Changes in histone acetylation during mouse oocyte meiosis.
ABSTRACT: We examined global changes in the acetylation of histones in mouse oocytes during meiosis. Immunocytochemistry with specific antibodies against various acetylated lysine residues on histones H3 and H4 showed that acetylation of all the lysines decreased to undetectable or negligible levels in the oocytes during meiosis, whereas most of these lysines were acetylated during mitosis in preimplantation embryos and somatic cells. When the somatic cell nuclei were transferred into enucleated oocytes, the acetylation of lysines decreased markedly. This type of deacetylation was inhibited by trichostatin A, a specific inhibitor of histone deacetylase (HDAC), thereby indicating that HDAC is able to deacetylate histones during meiosis but not during mitosis. Meiosis-specific deacetylation may be a consequence of the accessibility of HDAC1 to the chromosome, because HDAC1 colocalized with the chromosome during meiosis but not during mitosis. As histone acetylation is thought to play a role in propagating the gene expression pattern to the descendent generation during mitosis, and the gene expression pattern of differentiated oocytes is reprogrammed during meiosis to allow the initiation of a new program by totipotent zygotes of the next generation, our results suggest that the oocyte cytoplasm initializes a program of gene expression by deacetylating histones.
Project description:Histone deacetylase 1 (HDAC1) is an epigenetic enzyme that regulates key cellular processes, such as cell proliferation, apoptosis, and cell survival, by deacetylating histone substrates. Aberrant expression of HDAC1 is implicated in multiple diseases, including cancer. As a consequence, HDAC inhibitors have emerged as effective anti-cancer drugs. HDAC inhibitor-induced G0/G1 cell-cycle arrest has been attributed to epigenetic transcriptional changes mediated by histone acetylation. However, the mechanism of G2/M arrest remains poorly understood. Here, we identified mitosis-related protein Eg5 (KIF11) as an HDAC1 substrate using a trapping mutant strategy. HDAC1 colocalized with Eg5 during mitosis and influenced the ATPase activity of Eg5. Importantly, an HDAC1- and HDAC2-selective inhibitor caused mitotic arrest and monopolar spindle formation, consistent with a model in which Eg5 deacetylation by HDAC1 is critical for mitotic progression. These findings revealed a previously unknown mechanism of action of HDAC inhibitors involving Eg5 acetylation, and provide a compelling mechanistic hypothesis for HDAC inhibitor-mediated G2/M arrest.
Project description:Changes in histone acetylation occur during oocyte development and maturation, but the role of specific histone deacetylases in these processes is poorly defined. We report here that mice harboring Hdac1(-/+)/Hdac2(-/-) or Hdac2(-/-) oocytes are infertile or sub-fertile, respectively. Depleting maternal HDAC2 results in hyperacetylation of H4K16 as determined by immunocytochemistry--normal deacetylation of other lysine residues of histone H3 or H4 is observed--and defective chromosome condensation and segregation during oocyte maturation occurs in a sub-population of oocytes. The resulting increased incidence of aneuploidy likely accounts for the observed sub-fertility of mice harboring Hdac2(-/-) oocytes. The infertility of mice harboring Hdac1(-/+)/Hdac2(-/-)oocytes is attributed to failure of those few eggs that properly mature to metaphase II to initiate DNA replication following fertilization. The increased amount of acetylated H4K16 likely impairs kinetochore function in oocytes lacking HDAC2 because kinetochores in mutant oocytes are less able to form cold-stable microtubule attachments and less CENP-A is located at the centromere. These results implicate HDAC2 as the major HDAC that regulates global histone acetylation during oocyte development and, furthermore, suggest HDAC2 is largely responsible for the deacetylation of H4K16 during maturation. In addition, the results provide additional support that histone deacetylation that occurs during oocyte maturation is critical for proper chromosome segregation.
Project description:Errors in meiotic chromosome segregation are the leading cause of spontaneous abortions and birth defects. Almost all such aneuploidy derives from meiotic errors in females, with increasing maternal age representing a major risk factor. It was recently reported that histones are globally deacetylated in mammalian oocytes during meiosis but not mitosis. In the present study, inhibition of meiotic histone deacetylation was found to induce aneuploidy in fertilized mouse oocytes, which resulted in embryonic death in utero at an early stage of development. In addition, a histone remained acetylated in the oocytes of older (10-month-old) female mice, suggesting that the function for histone deacetylation is decreased in the oocytes of such mice. Thus, histone deacetylation may be involved in the fair distribution of chromosomes during meiotic division. The high incidence of aneuploidy in the embryos of older females may be due to inadequate meiotic histone deacetylation.
Project description:Posttranslational modifications, including acetylation and deacetylation of histones and other proteins, modulate hormone action. In Tribolium castaneum TcA cells, Trichostatin A, a histone deacetylase (HDAC) inhibitor, mimics juvenile hormone (JH) in inducing JH response genes (e.g., Kr-h1), suggesting that HDACs may be involved in JH action. To test this hypothesis, we identified genes coding for HDACs in T. castaneum and studied their function. Knockdown of 12 HDAC genes showed variable phenotypes; the most severe phenotype was detected in insects injected with double-stranded RNA targeting HDAC1 (dsHDAC1). The dsHDAC1-injected insects showed arrested growth and development and eventually died. Application of JH analogs hydroprene to T. castaneum larvae and JH III to TcA cells suppressed HDAC1 expression. Sequencing of RNA isolated from control and dsHDAC1-injected larvae identified 1,720 differentially expressed genes, of which 1,664 were up-regulated in dsHDAC1-treated insects. The acetylation levels of core histones were increased in TcA cells exposed to dsHDAC1 or JH III. ChIP assays performed using histone H2BK5ac antibodies showed an increase in acetylation in the Kr-h1 promoter region of cells exposed to JH III or dsHDAC1. Overexpression or knockdown of HDAC1, SIN3, or both resulted in a decrease or increase in Kr-h1 mRNA levels and its promoter activity, respectively. Overexpression of the JH receptor Methoprene tolerant (Met) was unable to induce Kr-h1 in the presence of HDAC1 or SIN3. These data suggest that epigenetic modifications influence JH action by modulating acetylation levels of histones and by affecting the recruitment of proteins involved in the regulation of JH response genes.
Project description:In embryonic stem cells (ESCs), Wnt-responsive development-related genes are silenced to maintain pluripotency and their expression is activated during differentiation. Acetylation of histones by histone acetyltransferases stimulates transcription, whereas deacetylation of histones by HDACs is correlated with transcriptional repression. Although Wnt-mediated gene transcription has been intimately linked to the acetylation or deacetylation of histones, how Wnt signaling regulates this type of histone modification is poorly understood. Here, we report that Smek, a regulatory subunit of protein phosphatase 4 (PP4) complex, plays an important role in histone deacetylation and silencing of the Wnt-responsive gene, brachyury, in ESCs. Smek mediates recruitment of PP4c and HDAC1 to the Tcf/Lef binding site of the brachyury promoter and inhibits brachyury expression in ESCs. Activation of Wnt signaling during differentiation causes disruption of the Smek/PP4c/HDAC1 complex, resulting in an increase in histones H3 and H4 acetylation at the brachyury gene locus. These results suggest that the Smek-containing PP4 complex represses transcription of Wnt-responsive development-related genes through histone deacetylation, and that this complex is essential for ESC pluripotency maintenance.
Project description:Illicit drugs are known to affect central nervous system (CNS). Majorly psychostimulants such as cocaine, methamphetamine (METH) and opioids such as morphine are known to induce epigenetic changes of histone modifications and chromatin remodeling which are mediated by histone acetyltransferase (HAT) and histone deacetylase (HDAC). Aberrant changes in histone acetylation-deacetylation process further exacerbate dysregulation of gene expression and protein modification which has been linked with neuronal impairments including memory formation and synaptic plasticity. In CNS, astrocytes play a pivotal role in cellular homeostasis. However, the impact of psychostimulants and opioid mediated epigenetic changes of HAT/HADCs in astrocytes has not yet been fully elucidated. Therefore, we have investigated the effects of the psychostimulants and opioid on the acetylation-regulating enzymes- HAT and HDACs role in astrocytes. In this study, Class I and II HDACs and HATs gene expression, protein changes and global level changes of acetylation of H3 histones at specific lysines were analyzed. In addition, we have explored the neuroprotective "nootropic" drug piracetam were exposed with or without psychostimulants and opioid in the human primary astrocytes. Results revealed that psychostimulants and opioid upregulated HDAC1, HDAC4 and p300 expression, while HDAC5 and GCN5 expression were downregulated. These effects were reversed by piracetam coexposure. Psychostimulants and opioid exposure upregulated global acetylation levels of all H3Ks, except H3K14. These results suggest that psychostimulants and opioids differentially influence HATs and HDACs.
Project description:HDAC11, the sole member of HDAC class IV family, plays vital roles in activating mitosis and apoptosis of tumor cells, but its functions in meiosis are rarely investigated. In the present study, the effect of HDAC11 on meiosis during porcine oocytes maturation was fully studied. The results showed that HDAC11 inhibition by its specific inhibitor JB-3-22 dramatically decreased the porcine oocyte maturation rate by disturbing spindle organization and chromosomes alignment without affecting the cytoplasmic maturation. Further study indicated that HDAC11 inhibition significantly elevated the acetylation levels of α-tubulin and H4K16, which are crucial for spindle organization and chromosomes alignment. Moreover, immunofluorescence staining results showed that HDAC11 inhibition also disturbed other meiosis-related histone modifications, such as increased H3S10pho, H4K5ac and H4K12ac levels and reduced H3T3pho level. Furthermore, RNA-seq analysis results indicated that HDAC11 inhibition disturbed porcine oocytes transcriptome (157 up-regulation, 106 down-regulation). In addition, HDAC11 inhibition compromised oocytes quality and subsequent development after parthenogenetic activation, which may be caused by the aberrant nuclear maturation and transcriptome expression profile during oocytes maturation. Therefore, our results elucidate the function of HDAC11 in porcine oocytes maturation and embryos development through regulating α-tubulin acetylation, meiosis-related histone modifications and transcriptome.
Project description:Mitosis is largely driven by posttranslational modifications of proteins. Recent studies suggest that protein acetylation is prevalent in mitosis, but how protein acetylation/deacetylation regulates mitotic progression remains unclear. Nuclear distribution protein C (NudC), a conserved protein that regulates cell division, was previously shown to be acetylated. We found that NudC acetylation was decreased during mitosis. Using mass spectrometry analysis, we identified K39 to be an acetylation site on NudC. Reconstitution of NudC-deficient cells with wild-type or K39R acetylation-defective NudC rescued mitotic phenotypes, including chromosome misalignment, chromosome missegregation, and reduced spindle width, observed after NudC protein knockdown. In contrast, the K39Q acetylation-mimetic NudC was unable to rescue these mitotic phenotypes, suggesting that NudC deacetylation is important for mitotic progression. To examine proteins that may play a role in NudC deacetylation during mitosis, we found that NudC co-localizes on the mitotic spindle with the histone deacetylase HDAC3, an HDAC shown to regulate mitotic spindle stability. Further, NudC co-immunoprecipitates with HDAC3 and loss of function of HDAC3 either by protein knockdown or inhibition with a small molecule inhibitor increased NudC acetylation. These observations suggest that HDAC3 may be involved in NudC deacetylation during mitosis. Cells with NudC or HDAC3 knockdown exhibited overlapping mitotic abnormalities, including chromosomes arranged in a "dome-like" configuration surrounding a collapsed mitotic spindle. Our studies suggest that NudC acetylation/deacetylation regulates mitotic progression and NudC deacetylation, likely through HDAC3, is critical for spindle function and chromosome congression.
Project description:Acetylation and deacetylation of histones and other proteins depend on the opposing activities of histone acetyltransferases and histone deacetylases (HDACs), leading to either positive or negative gene expression changes. The use of HDAC inhibitors (HDACi) has uncovered a role for HDACs in the control of proliferation, apoptosis and inflammation. However, little is known of the roles of specific HDACs in intestinal epithelial cells (IEC). We investigated the consequences of ablating both Hdac1 and Hdac2 in murine IECs gene expression. HDAC1 and HDAC2 conditionally mutated mice were provided by Dr EN Olson (University of Texas Southwestern Medical Center, Dallas, TX) (Montgomery et al., 2007). Floxed HDAC1 and HDAC2 mice were crossed with villin-Cre transgenic mice to insure specific intestinal epithelial cell gene deletion (Madison et al., 2002). Total RNAs from the colon of three control and three HDAC1/2 IEC-specific knockout mice were isolated with the Rneasy kit (Qiagen, Mississauga, ON, Canada).
Project description:Lymphoid specific helicase (Lsh) belongs to the family of SNF2/helicases. Disruption of Lsh leads to developmental growth retardation and premature aging in mice. However, the specific effect of Lsh on human cellular senescence remains unknown. Herein, we report that Lsh overexpression delays cell senescence by silencing p16(INK4a) in human fibroblasts. The patterns of p16(INK4a) and Lsh expression during cell senescence present the inverse correlation. We also find that Lsh requires histone deacetylase (HDAC) activity to repress p16(INK4a) and treatment with trichostatin A (TSA) is sufficient to block the repressor effect of Lsh. Moreover, overexpression of Lsh is correlated with deacetylation of histone H3 at the p16 promoter, and TSA treatment in Lsh-expressing cells reverses the acetylation status of histones. Additionally, we demonstrate an interaction between Lsh, histone deacetylase 1 (HDAC1) and HDAC2 in vivo. Furthermore, we demonstrate that Lsh interacts in vivo with the p16 promoter and recruits HDAC1. Our data suggest that Lsh represses endogenous p16(INK4a) expression by recruiting HDAC to establish a repressive chromatin structure at the p16(INK4a) promoter, which in turn delays cell senescence.