Trichostatin A Relieves Growth Suppression and Restores Histone Acetylation at Specific Sites in a FUS ALS/FTD Yeast Model

The embryogenic transition of somatic cells requires an extensive reprogramming of the cell transcriptome. Relevantly, the extensive modulation of the genes that have a regulatory function, in particular the genes encoding the transcription factors (TFs) and miRNAs, have been indicated as controlling somatic embryogenesis (SE) that is induced in vitro in the somatic cells of plants. Identifying the regulatory relationships between the TFs and miRNAs during SE induction is of central importance for understanding the complex regulatory interplay that fine-tunes a cell transcriptome during the embryogenic transition. Hence, here, we analysed the regulatory relationships between AGL15 (AGAMOUS-LIKE 15) TF and miR156 in an embryogenic culture of Arabidopsis. Both AGL15 and miR156 control SE induction and AGL15 has been reported to target the MIR156 genes in planta. The results showed that AGL15 contributes to the regulation of miR156 in an embryogenic culture at two levels that involve the activation of the MIR156 transcription and the containment of the abundance of mature miR156 by repressing the miRNA biogenesis genes DCL1 (DICER-LIKE1), SERRATE and HEN1 (HUA-ENHANCER1). To repress the miRNA biogenesis genes AGL15 seems to co-operate with the TOPLESS co-repressors (TPL and TPR1-4), which are components of the SIN3/HDAC silencing complex. The impact of TSA (trichostatin A), an inhibitor of the HDAC histone deacetylases, on the expression of the miRNA biogenesis genes together with the ChIP results implies that histone deacetylation is involved in the AGL15-mediated repression of miRNA processing. The results indicate that HDAC6 and HDAC19 histone deacetylases might co-operate with AGL15 in silencing the complex that controls the abundance of miR156 during embryogenic induction. This study provides new evidence about the histone acetylation-mediated control of the miRNA pathways during the embryogenic reprogramming of plant somatic cells and the essential role of AGL15 in this regulatory mechanism.

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P1817DOES EPIGENETIC FACTORS LIKE HDAC2 PLAYS ROLE IN STEROID RESISTANCE VIA REGULATION OF P-GP AND MRP-1 IN CHILDHOOD IDIOPATHIC NEPHROTIC SYNDROME?

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfaa142.p1817 ◽

2020 ◽

Vol 35 (Supplement_3) ◽

Author(s):

HARSHIT SINGH ◽

Narayan Prasad ◽

Vikas Agarwal ◽

Akhilesh Kumar Jaiswal

Keyword(s):

Nephrotic Syndrome ◽

Histone Acetylation ◽

Mrna Expression ◽

Idiopathic Nephrotic Syndrome ◽

Trichostatin A ◽

Fold Change ◽

Steroid Resistance ◽

Mrna Levels ◽

Inflammatory Genes ◽

Maximal Induction

Abstract Background and Aims The action of glucocorticoids is to switch off activated inflammatory genes. The activated glucocorticoid receptors (GR) interact with co-repressor molecules to impair NFκB-associated co-activator activity, reducing histone acetylation, chromatin remodelling. Reduction in histone acetylation occurs via recruitment of histone deacetylase (HDAC) 2 to the activated inflammatory gene complex by activated GR, resulting in efficacious suppression of activated inflammatory genes within the nucleus. To evaluate the effect HDAC 2 on P-gp and MRP-1 expression and function. Method Total of 78 subjects were considered in the study out of which 50 were steroid sensitive nephrotic syndrome (SSNS), and 28 were steroid resistant nephrotic syndrome (SRNS) patients. mRNA expression was analyzed on peripheral blood mononuclear cells (PBMCs) in SRNS patients (mean age 8.43±3.8 years), SSNS patients (mean age 7.54±3.5 years). PBMCs were treated with 1µM of Theophylline (HDAC2 stimulator) and 0.8µM of Trichostatin A (HDAC2 inhibitor) for a period of 48 hours. Quantitative PCR was performed using light cycler LC480 using SYBR green PCR technology with SYBR premix relative gene expression levels were calculated and normalized to the corresponding levels of the housekeeping gene (GAPDH). Results Expression of P-gp (4.79±0.970 v/s 2.13±0.72, p<0.0001) and MRP-1 (3.99 ±0.8 v/s 1.99 ±0.91, p<0.0001) on PBMCs was increased in SRNS as compared to that of SSNS. HDAC2 mRNA levels were significantly decreased in SRNS patients as compared to that of SSNS patients (2.97 ± 0.95 v/s 6.02 ± 1.13, p<0.0001). Theophylline(HDAC stimulator) for a period of 48 hours decreased mRNA levels of P-gp and MRP-1 in PBMCs of SRNS with maximal induction at 1µM (fold change 2.65 and 2.21, *p<0.0001) However HDAC2 mRNA expression increased significantly (fold change5.67, *p<0.0001). In SSNS patients P-gp and MRP-1 mRNA expression decreased at1µM (fold change 1.25, 1.24, *p<0.0001) while the mRNA expression was increased (fold change 6.93, *p<0.0001). TSA(HDAC inhibitor) for a period of 48 hours increased mRNA levels of P-gp and MRP-1 in PBMCs of SRNS with maximal induction at 0.8µM (fold change 7.51, 7.31, *p<0.0001) and significantly decreased the level of HDAC2 (fold change1.50, *p<0.0001) similarly in SSNS patients P-gp and MRP-1 mRNA expression increased at 0.8µM (fold change 3.49, 3.35, *p<0.0001) and HDAC2 decreased (fold change2.53, *p<0.0001) at 0.8µM. The functional activity of P-gp and MRP-1 was significantly higher in SRNS group as compared to SSNS group (p<0.001), whereas enzymatic activity of HDAC2 was increased in SSNS group as compared to SRNS group (p<0.001) Conclusion As we observed that HDAC2 regulates P-gp and MRP-1 efflux pumps, Inducer of HDAC2 may be a probable treatment stratergy for patients of Idiopathic Nephrotic Syndrome.

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Effects of histone deacetylase inhibitors on estradiol-induced proliferation and hyperplasia formation in the mouse uterus

Journal of Endocrinology ◽

10.1677/joe.1.06118 ◽

2005 ◽

Vol 185 (3) ◽

pp. 539-549 ◽

Cited By ~ 18

Author(s):

Andrei G Gunin ◽

Irina N Kapitova ◽

Nina V Suslonova

Keyword(s):

Estrogen Receptor ◽

Histone Acetylation ◽

Histone Deacetylase ◽

Sodium Butyrate ◽

Histone Deacetylase Inhibitors ◽

Trichostatin A ◽

Progesterone Receptors ◽

Estrogen Receptor Α ◽

Mouse Uterus ◽

Myometrial Cells

It is suggested that estrogen hormones recruit mechanisms controlling histone acetylation to bring about their effects in the uterus. However, it is not known how the level of histone acetylation affects estrogen-dependent processes in the uterus, especially proliferation and morphogenetic changes. Therefore, this study examined the effects of histone deacetylase blockers, trichostatin A and sodium butyrate, on proliferative and morphogenetic reactions in the uterus under long-term estrogen treatment. Ovari-ectomized mice were treated with estradiol dipropionate (4 μg per 100 g; s.c., once a week) or vehicle and trichostatin A (0.008 mg per 100 g; s.c., once a day) or sodium butyrate (1% in drinking water), or with no additional treatments for a month. In animals treated with estradiol and trichostatin A or sodium butyrate, uterine mass was increased, and abnormal uterine glands and atypical endometrial hyperplasia were found more often. Both histone deacetylase inhibitors produced an increase in the numbers of mitotic and bromodeoxyuridine-labelled cells in luminal and glandular epithelia, in stromal and myometrial cells. Levels of estrogen receptor-α and progesterone receptors in uterine epithelia, stromal and myometrial cells were decreased in mice treated with estradiol and trichostatin A or sodium butyrate. Expression of β-catenin in luminal and glandular epithelia was attenuated in mice treated with estradiol with trichostatin A or sodium butyrate. Both histone deacetylase inhibitors have similar unilateral effects; however the action of trichostatin A was more expressed than that of sodium butyrate. Thus, histone deacetylase inhibitors exert proliferative and morphogenetic effects of estradiol. The effects of trichostatin A and sodium butyrate are associated with changes in expression of estrogen receptor-α, progesterone receptors and β-catenin in the uterus.

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Molecular inhibition of histone deacetylation results in major enhancement of the production of IL-1β in response to LPS

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00406.2005 ◽

2006 ◽

Vol 290 (3) ◽

pp. E490-E493 ◽

Cited By ~ 13

Author(s):

Timothy A. Sato ◽

Murray D. Mitchell

Keyword(s):

Dna Methylation ◽

Receptor Antagonist ◽

Histone Acetylation ◽

Trichostatin A ◽

Cell Types ◽

Causative Factor ◽

Histone Deacetylation ◽

The Other ◽

Acetylation Status ◽

Tnf Α

It has been postulated that the progression of human pregnancy to term is, in part, the result of a relative maternal Th2 immunological state. This can be activated in some cell types by modifying DNA methylation and histone acetylation status. We demonstrate that the molecular inhibition of histone deacetylation, using trichostatin A (TSA), in human choriodecidual explants leads to a massive increase in lipopolysaccharide (LPS)-stimulated IL-1β. The inhibition of histone deacetylation had no effect on LPS-stimulated TNF-α production or production of the other cytokines studied (IL-10, IL-1 receptor antagonist). The molecular inhibition of DNA methylation and histone deacetylation, using 5-aza-2′-deoxycytidine and TSA, respectively, in human choriodecidual explants also results in an increase in the basal production of TNF-α but not that of IL-1β. The differential response is unique, and the relative uncoupling of IL-1β and TNF-α responsiveness may have importance in other biological systems and provide new therapeutic targets for pathologies where upregulation of IL-1β is known to be a causative factor.

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Circadian and Light-Induced Transcription of Clock Gene Per1 Depends on Histone Acetylation and Deacetylation

Molecular and Cellular Biology ◽

10.1128/mcb.24.14.6278-6287.2004 ◽

2004 ◽

Vol 24 (14) ◽

pp. 6278-6287 ◽

Cited By ~ 144

Author(s):

Yoshihisa Naruse ◽

Kentaro Oh-hashi ◽

Norio Iijima ◽

Midori Naruse ◽

Hideyo Yoshioka ◽

...

Keyword(s):

Histone Acetylation ◽

Histone Deacetylases ◽

Feedback Loop ◽

Clock Gene ◽

Clock Genes ◽

Trichostatin A ◽

Chromatin Modification ◽

Histone Acetyltransferases ◽

Fibroblast Cells ◽

Circadian Clock Genes

ABSTRACT Circadian clock genes are regulated through a transcriptional-translational feedback loop. Alterations of the chromatin structure by histone acetyltransferases and histone deacetylases (HDACs) are commonly implicated in the regulation of gene transcription. However, little is known about the transcriptional regulation of mammalian clock genes by chromatin modification. Here, we show that the state of acetylated histones fluctuated in parallel with the rhythm of mouse Per1 (mPer1) or mPer2 expression in fibroblast cells and liver. Mouse CRY1 (mCRY1) repressed transcription with HDACs and mSin3B, which was relieved by the HDAC inhibitor trichostatin A (TSA). In turn, TSA induced endogenous mPer1 expression as well as the acetylation of histones H3 and H4, which interacted with the mPer1 promoter region in fibroblast cells. Moreover, a light pulse stimulated rapid histone acetylation associated with the promoters of mPer1 or mPer2 in the suprachiasmatic nucleus (SCN) and the binding of phospho-CREB in the CRE of mPer1. We also showed that TSA administration into the lateral ventricle induced mPer1 and mPer2 expression in the SCN. Taken together, these data indicate that the rhythmic transcription and light induction of clock genes are regulated by histone acetylation and deacetylation.

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NuA4 Subunit Yng2 Function in Intra-S-Phase DNA Damage Response

Molecular and Cellular Biology ◽

10.1128/mcb.22.23.8215-8225.2002 ◽

2002 ◽

Vol 22 (23) ◽

pp. 8215-8225 ◽

Cited By ~ 85

Author(s):

John S. Choy ◽

Stephen J. Kron

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Histone Acetylation ◽

Trichostatin A ◽

S Phase ◽

Open Chromatin ◽

Histone H4 ◽

Histone H4 Acetylation ◽

Histone Deacetylase Inhibitor Trichostatin ◽

Nucleosomal Histone

ABSTRACT While regulated transcription requires acetylation of histone N-terminal tails to promote an open chromatin conformation, a similar role for histone acetylation in DNA replication and/or repair remains to be established. Cells lacking the NuA4 subunit Yng2 are viable but critically deficient for genome-wide nucleosomal histone H4 acetylation. We found that yng2 mutants are specifically sensitized to DNA damage in S phase induced by cdc8 or cdc9 mutations, hydroxyurea, camptothecin, or methylmethane sulfonate (MMS). In yng2, MMS treatment causes a persistent Mec1-dependent intra-S-phase checkpoint delay characterized by slow DNA repair. Restoring H4 acetylation with the histone deacetylase inhibitor trichostatin A promotes checkpoint recovery. In turn, mutants lacking the histone H3-specific acetyltransferase GCN5 are similarly sensitive to intra-S-phase DNA damage. The inviability of gcn5 yng2 double mutants suggests overlapping roles for H3 and H4 acetylation in DNA replication and repair. Paradoxically, haploid yng2 mutants do not tolerate mutations in genes important for nonhomologous end joining repair yet remain proficient for homologous recombination. Our results implicate nucleosomal histone acetylation in maintaining genomic integrity during chromosomal replication.

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Acetylation of Histone H3 and Adrenergic-Regulated Gene Transcription in Rat Pinealocytes

Endocrinology ◽

10.1210/en.2007-0578 ◽

2007 ◽

Vol 148 (10) ◽

pp. 4592-4600 ◽

Cited By ~ 13

Author(s):

A. K. Ho ◽

D. M. Price ◽

W. G. Dukewich ◽

N. Steinberg ◽

T. G. Arnason ◽

...

Keyword(s):

Histone Acetylation ◽

Histone Deacetylase ◽

Gene Transcription ◽

Histone Deacetylase Inhibitor ◽

Trichostatin A ◽

Histone H3 ◽

Dna Recovery ◽

Inducible Camp Early Repressor ◽

Deacetylase Inhibitor ◽

Induced Genes

In this study we investigated the effect of histone acetylation on the transcription of adrenergic-induced genes in rat pinealocytes. We found that treatment of pinealocytes with trichostatin A (TSA), a histone deacetylase inhibitor, caused hyperacetylation of histone H3 (H3) Lys14 at nanomolar concentrations. Hyperacetylation of H3 was also observed after treatment with scriptaid, a structurally unrelated histone deacetylase inhibitor. The effects of TSA and scriptaid were inhibitory on the adrenergic induction of arylalkylamine-n-acetyltransferase (aa-nat) mRNA, protein, and enzyme activity, and on melatonin production. TSA at higher concentrations also inhibited the adrenergic induction of mapk phosphatase-1 (mkp-1) and inducible cAMP early repressor mRNAs. In contrast, the effect of TSA on the norepinephrine induction of the c-fos mRNA was stimulatory. Moreover, the effect of TSA on adrenergic-induced gene transcription was dependent on the time of its addition; its effect was only observed during the active phase of transcription. Chromatin immunoprecipitation with antibodies against acetylated Lys14 of H3 showed an increase in DNA recovery of the promoter regions of aa-nat, mkp-1, and c-fos after treatment with TSA. Together, our results demonstrate that histone acetylation differentially influences the transcription of adrenergic-induced genes, an enhancing effect for c-fos but inhibitory for aa-nat, mkp-1, and inducible cAMP early repressor. Moreover, both inhibitory and enhancing effects appear to be mediated through specific modification of promoter-bound histones during gene transcription.

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Potentiation of androgen receptor transcriptional activity by inhibition of histone deacetylation--rescue of transcriptionally compromised mutants

Journal of Endocrinology ◽

10.1677/joe.0.1820377 ◽

2004 ◽

Vol 182 (3) ◽

pp. 377-389 ◽

Cited By ~ 33

Author(s):

CG Korkmaz ◽

K Fronsdal ◽

Y Zhang ◽

PI Lorenzo ◽

F Saatcioglu

Keyword(s):

Androgen Receptor ◽

Histone Acetylation ◽

Nuclear Receptor ◽

Transcriptional Activation ◽

Transcriptional Activity ◽

Trichostatin A ◽

Specific Antigen ◽

Histone Deacetylation ◽

Lncap Cells ◽

Viral Oncoprotein

Androgens are critical in the development and maintenance of the male reproductive system and important in the progression of prostate cancer. The effects of androgens are mediated by the androgen receptor (AR), which is a ligand-modulated transcription factor that belongs to the nuclear receptor superfamily. We and others have previously shown that CREB-binding protein (CBP) can function as a coactivator for AR. Similar to some other nuclear receptor coactivators and/or the proteins that they interact with, CBP has histone acetyl transferase (HAT) activity that is thought to contribute to transcriptional activation by nuclear receptors. We have therefore assessed whether an increase in the histone acetylation status in the cell can influence AR transcriptional activity, by using the histone deacetylase (HDAC) inhibitors (HDACIs) trichostatin A (TSA), sodium butyrate (Na-But) and depsipeptide (FR901228). We found that inhibition of HDAC activity significantly increased the ability of endogenous AR in LNCaP cells, or ectopically expressed AR in HeLa cells, to activate transcription from AR-dependent reporter constructs. In addition, HDACIs increased the androgen-dependent activation of the prostate-specific antigen (PSA) gene in LNCaP cells, an increase that was not due to an increase in nuclear AR protein levels. Moreover, the viral oncoprotein E1A that inhibits CBP HAT activity fully repressed the ability of HDACIs to stimulate AR-mediated transcription, indicating that CBP is involved in this process. Deletional mutagenesis of AR indicated that whereas the AF-2 domain in the C-terminus is dispensable, the AF-1 domain in the N-terminus is required for augmentation of AR action by HDACIs, an observation which is in concordance with the reduced ability of CBP to activate AR N-terminal deletion mutants. Furthermore, HDACI treatment rescued the deficiency in the transactivation potential of AF-2 mutants. Taken together, our findings suggest that a change in the level of histone acetylation of target genes is an important determinant of AR action, possibly mediated by CBP.

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134INCOMPLETE HISTONE ACETYLATION OF SOMATIC CHROMATIN IN BOVINE OOCYTES

Reproduction Fertility and Development ◽

10.1071/rdv16n1ab134 ◽

2004 ◽

Vol 16 (2) ◽

pp. 189

Author(s):

G. Wee ◽

S.-H. Kim ◽

K.P. Kim ◽

S. Yeo ◽

D.-B. Koo ◽

...

Keyword(s):

Gene Expression ◽

Somatic Cell ◽

Histone Acetylation ◽

Transcriptional Activation ◽

Linear Models ◽

Trichostatin A ◽

Specific Inhibitor ◽

Early Embryo ◽

Image Analyzer ◽

Cell Nuclei

Histone acetylation as an important regulatory mechanism of chromatin structure preceeding zygotic gene expression in early embryo development. After fertilization, transcriptional activation of the embryo begins during the S/G2 phase of the first cell cycle. However, the precise mechanism underlying activation of zygotic transcription remains to be understood, especially in bovine nuclear transfer (NT) embryos. It is known that acetylation of histone H4 lysine 5 (H4K5) represents hyperacetylation state, which is correlated with gene expression. In this study, the acetylation of H4K5 was observed during pronuclear formation by using immunofluorescence analysis with anti-AcH4K5. Our data were analyzed by the general linear models (GLM) procedure of the SAS. In IVF embryos, acetylation of H4K5 occurred on the paternal chromatin at 8h after fertilization but did not occur on the maternal chromatin until 10h after fertilization. Reconstructed oocytes with deactylated somatic cell nuclei began to show signs of acetylation on chromatin at 3h after fusion. When acetylation intensity was calculated using an image analyzer, IVF embryos presented a higher acetylation signal than NT embryos (P<0.05). To induce hyperacetylation in NT embryos, somatic cells were exposed to trichostatin A (TSA, 1μM for 60h), a specific inhibitor of histone deacetylase (HDAC), prior to NT. Acetylated signals of H4K5 increased significantly in TSA-treated cells as compared with non-treated cells (P<0.05). The reconstructed embryos with TSA-treated cells showed a higher fluorescence intensity than the oocytes with non-treated cells (P<0.05), but weak signals compared to IVF embryos. Thus, the results demonstrated low histone acetylation level of somatic cell nuclei after NT during the zygotic progress. Our findings suggest that developmental failures of NT embryos may be due to incomplete chromatin remodeling of somatic cell nuclei during early embryonic development.

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96 TRICHOSTATIN A IMPROVED THE QUALITY OF RABBIT NUCLEAR TRANSFER EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv19n1ab96 ◽

2007 ◽

Vol 19 (1) ◽

pp. 165 ◽

Cited By ~ 3

Author(s):

J. Xu ◽

L.-Y. Sung ◽

J. Zhang ◽

X. Tian ◽

Y. E. Chen ◽

...

Keyword(s):

Histone Acetylation ◽

Trichostatin A ◽

Donor Cell ◽

Cumulus Cells ◽

Cell Number ◽

Developmental Potential ◽

Number Of Factors ◽

Nuclear Transplant ◽

Blastocyst Quality ◽

And Control

Nuclear reprogramming is dependent upon a number of factors, including chromatin organization and modification. Trychostatin A (TSA), a histone deacetylase inhibitor, was used to increase histone acetylation and to improve reprogrammability in both cattle and mice. The objective of the study was to determine whether TSA could improve the pre-implantational development potential of rabbit nuclear transplant (NT) embryos. Rabbit oocytes were flushed from the oviducts of superovulated donors treated with the regime of FSH and hCG. Cumulus cells were then denuded from the oocytes by incubation in 0.5% hyaluronidase and pipetting. Oocyte enucleation was conducted in 10% FBS M199 and confirmed under fluorescence microscopy. Cumulus cells were prepared as nuclear donors for NT; a donor cell with the diameter approximately 15–19 µm was transferred into the perivitelline space of an enucleated oocyte, and subsequently fused with the oocyte recipient by application of 3 direct current pulses at 3.2 kV cm−1 for a duration of 20 µs/pulse. Fused embryos were activated by the same electrical stimulation regime described above, and subsequently cultured in M199 + 10% FBS containing 2.0 mM 6-dimethylaminopurine (DMAP) and 5 µg mL−1 cycloheximide for 1 h. Rabbit NT embryos were cultured in 5 nM TSA-2.5% FBS-B2 medium for 10 h before being transferred into regular medium (FBS-B2). The TSA-treated embryos (5 nM vs. 0 nM) were cultured in 400 µL FBS-B2 medium for 5 days in 5% CO2 in a humidified atmosphere at 38.5°C (initiation of activation = Day 0). NT embryo development to cleaved (2 to 4 cell), morula, and blastocyst stages was evaluated on Day 2, Day 3, and Day 5, respectively. The selected NT blastocysts were counted for cell numbers following Hoechst 33342 epifluorescenin staining. The results (Table 1) showed that there was no difference on pre-implantational development of cloned embryos between TSA-added and control groups (P > 0.05). However, a significantly higher cell number per NT blastocyst was found in the TSA-added group (357 vs. 113; P < 0.05). This indicated that the blastocyst quality in NT embryos was improved with the addition of TSA by increasing histone acetylation activity. The developmental potential of TSA-treated NT embryos to term is under investigation. Table 1.Effects of TSA on the pre-implantational development of cloned rabbit embryos This work was supported by NIH/NCRR-SBIR grant: 1R43RR020261-01.

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