADAMTS1 promoter were sustained in cancer cell-precocultured NAFs after removal of cancer cells. These results suggest that cancer cells are capable of inducing stromal fibroblasts to secrete ADAMTS1 persistently for their invasion and the effect is epigenetically inheritable.
|A transcriptional repressor co-regulatory network governing androgen response in prostate cancers.|
Chng, KR; Chang, CW; Tan, SK; Yang, C; Hong, SZ; Sng, NY; Cheung, E
The EMBO journal 31 2810-23 2012
Transcriptional corepressors are frequently aberrantly over-expressed in prostate cancers. However, their crosstalk with the Androgen receptor (AR), a key player in prostate cancer development, is unclear. Using ChIP-Seq, we generated extensive global binding maps of AR, ERG, and commonly over-expressed transcriptional corepressors including HDAC1, HDAC2, HDAC3, and EZH2 in prostate cancer cells. Surprisingly, our results revealed that ERG, HDACs, and EZH2 are directly involved in androgen-regulated transcription and wired into an AR centric transcriptional network via a spectrum of distal enhancers and/or proximal promoters. Moreover, we showed that similar to ERG, these corepressors function to mediate repression of AR-induced transcription including cytoskeletal genes that promote epithelial differentiation and inhibit metastasis. Specifically, we demonstrated that the direct suppression of Vinculin expression by ERG, EZH2, and HDACs leads to enhanced invasiveness of prostate cancer cells. Taken together, our results highlight a novel mechanism by which, ERG working together with oncogenic corepressors including HDACs and the polycomb protein, EZH2, could impede epithelial differentiation and contribute to prostate cancer progression, through directly modulating the transcriptional output of AR.
|Neural differentiation of P19 carcinoma cells and primary neurospheres: cell morphology, proliferation, viability, and functionality.|
Priscilla D Negraes,Telma T Schwindt,Cleber A Trujillo,Henning Ulrich
Current protocols in stem cell biology Chapter 2 2012
This unit describes the culture and induction of in vitro models of neural differentiation and strategies to evaluate the participation of extrinsic and intrinsic factors in modulation of this process. Protocols focus on large-scale expansion of pluripotent P19 murine embryonic carcinoma cells and their induction to neural differentiation in the presence of retinoic acid, closely resembling conditions of early neuroectodermal differentiation. Procedures are also described for obtaining rat neural precursor cells (NPCs) or neurospheres and for differentiating them in the absence of growth factors. Experimental strategies are reported using P19 cells and NPCs as in vitro models for studying the actions of extrinsic and intrinsic factors on morphology, proliferation, viability, neural phenotype determination, and progress of differentiation, as well as the functionality of ion channels and metabotropic receptors in inducing calcium fluxes at different developmental stages. The methods described here may be useful for optimizing in vitro protocols for stem cell differentiation into defined neural populations, as well as for studying mechanisms that underlie neurogenesis and gliogenesis.
|A radiosensitizing effect of artesunate in glioblastoma cells is associated with a diminished expression of the inhibitor of apoptosis protein survivin.|
Sebastian Reichert,Vera Reinboldt,Stephanie Hehlgans,Thomas Efferth,Claus RÃ¶del,Franz RÃ¶del
Radiotherapy and oncology : journal of the European Society for Therapeutic Radiology and Oncology 103 2012
Novel strategies to overcome an irradiation resistant phenotype may help to increase therapeutic efficacy in glioblastoma multiforme. The present study aimed to elucidate radiation sensitizing properties of artesunate, a semi synthetic derivate of artemisinin and to assess factors involved in this effect.
|How do I prepare a non-reducing sample buffer?||Non-reducing sample buffer can be made and stored for months at room temperature. Here is the formulation for a 2X Non-reducing sample buffer: Water (37ml), 0.5M Tris-HCl (10ml), Glycerol (8ml), 10% SDS (16ml), 0.05% Bromophenol Blue (5ml).|