Resveratrol protects intestinal epithelial cells against radiation-induced damage by promoting autophagy and inhibiting apoptosis through SIRT1 activation.

Intrinsic autophagy is important for the maintenance of intestinal homeostasis and intestinal regeneration. Ionizing radiation suppresses intrinsic autophagy and reduces damage-induced regeneration in the intestine, resulting in intestinal injury. Resveratrol, a sirtuin 1 (SIRT1) agonist, promotes autophagy and exerts radioprotective effect. In this study, the protective effect of resveratrol against radiation-induced intestinal injury and its potential mechanism were investigated. Intestinal epithelial cells (IEC-6) were exposed to 10 Gy ionizing radiation and resveratrol (0.1-40.0 µM). Cell viability was investigated using Cell Counting Kit 8 (CCK8), apoptosis was observed by Annexin V-fluorescein isothiocyanate/propidium iodide (PI) staining and flow cytometry, and the expression of apoptotic and autophagic proteins was determined by western blotting. Resveratrol exerted a high toxicity against IEC-6 cells, but at low concentrations, it inhibited ionizing radiation-induced apoptosis. Resveratrol increased SIRT1 expression after irradiation and inhibited ionizing radiation-induced p53 acetylation and pro-apoptotic protein, Bax, expression. Furthermore, resveratrol promoted autophagy via the phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway, thereby protecting IEC-6 cells against radiation-induced damage. These results suggest that resveratrol reduces radiation-induced IEC-6 cell damage by inhibiting apoptosis and promoting autophagy via the activation of SIRT1, and that the PI3K/AKT/mTOR signaling pathway is involved in the induction of autophagy.

Increased Histone Acetylation and Decreased Expression of Specific Histone Deacetylases in Ultraviolet-Irradiated and Intrinsically Aged Human Skin In Vivo.

Histone deacetylases (HDACs) are conserved enzymes that remove acetyl groups from lysine side chains in histones and other proteins and play a crucial role in epigenetic regulation. Previously, we showed that histone acetylation is implicated in ultraviolet (UV)-induced inflammation and matrix impairment. To elucidate the histone acetylation status and specific HDACs involved in skin aging, we examined the changes in histone acetylation, global HDAC activity, and the expression of HDACs and sirtuins (SIRTs) in intrinsically aged and photoaged human skin as well as in UV-irradiated human skin in vivo. Following acute UV irradiation, the acetylated histone H3 (AcH3) level was increased, but HDAC activity and the expression levels of HDAC4, HDAC11, and SIRT4 were significantly decreased. In intrinsically aged skin, AcH3 levels were increased, but HDAC activity and the expression levels of HDAC4, HDAC5, HDAC10, HDAC11, SIRT6, and SIRT7 were significantly decreased. However, histone acetylation and HDAC expression in photoaged skin were not significantly different from those in intrinsically aged skin. Collectively, HDAC4 and HDAC11 were decreased in both UV-irradiated and intrinsically aged skin, suggesting that they may play a universal role in increased histone acetylation associated with skin aging.

SIRT7-dependent deacetylation of NPM promotes p53 stabilization following UV-induced genotoxic stress.

Adaptation to different forms of environmental stress is crucial for maintaining essential cellular functions and survival. The nucleolus plays a decisive role as a signaling hub for coordinating cellular responses to various extrinsic and intrinsic cues. p53 levels are normally kept low in unstressed cells, mainly due to E3 ubiquitin ligase MDM2-mediated degradation. Under stress, nucleophosmin (NPM) relocates from the nucleolus to the nucleoplasm and binds MDM2, thereby preventing degradation of p53 and allowing cell-cycle arrest and DNA repair. Here, we demonstrate that the mammalian sirtuin SIRT7 is an essential component for the regulation of p53 stability during stress responses induced by ultraviolet (UV) irradiation. The catalytic activity of SIRT7 is substantially increased upon UV irradiation through ataxia telangiectasia mutated and Rad3 related (ATR)-mediated phosphorylation, which promotes efficient deacetylation of the SIRT7 target NPM. Deacetylation is required for stress-dependent relocation of NPM into the nucleoplasm and MDM2 binding, thereby preventing ubiquitination and degradation of p53. In the absence of SIRT7, stress-dependent stabilization of p53 is abrogated, both in vitro and in vivo, impairing cellular stress responses. The study uncovers an essential SIRT7-dependent mechanism for stabilization of the tumor suppressor p53 in response to genotoxic stress.

Light-Driven Activation of RNA-Guided Nucleic Acid Cleavage.

As one of the most favorable stimuli, photoactivation provides an advantageous way to manipulate biological objects. In the current study, we have successfully demonstrated the use of light activation guide RNA (gRNA) strategy for controlling CRISPR systems. By conjugating photolabile protecting groups, the CRISPR functions became minimal, but exposure of acylated gRNAs to 365 nm light triggers the removal of masking groups, leading to the rescue of CRISPR functions. Furthermore, our strategy has been successfully used to control gene editing in human cells. This proof-of-concept study therefore demonstrates the promising potential of our strategy to versatile applications in chemical biology.

Photobiomodulation therapy improves human dental pulp stem cell viability and migration in vitro associated to upregulation of histone acetylation.

This in vitro study evaluated the role of photobiomodulation therapy (PBMT) on viability and migration of human dental pulp stem cells (hDPSCs) and its association to epigenetic mechanisms such as histone acetylation. The hDPSCs were characterized and assigned into control and PBMT groups. For the PBMT, five laser irradiations at 6-h intervals were performed using a continuous-wave InGaAlP diode laser. Viability (MTT), migration (scratch), and histone acetylation H3 (H3K9ac immunofluorescence) were evaluated immediately after the last irradiation. PBMT significantly increased the viability (P = 0.004). Also, PBMT group showed significantly increased migration of cells in the wound compared to the control in 6 h (P = 0.002), 12 h (P = 0.014) and 18 h (P = 0.083) being faster than the control, which only finished the process at 24 h. PBMT induced epigenetic modifications in hDPSC due to increased histone acetylation (P = 0.001). PBMT increased viability and migration of hDPSCs, which are related with the upregulation of histone acetylation and could be considered a promising adjuvant therapy for regenerative endodontic treatment.

The Histone Deacetylase Inhibitor Romidepsin Spares Normal Tissues While Acting as an Effective Radiosensitizer in Bladder Tumors in Vivo.

PURPOSE: Muscle-invasive bladder cancer has a 40% to 60% 5-year survival rate with radical treatment by surgical removal of the bladder or radiation therapy-based bladder preservation techniques, including concurrent chemoradiation. Elderly patients cannot tolerate current chemoradiation therapy regimens and often receive only radiation therapy, which is less effective. We urgently need effective chemotherapy agents for use with radiation therapy combinations that are nontoxic to normal tissues and tolerated by elderly patients. METHODS AND MATERIALS: We have identified histone deacetylase (HDAC) inhibitors as promising agents to study. Pan-HDAC inhibition, using panobinostat, is a good strategy for radiosensitization, but more selective agents may be more useful radiosensitizers in a clinical setting, resulting in fewer systemic side effects. Herein, we study the HDAC class I-selective agent romidepsin, which we predict to have fewer off-target effects than panobinostat while maintaining an effective level of tumor radiosensitization. RESULTS: In vitro effects of romidepsin were assessed by clonogenic assay and showed that romidepsin was effective in the nanomolar range in different bladder cancer cells and radiosensitized these cells. The radiosensitizing effect of romidepsin was confirmed in vivo using superficial xenografts. The drug/irradiation combination treatment resulted in significant tumor growth delay but did not increase the severity of acute (3.75 days) intestinal normal tissue toxicity or late toxicity at 29 weeks. Moreover, we showed that romidepsin treatment impaired both homologous recombination and nonhomologous end joining DNA repair pathways, suggesting that the disruption of DNA repair pathways caused by romidepsin is a key mechanism for its radiosensitizing effect in bladder cancer cells. CONCLUSIONS: This study demonstrates that romidepsin is an effective radiosensitizer in vitro and in vivo and does not increase the acute and late toxicity after ionizing radiation. Romidepsin is already in clinical use for the cutaneous T-cell lymphoma, but a phase 1 clinical trial of romidepsin as a radiosensitizer could be considered in muscle-invasive bladder cancer.

Enhancement of chitin suspension hydrolysis by a combination of ultrasound and chitinase.

This study evaluated the degradation kinetics and structural characteristics of chitin suspension (CS) with a combination of ultrasound and chitinase. Compared with the enzymolysis, the degradation degree of sonoenzymolysis was enhanced to the maximum by 27.93 % at an intensity of 25 W/mL for 20 min. According to degradation kinetics, ultrasound intensified molecular collision rate between chitinase and substrate, thereby increasing the degradation degree. What's more, combined with chitinase, ultrasound intensified the rate of the breakage of glycosidic bond and viscosity-average molecular weight (Mv) decrease, but no obvious change in acetylation degree (DA). Additionally, the intra- or inter-hydrogen bindings were weakened by ultrasound during sonoenzymolysis, leading to a slight decrease in crystalline index and a more ordered structure, which increased the accessibility of the substrate to enzyme. In conclusion, combination of chitinase and ultrasound could enhance the hydrolysis of CS while without changing its primary structure.

Elevated HDAC activity and altered histone phospho-acetylation confer acquired radio-resistant phenotype to breast cancer cells.

BACKGROUND: Poor-responsiveness of tumors to radiotherapy is a major clinical problem. Owing to the dynamic nature of the epigenome, the identification and targeting of potential epigenetic modifiers may be helpful to curb radio-resistance. This requires a detailed exploration of the epigenetic changes that occur during the acquirement of radio-resistance. Such an understanding can be applied for effective utilization of treatment adjuncts to enhance the efficacy of radiotherapy and reduce the incidence of tumor recurrence. RESULTS: This study explored the epigenetic alterations that occur during the acquirement of radio-resistance. Sequential irradiation of MCF7 breast cancer cell line up to 20 Gy generated a radio-resistant model. Micrococcal nuclease digestion demonstrated the presence of compact chromatin architecture coupled with decreased levels of histone PTMs H3K9ac, H3K27 ac, and H3S10pK14ac in the G0/G1 and mitotic cell cycle phases of the radio-resistant cells. Further investigation revealed that the radio-resistant population possessed high HDAC and low HAT activity, thus making them suitable candidates for HDAC inhibitor-based radio-sensitization. Treatment of radio-resistant cells with HDAC inhibitor valproic acid led to the retention of γH2AX and decreased H3S10p after irradiation. Additionally, an analysis of 38 human patient samples obtained from 8 different tumor types showed variable tumor HDAC activity, thus demonstrating inter-tumoral epigenetic heterogeneity in a patient population. CONCLUSION: The study revealed that an imbalance of HAT and HDAC activities led to the loss of site-specific histone acetylation and chromatin compaction as breast cancer cells acquired radio-resistance. Due to variation in the tumor HDAC activity among patients, our report suggests performing a prior assessment of the tumor epigenome to maximize the benefit of HDAC inhibitor-based radio-sensitization.

Regulation of acetylation of plant cell wall components is complex and responds to external stimuli.

Previously, we reported that the allelic de-etiolated by zinc (dez) and trichome birefringence (tbr) mutants exhibit photomorphogenic development in the dark, which is enhanced by high Zn. TRICHOME BIREFRINGENCE-LIKE proteins had been implicated in transferring acetyl groups to various hemicelluloses. Pectin O-acetylation levels were lower in dark-grown dez seedlings than in the wild type. We observed Zn-enhanced photomorphogenesis in the dark also in the reduced wall acetylation 2 (rwa2-3) mutant, which exhibits lowered O-acetylation levels of cell wall macromolecules including pectins and xyloglucans, supporting a role for cell wall macromolecule O-acetylation in the photomorphogenic phenotypes of rwa2-3 and dez. Application of very short oligogalacturonides (vsOGs) restored skotomorphogenesis in dark-grown dez and rwa2-3. Here we demonstrate that in dez, O-acetylation of non-pectin cell wall components, notably of xyloglucan, is enhanced. Our results highlight the complexity of cell wall homeostasis and indicate against an influence of xyloglucan O-acetylation on light-dependent seedling development.

Photobiomodulation therapy modulates epigenetic events and NF-κB expression in oral epithelial wound healing.

The aim of this study was to evaluate the effect of photobiomodulation therapy (PBMT) on histone 3 acetylation (acH3) and NF-κB expression during oral ulcer healing. A total of 48 male Wistar rats were divided into control group (CG) and PBMT group (n = 24 each). Traumatic ulcers were created in the dorsum of the rats' tongue with a punch tool. Irradiation with InGaAlP laser, 660 nm, 40 mW, 0.04 cm2 spot size, 4 J/cm2, 4 s, and 0.16 J per spot was performed once a day in close contact for 10 consecutive days. CG received only daily handling. Rats were euthanized on days 3, 5, and 10 (n = 8) and were monitored daily to assess wound status. Immunohistochemical analysis for acH3 and NF-κB detection was performed. One thousand epithelial cells were counted, and mean acH3- and NF-κB-positive cells were calculated and compared between the groups. PBMT accelerated the repair of oral ulcers. On day 3, PBMT showed significantly higher means for acH3- and NF-κB-positive cells than CG. On day 5, no difference was observed between the groups concerning both markers. On day 10, PBMT presented lower acH3 and NF-κB means than the control group. We concluded that PBMT stimulates keratinocyte migration in the early stage of oral wound healing and keratinocyte differentiation at the final stage by modulating histone acetylation and NF-κB expression.

Radiation biology and oncology in the genomic era.

Radiobiology research is building the foundation for applying genomics in precision radiation oncology. Advances in high-throughput approaches will underpin increased understanding of radiosensitivity and the development of future predictive assays for clinical application. There is an established contribution of genetics as a risk factor for radiotherapy side effects. An individual's radiosensitivity is an inherited polygenic trait with an architecture that includes rare mutations in a few genes that confer large effects and common variants in many genes with small effects. Current thinking is that some will be tissue specific, and future tests will be tailored to the normal tissues at risk. The relationship between normal and tumor cell radiosensitivity is poorly understood. Data are emerging suggesting interplay between germline genetic variation and epigenetic modification with growing evidence that changes in DNA methylation regulate the radiosensitivity of cancer cells and histone acetyltransferase inhibitors have radiosensitizing effects. Changes in histone methylation can also impair DNA damage response signaling and alter radiosensitivity. An important effort to advance radiobiology in the genomic era was establishment of the Radiogenomics Consortium to enable the creation of the large radiotherapy cohorts required to exploit advances in genomics. To address challenges in harmonizing data from multiple cohorts, the consortium established the REQUITE project to collect standardized data and genotyping for ~5,000 patients. The collection of detailed dosimetric data is important to produce validated multivariable models. Continued efforts will identify new genes that impact on radiosensitivity to generate new knowledge on toxicity pathogenesis and tests to incorporate into the clinical decision-making process.

Alterations in histone acetylation following exposure to 60Co γ-rays and their relationship with chromosome damage in human lymphoblastoid cells.

Chromosome damage is related to DNA damage and erroneous repair. It can cause cell dysfunction and ultimately induce carcinogenesis. Histone acetylation is crucial for regulating chromatin structure and DNA damage repair. Ionizing radiation (IR) can alter histone acetylation. However, variations in histone acetylation in response to IR exposure and the relationship between histone acetylation and IR-induced chromosome damage remains unclear. Hence, this study investigated the variation in the total acetylation levels of H3 and H4 in human lymphocytes exposed to 0-2 Gy 60Co γ-rays. Suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor, was added to modify the histone acetylation state of irradiated cells. Then, the total acetylation level, enzyme activity, dicentric plus centric rings (dic + r) frequencies, and micronucleus (MN) frequencies of the treated cells were analyzed. Results indicated that the acetylation levels of H3 and H4 significantly decreased at 1 and 24 h, respectively, after radiation exposure. The acetylation levels of H3 and H4 in irradiated groups treated with SAHA were significantly higher than those in irradiated groups that were not treated with SAHA. SAHA treatment inhibited HDAC activity in cells exposed to 0-1 Gy 60Co γ-rays. SAHA treatment significantly decreased dic + r/cell and MN/cell in cells exposed to 0.5 or 1.0 Gy 60Co γ-rays relative to that in cells that did not receive SAHA treatment. In conclusion, histone acetylation is significantly affected by IR and is involved in chromosome damage induced by 60Co γ-radiation.

Quantitative proteomics analysis reveals alterations of lysine acetylation in mouse testis in response to heat shock and X-ray exposure.

Environmental stresses are important factors causing male infertility which attracts broad attention. Protein acetylation is a pivotal post-translational modification and modulates diverse physiological processes including spermatogenesis. In this study, we employed quantitative proteomic techniques and bioinformatics tools to analyze the alterations of acetylome profile of mouse testis after heat shock and X-irradiation. Overall, we identified 1139 lysine acetylation sites in 587 proteins in which 1020 lysine acetylation sites were quantified. The Gene Ontology analysis showed that the major acetylated protein groups were involved in generation of precursor metabolites and metabolic processes, and were localized predominantly in cytosolic and mitochondrial. Compared to the control group, 36 sites of 28 acetylated proteins have changed after heat shock, and 49 sites of 43 acetylated proteins for X-ray exposure. Some of the differentially acetylated proteins have been reported to be associated with the progression of spermatogenesis and male fertility. We observed the up-regulated acetylation level change on testis specific histone 2B and heat shock protein upon heat treatment and a sharp decline of acetylation level on histone H2AX under X-ray treatment, suggesting their roles in male germ cells. Notably, the acetylation level on K279 of histone acetyltransferase (Kat7) was down-regulated in both heat and X-ray treatments, indicating that K279 may be a key acetylated site and affect its functions in spermatogenesis. Our results reveal that protein acetylation might add another layer of complexity to the regulation for spermatogenesis, and further functional studies of these proteins will help us elucidate the mechanisms of abnormal spermatogenesis.

Chronic sun exposure is associated with distinct histone acetylation changes in human skin.

BACKGROUND: Photoageing is attributed to continuous sunlight or artificial ultraviolet exposure and manifests as clinical and histological changes in skin. Epigenetic changes have been found to be involved in the pathogenesis of photoageing. However, the underlying mechanisms are unclear. OBJECTIVES: To analyse histone modification patterns in sun-exposed and nonexposed skin, and to identify the abnormally histone-modified genes related to photoageing. METHODS: Skin biopsies were collected from both the outer forearm (sun-exposed area) and the buttock (sun-protected area) in 20 healthy middle-aged female volunteers. Global histone H3/H4 acetylation and H3K4/H3K9 methylation statuses were assessed by enzyme-linked immunosorbent assay. Expression levels of histone acetyltransferases and histone deacetylases were measured by reverse-transcriptase quantitative polymerase chain reaction (qPCR) and Western blot. Chromatin immunoprecipitation combined with DNA microarray (ChIP-chip) assay with anti-acetyl-histone H3 antibody in a sun-exposed pool (combining six sun-exposed skin samples) and a nonexposed pool (combining six nonexposed skin samples) was conducted to explore the abnormally acetylated histone H3 genes related to photoageing; ChIP-qPCR was then used to verify the results of ChIP-chip. RESULTS: We observed higher global histone H3 acetylation levels increased EP300 and decreased HDAC1 and SIRT1 expression in sun-exposed skin compared with matched nonexposed skin. Furthermore, the ChIP-chip assay showed that 227 genes displayed significant hyperacetylation of histone H3, and 81 genes displayed significant hypoacetylation of histone H3 between the two groups. Histone H3 acetylation levels on the promoters of PDCD5, ITIH5, MMP1 and AHR were positively correlated with the mRNA expression of the corresponding gene. CONCLUSIONS: Chronic sun exposure-induced histone H3 hyperacetylation may play a critical role in the pathogenesis of skin photoageing.

UV-Induced RPA1 Acetylation Promotes Nucleotide Excision Repair.

Replication protein A (RPA) is a multifunctional, single-stranded DNA-binding protein complex and plays a critical role in DNA replication and damage response. Herein, we show that the 70-kDa subunit of RPA (RPA1) is acetylated on lysine 163 by the acetyltransferases GCN5 and PCAF and that such acetylation is reversed principally via the action of the deacetylase HDAC6. UV irradiation promotes cytoplasmic translocation of HDAC6, thereby disrupting the interaction of HDAC6 with RPA1 and increasing RPA1 acetylation. Mutation of the acetylation site of RPA1 specifically impairs the ability of the protein to interact with the key nucleotide excision repair (NER) protein XPA, reduces XPA retention at sites of DNA damage caused by UV, compromises NER, and renders the cell hypersensitive to UV irradiation. Our data suggest that the acetylation status of RPA1 played a crucial role in repair of DNA damage via NER.

Low-dose radiation differentially regulates protein acetylation and histone deacetylase expression in human coronary artery endothelial cells.

PURPOSE: Ionizing radiation induces cardiovascular disease, the endothelium being the main target. The exact mechanism of the damage is unclear but the involvement of multiple signaling pathways is probable. Reversible lysine acetylation is a posttranslational protein modification that regulates activity across a broad range of signaling pathways. The aim of this study was to determine if a low radiation dose results in acetylome alteration in endothelial cells. MATERIALS AND METHODS: Human coronary artery endothelial cell line was irradiated with Cs-137 gamma-rays (0.5 Gy) and proteomics analysis was performed using enriched acetylated peptides and all peptides. Data were validated using immunoblotting, deacetylase activity assay, and RhoA activity assay. RESULTS: Nearly a hundred proteins were found to have an altered acetylation status 24 h after irradiation, primarily due to an overall decrease in acetylation. The expression of specific deacetylases was significantly increased, coinciding with an enhancement in global deacetylase activity. Proteins changed in their acetylation status belonged to several pathways including protein synthesis, cytoskeleton-related processes, protein folding and calcium signaling. The predicted changes in the RhoA/actin cytoskeleton pathway were validated by immunoassay. CONCLUSIONS: This study shows that protein acetylation is an important mediator of radiation responses in human cardiac coronary endothelial cells. Increased knowledge of the endothelial response to radiation is crucial for the development of normal tissue-sparing modalities during radiation therapy.

Calreticulin attenuated microwave radiation-induced human microvascular endothelial cell injury through promoting actin acetylation and polymerization.

Recent work reveals that actin acetylation modification has been linked to different normal and disease processes and the effects associated with metabolic and environmental stressors. Herein, we highlight the effects of calreticulin on actin acetylation and cell injury induced by microwave radiation in human microvascular endothelial cell (HMEC). HMEC injury was induced by high-power microwave of different power density (10, 30, 60, 100 mW/cm2, for 6 min) with or without exogenous recombinant calreticulin. The cell injury was assessed by lactate dehydrogenase (LDH) activity and Cell Counting Kit-8 in culture medium, migration ability, intercellular junction, and cytoskeleton staining in HMEC. Western blotting analysis was used to detected calreticulin expression in cytosol and nucleus and acetylation of globular actin (G-actin). We found that HMEC injury was induced by microwave radiation in a dose-dependent manner. Pretreatment HMEC with calreticulin suppressed microwave radiation-induced LDH leakage and increased cell viability and improved microwave radiation-induced decrease in migration, intercellular junction, and cytoskeleton. Meanwhile, pretreatment HMEC with exogenous calreticulin upregulated the histone acetyltransferase activity and the acetylation level of G-actin and increased the fibrous actin (F-actin)/G-actin ratio. We conclude that exogenous calreticulin protects HMEC against microwave radiation-induced injury through promoting actin acetylation and polymerization.

Lysine Acetylation and Succinylation in HeLa Cells and their Essential Roles in Response to UV-induced Stress.

Lysine acetylation and succinylation are major types of protein acylation that are important in many cellular processes including gene transcription, cellular metabolism, DNA damage response. Malfunctions in these post-translational modifications are associated with genome instability and disease in higher organisms. In this study, we used high-resolution nano liquid chromatography-tandem mass spectrometry combined with affinity purification to quantify the dynamic changes of protein acetylation and succinylation in response to ultraviolet (UV)-induced cell stress. A total of 3345 acetylation sites in 1440 proteins and 567 succinylation sites in 246 proteins were identified, many of which have not been reported previously. Bioinformatics analysis revealed that these proteins are involved in many important biological processes, including cell signalling transduction, protein localization and cell metabolism. Crosstalk analysis between these two modifications indicated that modification switches might regulate protein function in response to UV-induced DNA damage. We further illustrated that FEN1 acetylation at different sites could lead to different cellular phenotypes, suggesting the multiple function involvement of FEN1 acetylation under DNA damage stress. These systematic analyses provided valuable resources and new insight into the potential role of lysine acetylation and succinylation under physiological and pathological conditions.

Ultraviolet-B induces ERCC6 repression in lens epithelium cells of age-related nuclear cataract through coordinated DNA hypermethylation and histone deacetylation.

BACKGROUND: Ultraviolet-B (UVB) exposure attributes to the formation of age-related nuclear cataract (ARNC), which is mediated with DNA damage. DNA damage, an important factor for pathogenesis of ARNC, is induced by UVB, and is generally resolved by the nucleotide excision repair (NER) repair mechanism. Cockayne syndrome complementation group B (CSB) protein coded by ERCC6 is a vital component for NER. However, we found no association between selected ERCC6 polymorphisms and ARNC. In this study, we investigated whether UVB exposure could alter ERCC6 expression and the process could involve epigenetic changes of DNA methylation and/or histone acetylation of ERCC6 in the lens epithelial cells (LECs). We also assessed the involvement of those coordinated changes in lens tissue from ARNC patients. RESULTS: mRNA and protein expression of ERCC6 in lens tissue (LECs) were lower in ARNCs than those in the controls. This reduction corresponded to methylation of a CpG site at the ERCC6 promoter and histone modifications (methylation and acetylation) nearby this site. UVB-treated human lens epithelium B3 (HLE-B3) and 239T cell presented (1) increased apoptosis, suggesting reduced UV-damage repair, (2) hypermethylation of the CpG site located at position -441 (relative to transcription start site) within the binding region for transcriptional factor Sp1 in the ERCC6 promoter, (3) the enhancement of histone H3K9 deacetylation, (4) induction in DNA methyltransferases 3b (DNMT3b) and histone deacetylase1 (HDAC1) associated to the CpG site of ERCC6 by CHIP assay. CONCLUSIONS: These findings suggest an orchestrated mechanism triggered by UVB radiation where the concurrent association of specific hypermethylation CpG site, H3K9 deacetylation of ERCC6, and repression of ERCC6 gene expression. Taken together, with the similar changes in the lens tissue from ARNC patients, our data unveiled a possible mechanism of epigenetic modification of DNA repair gene in the pathogenesis of ARNC.