Haoli Hou

Epigenetic Changes are Associated with Programmed Cell Death Induced by Heat Stress in Seedling Leaves of Zea mays

Histone modification plays a crucial role in regulation of chromatin architecture and function, responding to adverse external stimuli. However, little is known about a possible relationship between epigenetic modification and programmed cell death (PCD) in response to environmental stress. Here, we found that heat stress induced PCD in maize seedling leaves which was characterized by chromatin DNA laddering and DNA strand breaks detected by a terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) test. The activities of the reactive oxygen species (ROS)-related enzymes superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were progressively increased over time in the heat-treated seedlings. However, the concentration of H2O2 remained at relatively lower levels, while the concentration of superoxide anion ([Formula: see text]) was increased, accompanied by the occurrence of higher ion leakage rates after heat treatment. The total acetylation levels of histones H3K9, H4K5 and H3 were significantly increased, whereas the di-methylation level of histone H3K4 was unchanged and the di-methylation level of histone H3K9 was decreased in the seedling leaves exposed to heat stress compared with the control seedlings, accompanied by increased nucleolus size indicative of chromatin decondensation. Furthermore, treatment of seedlings with trichostatin A (TSA), which always results in genomic histone hyperacetylation, caused an increase in the [Formula: see text] level within the cells. The results suggested that heat stress persistently induced [Formula: see text], leading to PCD in association with histone modification changes in the maize leaves.

Transcriptional Regulation of Cell Cycle Genes in Response to Abiotic Stresses Correlates with Dynamic Changes in Histone Modifications in Maize

The histone modification level has been shown to be related with gene activation and repression in stress-responsive process, but there is little information on the relationship between histone modification and cell cycle gene expression responsive to environmental cues. In this study, the function of histone modifications in mediating the transcriptional regulation of cell cycle genes under various types of stress was investigated in maize (Zea mays L.). Abiotic stresses all inhibit the growth of maize seedlings, and induce total acetylation level increase compared with the control group in maize roots. The positive and negative regulation of the expression of some cell cycle genes leads to perturbation of cell cycle progression in response to abiotic stresses. Chromatin immunoprecipitation analysis reveals that dynamic histone acetylation change in the promoter region of cell cycle genes is involved in the control of gene expression in response to external stress and different cell cycle genes have their own characteristic patterns for histone acetylation. The data also showed that the combinations of hyperacetylation and hypoacetylation states of specific lysine sites on the H3 and H4 tails on the promoter regions of cell cycle genes regulate specific cell cycle gene expression under abiotic stress conditions, thus resulting in prolonged cell cycle duration and an inhibitory effect on growth and development in maize seedlings.

Zinc oxide nanoparticles-induced epigenetic change and G2/M arrest are associated with apoptosis in human epidermal keratinocytes

As an engineered nanomaterial, zinc oxide nanoparticles (ZnO NPs) are used frequently in biological applications and can make contact with human skin. Here, we systematically investigated the effects of ZnO NPs on non-tumorigenic human epidermal keratinocytes, which were used as a test model for this in vitro study, at the epigenetic and molecular levels. Our results showed that ZnO NPs induced cell cycle arrest at the G2/M checkpoint before the viability of human epidermal keratinocytes was reduced, which was associated with the chromatin changes at the epigenetic level, including increased methylation of histone H3K9 and decreased acetylation of histone H4K5 accompanied by chromatin condensation at 24 hours. The mRNA expression of the methyltransferase genes G9a and GLP was also increased upon treatment with ZnO NPs, and the acetyltransferase genes GCN5, P300, and CBP were downregulated. Reactive oxygen species were found to be more abundant after treatment with ZnO NPs for 6 hours, and DNA damage was observed at 24 hours. Transmission electron microscopy and flow cytometry confirmed that ZnO NPs were absorbed into the cell when they were added to the medium. Apoptotic human epidermal keratinocytes were detected, and the expression of the proapoptotic genes Bax, Noxa, and Puma increased significantly, while the expression of the antiapoptotic gene Bcl-xl decreased 24 hours after exposure to ZnO NPs. These findings suggest that the ZnO NPs induced cell cycle arrest at G2/M, which was associated with epigenetic changes and accompanied by p53-Bax mitochondrial pathway-mediated apoptosis.

Promoter-associated histone acetylation is involved in the osmotic stress-induced transcriptional regulation of the maize ZmDREB2A gene

Epigenetic modifications play a key role in the transcriptional regulation of stress-induced gene expression in plants. In this study, we showed that the overall acetylation levels of histone H3 lysine 9 (H3K9) and H4 lysine 5 (H4K5) in the genome were increased in maize seedlings after mannitol treatment (to mimic osmotic stress). Mannitol treatment significantly induced the upregulation of the maize osmotic stress responsive gene Zea mays dehydration-responsive element binding protein 2A (ZmDREB2A), whereas abscisic acid (ABA) did not result in the induction of this gene. The application of exogenous ABA under osmotic stress conditions strongly repressed the induction of the ZmDREB2A gene. Chromatin immunoprecipitation and chromatin accessibility by real-time PCR experiments revealed that the promoter region of the ZmDREB2A gene was quickly hyperacetylated and decondensed after the mannitol treatment, suggesting that the promoter region is poised for histone acetylation to allow for fast induction of the ZmDREB2A gene. However, under osmotic stress conditions, the ABA treatment decreased the acetylation status and chromatin accessibility to micrococcal nuclease. These results suggest that osmotic stress activates the transcription of the ZmDREB2A gene by increasing the levels of acetylated histones H3K9 and H4K5 associated with the ZmDREB2A promoter region.

Histone Acetylation is Involved in Gibberellin-Regulated sodCp Gene Expression in Maize Aleurone Layers.

The cereal aleurone layer plays an important role in seed germination, and reactive oxygen species (ROS) in aleurone layers act as crucial signal molecules in this progression. Recent studies have revealed that epigenetic modification is involved in plant development and seed germination. However, little is known about a possible relationship between histone modification and the ROS signaling pathway in cereal aleurone layers during seed germination. Here, we found that the expression of both histone acetyltransferases (HATs) and histone deacetylases (HDACs) was increased gradually during seed germination, accompanied by an increase in global acetylation levels of histones H3 and H4 in maize aleurone layers. The acetylation was found to be promoted by GA(3) and suppressed by ABA. However, when the HDAC inhibitor trichostatin A (TSA) was used, the increased H3K9ac and H4K5ac level correlated with an inhibition of the germination. These results indicated that the overall histone acetylation in the aleurone layers is not required for germination. Similarly these two hormones, GA(3) and ABA, exerted opposed effects on the expression of the ROS-related gene sodCp. Furthermore, chromatin immunoprecipitation experiments showed that the promoter region of the sodCp gene was hyperacetylated during germination, and this acetylation was promoted by GA(3) and inhibited by both ABA and TSA. These results suggested that GA(3)-mediated expression of the sodCp gene in aleurone layers is associated with histone hyperacetylation on the promoter and coding region of this gene, consequently leading to an accumulation of H(2)O(2) which regulated production of α-amylase during seed germination.

Single-wall and multi-wall carbon nanotubes promote rice root growth by eliciting the similar molecular pathways and epigenetic regulation.

Organisms are constantly exposed to environmental stimuli and have evolved mechanisms of protection and adaptation. Various effects of nanoparticles (NPs) on crops have been described and some results confirm that NPs could enhance plant growth at the physiological and genetic levels. This study comparatively analysed the effect of carbon nanotubes (CNTs) on rice growth. The results showed that single-wall CNTs were located in the intercellular space while multi-wall CNTs penetrated cell walls in roots. CNTs could promote rice root growth through the regulation of expression of the root growth related genes and elevated global histone acetylation in rice root meristem zones. These responses were returned to normal levels after CNTs were removed from medium. CNTs caused the similar histone acetylation and methylation statuses across the local promoter region of the Cullin-RING ligases 1 (CRL1) gene and increased micrococcal nuclease accessibility of this region, which enhanced this gene expression. The authors results suggested that CNTs could cause plant responses at the cellular, genetic, and epigenetic levels and these responses were independent on interaction modes between root cells and CNTs.

Histone acetylation and reactive oxygen species are involved in the preprophase arrest induced by sodium butyrate in maize roots.

Histone acetylation plays a critical role in controlling chromatin structure, and reactive oxygen species (ROS) are involved in cell cycle progression. To study the relationship between histone acetylation and cell cycle progression in plants, sodium butyrate (NaB), a histone deacetylase (HDAC) inhibitor that can cause a significant increase in histone acetylation in both mammal and plant genomes, was applied to treat maize seedlings. The results showed that NaB had significant inhibition effects on different root zones at the tissue level and caused cell cycle arrest at preprophase in the root meristem zones. This effect was accompanied by a dramatic increase in the total level of acetylated lysine 9 on histone H3 (H3K9ac) and acetylated lysine 5 on histone H4 (H4K5ac). The exposure of maize roots in NaB led to a continuous rise of intracellular ROS concentration, accompanied by a higher electrolyte leakage ratio and malondialdehyde (MDA) relative value. The NaB-treated group displayed negative results in both TdT-mediated dUTP nick end labelling (TUNEL) and γ-H2AX immunostaining assays. The expression of topoisomerase genes was reduced after treatment with NaB. These results suggested that NaB increased the levels of H3K9ac and H4K5ac and could cause preprophase arrest accompanied with ROS formation leading to the inhibition of DNA topoisomerase.

Overexpression of Ribosomal RNA in the Development of Human Cervical Cancer Is Associated with rDNA Promoter Hypomethylation.

The ribosomal RNA (rRNA) gene encodes rRNA for protein synthesis. Aberrant expression of the rRNA gene has been generally observed in tumor cells and levels of its promoter methylation as an epigenetic regulator affect rRNA gene transcription. The possible relationship between expression and promoter methylation of rDNA has not been examined in human clinical cervical cancer. Here we investigate rRNA gene expression by quantitative real time PCR, and promoter methylation levels by HpaII/MspI digestion and sodium bisulfite sequencing in the development of human cervical cancer. We find that indeed rRNA levels are elevated in most of cervical intraepithelial neoplasia (CIN) specimens as compared with non-cancer tissues. The rDNA promoter region in cervical intraepithelial neoplasia (CIN) tissues reveals significant hypomethylation at cytosines in the context of CpG dinucleotides, accompanied with rDNA chromatin decondensation. Furthermore treatment of HeLa cells with the methylation inhibitor drug 5-aza-2’-deoxycytidine (DAC) demonstrates the negative correlation between the expression of 45S rDNA and the methylation level in the rDNA promoter region. These data suggest that a decrease in rDNA promoter methylation levels can result in an increase of rRNA synthesis in the development of human cervical cancer.

Histone acetylation is involved in GA-mediated 45S rDNA decondensation in maize aleurone layers

Key messageThe aleurone layer is crucial to seed germination. Using dissected aleurone layers, we found that GA increased histone acetylation accompanied by rDNA decondensation in aleurone layers during maize seed germination.AbstractAleurone layers play an important role in cereal seed germination. In this study, we reported that rDNA chromatin was decondensed, accompanied with increased rDNA expression and genomic global hyperacetylation in gibberellin (GA)-treated maize-dissected aleurone layers. The activity analysis of histone acetyltransferase (HAT) and deacetylase (HDAC) showed that GA increased the level of histone acetylation by promoting the ratio of HAT/HDAC activity in aleurone layers. HDAC inhibitors TSA and CUDC-101 elevated the histone acetylation in aleurone layers accompanied by 45S rDNA decondensation. The further chromatin immunoprecipitation experiments showed that GA treatment promoted the level of histone acetylation in the promoter region of the rRNA and HAT/HDAC genes in aleurone layers. Taken together, these data indicated that histone acetylation mediates GA-regulated 45S rDNA chromatin decondensation in aleurone layers during maize seed germination.

Histone Modification Is Involved in Okadaic Acid (OA) Induced DNA Damage Response and G2-M Transition Arrest in Maize

Histone modifications are involved in regulation of chromatin structure. To investigate the relationship between chromatin modification and cell cycle regulation during plant cell proliferation, Okadaic acid (OA), a specific inhibitor of serine/threonine protein phosphatase, was applied in this study. The results showed that OA caused the cell cycle arrest at preprophase, leading to seedling growth inhibition. Western blotting assay revealed that the spatial distribution of phosphorylation of Ser10 histone H3 tails (H3S10ph) signals was altered under OA treatment. Reactive oxygen species (ROS) was found to be at higher levels and TdT-mediated dUTP nick end labeling (TUNEL) assay displayed DNA breaks happened at the chromatin after treatment with OA, companied with an increase in the acetylation of histone H4 at lysine 5 (H4K5ac) level. From these observations, we speculated that the alteration of the spatial distribution of H3S10ph and the level of H4K5ac was involved in the procedure that OA induced DNA breaks and G2-M arrested by the accumulation of ROS, and that the histone H3S10ph and H4K5ac might facilitate DNA repair by their association with the chromatin decondensation.