Effects of the omega-3 fatty acid DHA on histone and p53 acetylation in diffuse large B cell lymphoma.

Diffuse large B cell lymphoma (DLBCL) often develops resistance and/or relapses in response to immunochemotherapy. Epigenetic modifiers are frequently mutated in DLBCL, i.e., the lysine (histone) acetyltransferases CREBBP and EP300. Mutations in CBP/p300 can prevent the proper acetylation and activation of (i) enhancer sequences of genes required for essential functions (e.g., germinal center exit and differentiation) and (ii) the tumor suppressor p53. Based on evidence that omega-3 fatty acids (ω-3 FAs) affect histone acetylation in various cancers, we investigated whether ω-3 FA docosahexaenoic acid (DHA) could modify levels of histone and p53 acetylation in three DLBCL cell lines (at different CREBBP/EP300 mutational status) versus normal B cells. Exposure to DHA at clinically attainable doses was shown to significantly alter the genome-wide levels of histone posttranslational modifications in a cell-line-dependent and dose-dependent manner. Although histone acetylation did not increase uniformly, as initially expected, levels of p53 acetylation increased consistently. Quantitative reverse transcription polymerase chain reaction results revealed significant changes in expression of multiple genes, including increased expression of CREBBP and of PRDM1 (required for differentiation into plasma cells or memory B cells). Taken together, our results provide (to our knowledge) the first characterization of the epigenetic effects of ω-3 FAs in DLBCL.

Where Epigenetics Meets Food Intake: Their Interaction in the Development/Severity of Gout and Therapeutic Perspectives.

Gout is the most frequent form of inflammatory arthritis in the world. Its prevalence is particularly elevated in specific geographical areas such as in the Oceania/Pacific region and is rising in the US, Europe, and Asia. Gout is a severe and painful disease, in which co-morbidities are responsible for a significant reduction in life expectancy. However, gout patients remain ostracized because the disease is still considered "self-inflicted", as a result of unhealthy lifestyle and excessive food and alcohol intake. While the etiology of gout flares is clearly associated with the presence of monosodium urate (MSU) crystal deposits, several major questions remain unanswered, such as the relationships between diet, hyperuricemia and gout flares or the mechanisms by which urate induces inflammation. Recent advances have identified gene variants associated with gout incidence. Nevertheless, genetic origins of gout combined to diet-related possible uric acid overproduction account for the symptoms in only a minor portion of patients. Hence, additional factors must be at play. Here, we review the impact of epigenetic mechanisms in which nutrients (such as ω-3 polyunsaturated fatty acids) and/or dietary-derived metabolites (like urate) trigger anti/pro-inflammatory responses that may participate in gout pathogenesis and severity. We propose that simple dietary regimens may be beneficial to complement therapeutic management or contribute to the prevention of flares in gout patients.

Epigenetic Reprogramming Mediated by Maternal Diet Rich in Omega-3 Fatty Acids Protects From Breast Cancer Development in F1 Offspring.

Diets rich in omega-3 fatty acids (FA) have been associated with lowered risks of developing certain types of cancers. We earlier reported that in transgenic mice prone to develop breast cancer (BCa), a diet supplemented with canola oil, rich in omega-3-rich FA (as opposed to an omega-6-rich diet containing corn oil), reduced the risk of developing BCa, and also significantly reduced the incidence of BCa in F1 offspring. To investigate the underlying mechanisms of the cancer protective effect of canola oil in the F1 generation, we designed and performed the present study with the same diets using BALB/c mice to remove any possible effect of the transgene. First, we observed epigenetic changes at the genome-wide scale in F1 offspring of mothers fed diets containing omega-3 FAs, including a significant increase in acetylation of H3K18 histone mark and a decrease in H3K4me2 mark on nucleosomes around transcription start sites. These epigenetic modifications contribute to differential gene expressions associated with various pathways and molecular mechanisms involved in preventing cancer development, including p53 pathway, G2M checkpoint, DNA repair, inflammatory response, and apoptosis. When offspring mice were exposed to 7,12-Dimethylbenz(a)anthracene (DMBA), the group of mice exposed to a canola oil (with omega 3 FAs)-rich maternal diet showed delayed mortality, increased survival, reduced lateral tumor growth, and smaller tumor size. Remarkably, various genes, including BRCA genes, appear to be epigenetically re-programmed to poise genes to be ready for a rapid transcriptional activation due to the canola oil-rich maternal diet. This ability to respond rapidly due to epigenetic potentiation appeared to contribute to and promote protection against breast cancer after carcinogen exposure.

Genetic and epigenetic determinants of diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoma and is notorious for its heterogeneity, aggressive nature, and the frequent development of resistance and/or relapse after treatment with standard chemotherapy. To address these problems, a strong emphasis has been placed on researching the molecular origins and mechanisms of DLBCL to develop effective treatments. One of the major insights produced by such research is that DLBCL almost always stems from genetic damage that occurs during the germinal center (GC) reaction, which is required for the production of high-affinity antibodies. Indeed, there is significant overlap between the mechanisms that govern the GC reaction and those that drive the progression of DLBCL. A second important insight is that some of the most frequent genetic mutations that occur in DLBCL are those related to chromatin and epigenetics, especially those related to proteins that "write" histone post-translational modifications (PTMs). Mutation or deletion of these epigenetic writers often renders cells unable to epigenetically "switch on" critical gene sets that are required to exit the GC reaction, differentiate, repair DNA, and other essential cellular functions. Failure to activate these genes locks cells into a genotoxic state that is conducive to oncogenesis and/or relapse.

40th International Asilomar Chromatin, Chromosomes, and Epigenetics Conference.

The 40th International Asilomar Chromatin, Chromosomes, and Epigenetics Conference was held in the Asilomar Conference Grounds, Pacific Grove, California, USA, on 6-9 December 2018. The organizing committee consisted of established scientists in the fields of chromatin and epigenetics: Sally Pasion and Michael Goldman from the Biology Department, San Francisco State University, California, USA; Philippe Georgel from the Department of Biological Sciences, Marshal University, West Virginia, USA; Juan Ausió from the Department of Biochemistry and Microbiology, University of Victoria, British Columbia, Canada; and Christopher Eskiw from the Department of Biochemistry, University of Saskatchewan, Saskatchewan, Canada. The meeting had two keynote speakers: Jessica Tyler and Jennifer Mitchell, and it covered topics on transcription, replication and repair, epigenetics, cell differentiation and disease, telomeres, and centromeres and it had two sessions devoted to nuclear and genomic organization. It encompassed the enthusiastic presentations of excellent trainees within the breathtaking natural setting of Pacific Grove.

MeCP2 and CTCF: enhancing the cross-talk of silencers.

This paper provides a brief introductory review of the most recent advances in our knowledge about the structural and functional aspects of two transcriptional regulators: MeCP2, a protein whose mutated forms are involved in Rett syndrome; and CTCF, a constitutive transcriptional insulator. This is followed by a description of the PTMs affecting these two proteins and an analysis of their known interacting partners. A special emphasis is placed on the recent studies connecting these two proteins, focusing on the still poorly understood potential structural and functional interactions between the two of them on the chromatin substrate. An overview is provided for some of the currently known genes that are dually regulated by these two proteins. Finally, a model is put forward to account for their possible involvement in their regulation of gene expression.

Sulforaphane modulates telomerase activity via epigenetic regulation in prostate cancer cell lines.

Epidemiologic studies have revealed that diets rich in sulforaphane (SFN), an isothiocyanate present in cruciferous vegetables, are associated with a marked decrease in prostate cancer incidence. The chemo-preventive role of SFN is associated with its histone de-acetylase inhibitor activity. However, the effect of SFN on chromatin composition and dynamic folding, especially in relation to HDAC inhibitor activity, remains poorly understood. In this study, we found that SFN can inhibit the expression and activity of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, in 2 prostate cancer cell lines. This decrease in gene expression is correlated with SFN-induced changes in chromatin structure and composition. The SFN-mediated changes in levels of histone post-translational modifications, more specifically acetylation of histone H3 lysine 18 and di-methylation of histone H3 lysine 4, 2 modifications linked with high risk of prostate cancer recurrence, were associated with regulatory elements within the hTERT promoter region. Chromatin condensation may also play a role in SFN-mediated hTERT repression, since expression and recruitment of MeCP2, a known chromatin compactor, were altered in SFN treated prostate cancer cells. Chromatin immuno-precipitation (ChIP) of MeCP2 showed enrichment over regions of the hTERT promoter with increased nucleosome density. These combined results strongly support a role for SFN in the mediation of epigenetic events leading to the repression of hTERT in prostate cancer cells. This ability of SFN to modify chromatin composition and structure associated with target gene expression provides a new model by which dietary phytochemicals may exert their chemoprevention activity.

The danger of epigenetics misconceptions (epigenetics and stuff ).

Within the past two decades, the fields of chromatin structure and function and transcription regulation research started to fuse and overlap, as evidence mounted to support a very strong regulatory role in gene expression that was associated with histone post-translational modifications, DNA methylation, as well as various chromatin-associated proteins (the pillars of the "Epigenetics" building). The fusion and convergence of these complementary fields is now often simply referred to as "Epigenetics". During these same 20 years, numerous new research groups have started to recognize the importance of chromatin composition, conformation, and its plasticity. However, as the field started to grow exponentially, its growth came with the spreading of several important misconceptions, which have unfortunately led to improper or hasty conclusions. The goal of this short "opinion" piece is to attempt to minimize future misinterpretations of experimental results and ensure that the right sets of experiment are used to reach the proper conclusion, at least as far as epigenetic mechanisms are concerned.

Breaking the cycle: the role of omega-3 polyunsaturated fatty acids in inflammation-driven cancers.

Chronic inflammation is a cyclical, self-stimulating process. Immune cells called to sites of inflammation release pro-inflammatory signaling molecules that stimulate activation of inducible enzymes and transcription factors. These enzymes and transcription factors then stimulate production of signaling molecules that attract more immune cells and induce more enzymatic and transcriptional activity, creating a perpetual loop of inflammation. This self-renewing pool of inflammatory stimuli makes for an ideal tumor microenvironment, and chronic inflammation has been linked to oncogenesis, tumor growth, tumor cell survival, and metastasis. Three protein pathways in particular, nuclear factor kappa B (NF-kB), cyclooxygenase (COX), and lipoxygenase (LOX), provide excellent examples of the cyclical, self-renewing nature of chronic inflammation-driven cancers. NF-kB is an inducible transcription factor responsible for the expression of a vast number of inflammation and cancer related genes. COX and LOX convert omega-6 (n-6) and omga-3 (n-3) polyunsaturated fatty acids (PUFA) into pro- and anti-inflammatory signaling molecules. These signaling molecules stimulate or repress activity of all three of these pathways. In this review, we will discuss the pro- and anti-inflammatory functions of these fatty acids and their role in chronic inflammation and cancer progression.

The epigenetic potentials of dietary polyphenols in prostate cancer management.

Prostate cancer is a disease that is greatly affected by lifestyle, particularly diet, and is more prevalent in US and European countries compared with South and East Asia. Among several known causes and risk factors, nutrition plays an important role in prostate cancer pathogenesis. Various dietary components including polyphenols have been shown to possess anticancer properties. Dietary polyphenols have been the subject of extensive studies for the last decade because of their anticancer and chemopreventive potentials. Besides possessing various antitumor properties, dietary polyphenols also contribute to epigenetic changes associated with the fate of cancer cells and have emerged as potential drugs for therapeutic intervention. Various polyphenols have been shown to affect DNA methylation, histone posttranslational modifications, and microRNA expression patterns in prostate cancer. In this review, we discuss the contribution of dietary polyphenols to various epigenetic modifications in prostate cancer. Since prostate cancer and diet are intimately associated, polyphenol-rich diets that epigenetically modify tumor biology have great significance in the prevention and management of prostate cancer.

MeCP2 binds to nucleosome free (linker DNA) regions and to H3K9/H3K27 methylated nucleosomes in the brain.

Methyl-CpG-binding protein 2 (MeCP2) is a chromatin-binding protein that mediates transcriptional regulation, and is highly abundant in brain. The nature of its binding to reconstituted templates has been well characterized in vitro. However, its interactions with native chromatin are less understood. Here we show that MeCP2 displays a distinct distribution within fractionated chromatin from various tissues and cell types. Artificially induced global changes in DNA methylation by 3-aminobenzamide or 5-aza-2'-deoxycytidine, do not significantly affect the distribution or amount of MeCP2 in HeLa S3 or 3T3 cells. Most MeCP2 in brain is chromatin-bound and localized within highly nuclease-accessible regions. We also show that, while in most tissues and cell lines, MeCP2 forms stable complexes with nucleosome, in brain, a fraction of it is loosely bound to chromatin, likely to nucleosome-depleted regions. Finally, we provide evidence for novel associations of MeCP2 with mononucleosomes containing histone H2A.X, H3K9me(2) and H3K27me(3) in different chromatin fractions from brain cortex and in vitro. We postulate that the functional compartmentalization and tissue-specific distribution of MeCP2 within different chromatin types may be directed by its association with nucleosomes containing specific histone variants, and post-translational modifications.

MeCP2: structure and function.

Despite a vast body of literature linking chromatin structure to regulation of gene expression, the role of architectural proteins in higher order chromatin transitions required for transcription activation and repression has remained an under-studied field. To demonstrate the current knowledge of the role of such proteins, we have focused our attention on the methylated DNA binding and chromatin-associated protein MeCP2. Structural studies using chromatin assembled in vitro have revealed that MeCP2 can associate with nucleosomes in an N-terminus dependent manner and efficiently condense nucleosome arrays. The present review attempts to match MeCP2 structural domains, or lack thereof, and specific chromatin features needed for the proper recruitment of MeCP2 to its multiple functions as either activator or repressor. We specifically focused on MeCP2's role in Rett syndrome, a neurological disorder associated with specific MeCP2 mutations.

Chromatin stability at low concentration depends on histone octamer saturation levels.

Studies on the stability of nucleosome core particles as a function of concentration have indicated a lower limit of approximately 5 ng/microL, below which the complexes start to spontaneously destabilize. Until recently little information was available on the effect of low concentration on chromatin. Using the well-characterized array of tandemly repeated 5S rDNA reconstituted into chromatin, we have investigated the effect of dilution. In this study, we demonstrate that the stability of saturated nucleosomal arrays and that of nucleosome core particles are within the same order of magnitude, and no significant loss of histones is monitored down to a concentration of 2.5 ng/microL. We observed that levels of subsaturation of the nucleosomal arrays were directly correlated with an increased sensitivity to histone loss, suggesting a shielding effect. The loss of histones from our linear nucleosomal arrays was shown not to be random, with a significant likelihood to occur at the end of the template than toward the center. This observation indicates that centrally located nucleosomes are more stable than those close to the end of the DNA templates. Itis important to take this information into account for the proper design of experiments pertaining to histone composition and the folding ability of chromatin samples.

Role of nucleic acid binding in Sir3p-dependent interactions with chromatin fibers.

Recent studies of the mechanisms involved in the regulation of gene expression in eukaryotic organisms depict a highly complex process requiring a coordinated rearrangement of numerous molecules to mediate DNA accessibility. Silencing in Saccharomyces cerevisiae involves the Sir family of proteins. Sir3p, originally described as repressing key areas of the yeast genome through interactions with the tails of histones H3 and H4, appears to have additional roles in that process, including involvement with a DNA binding component. Our in vitro studies focused on the characterization of Sir3p-nucleic acid interactions and their biological functions in Sir3p-mediated silencing using binding assays, EM imaging, and theoretical modeling. Our results suggest that the initial Sir3p recruitment is partially DNA-driven, highly cooperative, and dependent on nucleosomal features other than histone tails. The initial step appears to be rapidly followed by the spreading of silencing using linker DNA as a track.

CHD proteins: a diverse family with strong ties.

Chromodomain/helicase/DNA-binding domain (CHD) proteins have been identified in a variety of organisms. Despite common features, such as their chromodomain and helicase domain, they have been described as having multiple roles and interacting partners. However, a common theme for the main role of CHD proteins appears to be linked to their ATP-dependent chromatin-remodeling activity. Their actual activity as either repressor or activator, and their cell or gene specificity, is connected to their interacting partner(s). In this minireview, we attempt to match the members of the CHD family with the presence of structural domains, cofactors, and cellular roles in the regulation of gene expression, recombination, genome organization, and chromatin structure, as well as their potential activity in RNA processing.

The use of Quantitative Agarose Gel Electrophoresis for rapid analysis of the integrity of protein-DNA complexes.

Recent biochemical studies evaluated the affinity of histones to DNA in the context of nucleosome core particle (NCP). These have indicated a concentration-dependence for nucleosome stability. However, when studying chromatin the preferred templates are nucleosome arrays (NA) and not the NCP. Biochemical methods are poorly suited for structural analysis of chromatin. To overcome that technical hindrance, and investigate the effect of concentration on stability of the histone-DNA interactions, we have applied the multigel Quantitative Agarose Gel Electrophoresis (QAGE) method to in vitro-assembled nucleosomal arrays. The results demonstrated the method to be extremely valuable for the evaluation of the effect of low concentration on NA. However, QAGE is a fairly time-demanding and complex method. To maximize the efficiency of use of this technology, we devised a protocol that allowed for multiple sets of templates to be analyzed simultaneously. Briefly, samples can be loaded at regular intervals and analyzed individually for their molecular composition. The technique presented in this study describes the calibration steps and proof of concept necessary to validate the use of multiple loading of multigel to evaluate the composition of nucleosomal arrays as a function of concentration.

Role of chromatin/epigenetic modifications on DNA accessibility.

From a therapeutic, and more precisely a DNA-drug interactions point of view, multiple factors can regulate DNA accessibility. In addition to various levels of chromatin compaction described in this review, DNA modifications such as methylation, the presence of specific DNA-binding proteins and chromatin-associated proteins, and, as was shown more recently, noncoding RNA can also contribute. The present review will discuss the importance of these epigenetic factors in modulating drug binding to DNA and the potential design of new therapeutic regimens aimed at improving DNA targeting.

HDAC1 acetylation is linked to progressive modulation of steroid receptor-induced gene transcription.

Although histone deacetylases (HDACs) are generally viewed as corepressors, we show that HDAC1 serves as a coactivator for the glucocorticoid receptor (GR). Furthermore, a subfraction of cellular HDAC1 is acetylated after association with the GR, and this acetylation event correlates with a decrease in promoter activity. HDAC1 in repressed chromatin is highly acetylated, while the deacetylase found on transcriptionally active chromatin manifests a low level of acetylation. Acetylation of purified HDAC1 inactivates its deacetylase activity, and mutation of the critical acetylation sites abrogates HDAC1 function in vivo. We propose that hormone activation of the receptor leads to progressive acetylation of HDAC1 in vivo, which in turn inhibits the deacetylase activity of the enzyme and prevents a deacetylation event that is required for promoter activation. These findings indicate that HDAC1 is required for the induction of some genes by the GR, and this activator function is dynamically modulated by acetylation.

Characterization of chromatin samples in the presence of Drosophila embryo extract by quantitative agarose gel electrophoresis.

Recent studies have focused attention on chromatin as both a negative and positive regulator of specific nuclear events. The vast majority of this research has been centered on chromatin remodeling and histone post-translational modifications over the regulatory regions of specific genes. However, due the technical difficulties of such studies, the contribution of the higher-order structure of chromatin on the regulation of gene expression has not been as thoroughly investigated and the majority of the initial studies have used biophysical methods or microscopy. Until recent technical developments, the main hindrance for these biophysical investigations of chromatin has been an almost absolute requirement for large amounts of highly purified material. The development of an agarose gel electrophoresis method (quantitative agarose gel electrophoresis), initially designed for the analysis of the three-dimensional structure of purified and in vivo-assembled chromatin over a promoter region, has been expanded to include studies of chromatin in the presence of a Drosophila crude extract. The technique presented in the study reported here will help in paving the way for subsequent analyses of structural modifications of chromatin that are linked with the recruitment of various chromatin-associated factors present in the provided extract(s).