The Australian Epigenetic Alliance put together quite the action packed affair on August 26th, 2011 in honor of Dr. Feinberg’s visit to Melbourne, and we were fortunate enough to convince PhD student Sebastian Lunke to cover the event so we could get all of the juicy details, and pass them on to our readers. Check out Sebastian’s report below to get filled in:
When Richard Saffery, Jeff Craig, David Martino, Mark Cruickshank and Nick Wong, members of the Victorian branch of the Australian Epigenetic Alliance, heard that Andy Feinberg would be visiting Australia, they did not hesitate long and invited him to give a talk in Melbourne. Their efforts accumulated in a one-day epigenetics symposium at Bio21, with a variety of talks by junior and senior scientists centered on a plenary lecture given by Professor Feinberg, as well as a poster display.
The free symposium attracted over 130 scientists from around Australia and impressed through good organization and a very high standard of the presented scientific work. The overall atmosphere of the meeting was relaxed and familiar, and although newest developments in the field of epigenetics and the underlying changes in technology were discussed in depth, there was still time for quick excursions into the epigenetics of the flowers and the bees, as well as the introduction of ideas inspired by travel epiphanies, lollies or the yeast residing in the bottom of a beer glass.
The selected topics ranged from epigenetics in disease to the latest developments in bioinformatics. One theme however was common to almost all presentations: Whole genome analysis and next generation sequencing. Throughout the presentations by Prof. Feinberg and others it became very clear that analyzing only one or two epigenetic marks in selected locations is often not sufficient. A more global and correlative approach is essential to explain variations in gene expression patterns in disease as well as development.
A large amount of the data presented during the day was derived from whole genome micro-array or next generation sequencing experiments, indicating exciting new opportunities in way we can nowadays analyze the epigenome. With new technology however new challenges arise, as was evident from the talks on bioinformatics. The large emerging datasets require a level of data analysis and informatics skills unprecedented in the field, and promise to keep discussions about proper normalization and interpretation heated for some time to come.
The Epigenetic Basis of Common Human Disease
Andy Feinberg, Johns Hopkins University
During his lecture, Prof. Feinberg introduced the idea that stochastic epigenetic variation is the underlying mechanism for phenotypic variation, as well as certain aspects of development and disease.
He started by describing how phenotypic variation can easily be attributed to differences in the underlying genetics when comparing different organisms. This however does not explain the large degree of variation we observe within a species, and even more so in a single multi-cellular organism. Every cell in the human body has the same genetic blueprint, yet there are was differences observed between different tissues and organs. The way the genetic code is interpreted is determined by the epigenome, which drives the specific expression patterns in each cell type. Moreover the epigenetic landscape is influences by the environment, and changes in epigenetic modifications like DNA methylation can cause a broad variety of diseases, including cancer.
Prof. Feinberg presented data derived from his work with Rafael Irizarry, which showed that the genome includes a large number of variable methylation regions (VMRs), which show heterogeneous levels of methylation even in inbreed laboratory mice. These VMRs could be responsible for phenotypic variation between specimens with similar genetic background, and their ability to be influenced by environmental stimuli potentially designates them an important role in adaptation and development. Furthermore they observed that VMRs are stable over time in normal tissue, but have a high degree of variation in cancer cells, which aids in the explanation of high levels of heterogeneity in cancer cells. Interestingly, these VMRs are the same that show high variability during development, underlining the importance of this variation in disease.
He also showed that stochastic variation in epigenetics is very likely to be necessary for the successful development of complex organisms, as mere genetic variation under the influence of selection pressure, the current standard model of adaptation, would in the long term not be able to efficiently respond to a rapidly changing environment. A combination of genetic variation with epigenetic variation, the degree of which is in turn genetically inherited, would be a more likely model while still concordant with Darwinian evolution.
Galaxy Framework for Data Analysis
Ross Lazarus, Harvard Medical School
The first talk of the day, presented by Ross Lazarus, introduced the Galaxy framework (http://usegalaxy.org) and its application in epigenetic data analysis to the audience. Galaxy is an open-source, web based bioinformatics project, designed to allow free and easy access to a large variety to complex bioinformatics and analysis tools. It is designed to allow reproducible data analysis, including grooming, alignment and analysis of next generation sequencing data, and aims to make commonly used bioinformatics tools available to scientist without requiring in depth bioinformatics skills. The open-source nature of the project allows for fast adaptation to a rapidly developing bioinformatics field as well as easy integration with third party software and databases.
Genome-Scale DNA Methylation Mapping
Aniruddha Chatterjee, University of Otago
The second talk was given by Aniruddha Chatterjee and described the challenges he faced in the analysis of genome wide reduced representation bisulfite sequencing (RRBS) data. He shared his experience in optimizing three of the publicly available alignment algorithms for RRBS derived data (BISMARK, RMAPBS and BSMAP) as well as a quality control script implemented by his lab. The aim of Aniruddha’s efforts is to unravel inter-individual variation in DNA methylation and he presented his initial analysis of RRBS data from three individuals.
Visualization and Interpretation of Chip-seq Data
Alicia Oshlack, Walter & Eliza Hall Institute
The last talk in this session was the keynote presentation by Alicia Oshlack, focused on the visualization, analysis and interpretation of epigenomic ChIP-seq data. In this keynote presentation Alicia pointed out the complexity of the epigenome, which to date comprises over 100 described epigenetic modifications, and the challenges in normalizing and analyzing genome wide ChIP-seq data. Her main focus was on bioinformatics tools developed by her and her co-workers (Repitools, edgeR), which aim to reduce noise inherent in ChIP-seq experiments as well as to correlate the vast amounts of epigenetic data with gene expression. After stressing the importance of analyzing the derived data in a genome-wide context, and not only in specific regions, she also emphasized the necessity of using input controls in all ChIP-seq experiments to enable correction for technical biases (such as shearing bias) introduced based on the variation in the epigenomic landscape of different cells or between treatment-control data sets.
Smchd1 in Cancer
Huei Sang Leong, Walter & Eliza Hall Institute
The second session was opened by Huei Sang Leong. Huei investigates the role of Smchd1 in cancer. Experiments in various tumor mouse models showed accelerated tumor. These results point to a role for Smchd1 as tumor suppressor, potentially linked to interaction with Myc and PCR2.
Epimutations of the MLH1 Gene
Megan Hitchins, Lowry Cancer Research Centre
Next Megan Hitchins gave a keynote presentation about Mendelian and non-Mendelian transmission patterns of MLH1 epimutations and potential underlying mechanisms. Megan analyzed several families with a history of Lynch syndrome, which could not be explained by loss of function through germline mutations or bi-allelic methylation. She was able to attribute the phenotype to epimutations, which followed different inheritance patterns in the various families. While the underlying mechanisms for the non-mendelian inheritance patterns could potentially be attributed to trans-effects or co-epimutations, Megan was able in one family to identify a single nucleotide substitution in the MLH1 promoter that caused a dominant inheritance of the epimutation.
This specific substitution caused a hyper-mosaic methylation pattern on the MLH1 promoter of the affected allele, resulting in diminished MLH1 expression. Moreover, she presented compelling evidence that the same nucleotide substitution is transmitted through the male germ line and capable of re-establishing the epimutation in the next generation despite a complete erasure of DNA-methylation in the sperm. Albeit the described variation is exceedingly rare, it goes to show that even smallest changes in the underlying sequence can have important effects on the surrounding epigenetic landscape and gene expression.
Effects of HDAC Inhibitors in AML
Jessica Salmon, Peter MacCallum Caner Centre
After lunch Jessica Salmon presented her work on targeting histone acetylation with HDAC inhibitors and the resulting effects on leukemic blasts in a mouse model of acute myeloid leukemia. The mice were treated with the HDACi Panobinostat, which resulted in decreased tumor progression and increased survival of the mice. Jessica and her co-workers were able to attribute this favorable effect to a decrease in the AML1-ETO fusion protein, which is responsible for accumulation of immature myeloid blasts in the blood stream of affected mice. Moreover they were able to show that Panobinostat does not induce apoptosis of the tumor cells, but rather pushes them towards differentiation and maturation.
Epigenetics Changes in Early Breast Carcinogenesis
Warwick James, The Garvan Institute of Medical Research
Following the theme of mouse cancer epigenetics, Warwick James next presented the effort of his lab to model the epigenetic change during early breast carcinogenesis. Studies were undertaken in cultured HMEC cells, which after several passages display a cancer associated phenotype, as well as cells derived from a novel mouse model of breast cancer. Initial analysis of ChIP-Seq, MBD-Seq and expression data reveals large-scale changes of the epigenome in vitro associated with early carcinogenesis, specifically through the silencing of cancer associated genes by increases in DNA methylation and repressive histone marks. A possible involvement of long range epigenetic silencing was also discussed.
Regulation of Cancer Associated miRNAs
Eugene Wee, University of Queensland
Also studying breast cancer, Eugene Wee and his colleagues aim to understand the regulation of cancer-associated miRNAs. To this end, they studied the regulatory regions of 93 known miRNAs and identified a correlation between DNA-methylation of two promoter regions and the expression of miR-200b. They were additionally able to correlate the cancer tissue specific methylation status of miR-200b promoters with the expression of various receptors.
Differentially Methylated and Expressed Genes in Food Allergies
David Martino, Murdoch Children’s Research Institute
The last session opened with a presentation by David Martino, who presented his work in elucidating the molecular and epigenetic basis of food allergies. David and his co-workers studied expression levels and DNA-methylation in CD4+ extracted at birth and twelve month after birth from children with and without food allergies. They identified life spread changes in RNA levels and DNA methylation in relation with age. They were also able to isolate a number of differentially expressed genes at birth in children with food allergies, that were associated with a reduced ability of T-cells to proliferate in in-vitro assays and accompanied by changes in DNA-methylation.
These changes resulted in differential baseline expression of Th2-associated genes in the same children at the 12 months time point. Their data provided a first molecular insight in how neonatal T-cell activation pathways can increase the risk of food allergy.
Fetal Vitamin D levels and CYP24A1 Regulation
Boris Novakovic, Murdoch Children’s Research Institute
Next Boris Novakovic studied the association between epigenetic CYP24A1 gene regulation and fetal vitamin D levels in twins. Fetal vitamin D levels are dependent on the supply of the hormone through the placenta. Studies have shown that the promoter of the CYP24A1 gene, the product of which encodes an enzyme that catabolises active vitamin D, is highly methylated in placental compared to other tissue. So far however this study however has failed to establish a direct link between variations in CYP24A1 promoter methylation and fetal/maternal vitamin D levels, with maternal vitamin D levels identified as the major contributor to the levels in the fetus.
Remodeling is at the Heart of Chromatin
Assam El-Osta, Baker IDI
The meeting concluded with a keynote presentation by Assam El-Osta, who introduced to the audience the “Sweet Genome”, a study evaluating changes in the epigenomic landscape in response to glucose. The findings of several clinical studies in diabetes (DCCT, EDICT, UKPDS) indicated lasting beneficial effects of tight glucose control even after the conclusion of the initial study, a phenomenon termed “legacy effect” or “hyperglycemic memory”. To investigate the molecular basis of these findings, Assam and his group established an in vitro model of hyperglycemic memory, where they exposed HAEC cells to high glucose conditions for 16 hours before returning the cells to normal glucose conditions for extended periods of time. Changes on gene expression and epigenetic marks were analyzed at several time points up to six days after return to normal conditions.
They were able to show a sustained increase in p65 expression in response to high glucose, which was maintained for several days after return to normal glucose conditions and associated with changes in histone 3 lysine 4 methylation as well as Set7 histone methyl-transferase expression. Genome wide analysis of changes in epigenetic marks in response to high glucose revealed significant changes in histone acetylation and DNA methylation in more than 5000 regions, most of which were associated with promoter regions. Gene expression profiling using RNA-Seq unveiled changes in a large number of genes associated with metabolic and cardiovascular disease. Based on preliminary results, these findings appear to transfer into an in-vitro setting as well and provide the first insight into a potential epigenetic basis of hyperglycemic memory.
Poster and presentation awards:
- Aniruddha Chatterjee from the University of Otago, New Zealand, won AGRF Young Investigator Award for an oral presentation (PhD student) for his talk “Genome-Scale DNA Methylation Mapping: Unravelling Inter-individual Variation”
- David Martino from the MCRI in Melbourne won the Illumina Young Investigator Award for oral presentation (Early Career Postdoc) for his talk “Genome-wide identification of differentially methylated and expressed genes in CD4+ T cells from children who develop IgE food allergy”
- Kevin Knower from Prince Henry’s Institute for Medical Research won the Monash Health Translation Precinct Poster Prize for his poster “Epigenetic influences regulating intratumoral estrogen production in postmenopausal breast cancer”
- Shamista Selvarajah from the Dept of Medicine, University of Melbourne, won the Monash Health Translation Precinct Poster Prize for her poster on “The histone variants H2Bv and H2A.Z cooperate in nucleosomes in gene promoters in the malaria parasite P. falciparum”
**EpiGenie would like to thank Sebastian Lunke for supplying such thorough coverage of this symposium. Sebastian is a PhD student in the Epigenetics in Human Health and Disease Lab at the Baker IDI Heart and Disease Institute in Melbourne, Austrailia.