Epigenetics in Human Health and Disease

Unit Code

01068

Department/Institution Offering Unit

The Alfred Medical Research and Education Precinct (AMREP)
Baker Medical Research Institute
Second Floor Commercial Road
Prahran
Victoria 3181 Australia

tel: + 61 3 8532 1389
fax: +61 3 8532 1100
web: www.baker.edu.au

Unit Points

Research Points: 100

Coursework Points: 0

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Contacts

Unit Coordinator/s:

Assam El-Osta (PhD)
Team Leader
Epigenetics in Human Health and Disease Laboratory
The Alfred Medical Research and Education Precinct (AMREP)
Baker Medical Research Institute
Second Floor Commercial Road
Prahran
Victoria 3181 Australia

tel: +61 3 8532 1389
fax: +61 3 8532 1100
email: assam.el-osta@baker.edu.au

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Overall Objectives

The unit is designed to foster curiousity in human molecular genetics by asking questions.

• Identify and to define the research problem.
• Review published materials related to the medical problem or scientific question and to evaluate possible solutions.
• Challenge and test hypotheses through careful and well thought-out experimentation, followed by data collection and analysis.
• Evaluation of experimental results, discussion with colleagues and detail how the data substantiate the conclusions.
• Prepare a report of the research findings and exhibit by presentation.

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Content

Coursework: N/A

Research: The term "epigenetics" which literally means "outside conventional genetics" is often used to describe the study of stable alterations that arise during development and cell proliferation. Changes in DNA methylation and chromatin remodelling are major determinants in the epigenetic control of genes and have profound roles in transcriptional regulation and DNA behaviour. The mechanisms underlying the targeting of DNA methylation and the subsequent regulation of transcription are relevant to human development and disease, as well as for attempts at somatic gene therapy. We are interested in examining epigenetic events such as DNA methylation, histone modification and the determinants involved on chromatin to further our understanding of endogenous gene transcription. These studies are now challenging the way we view gene regulation beyond our simple understanding of "textbook" operations. Our desire to dissect the molecular details allows us to determine the roles of transcriptional regulators and provide us with a greater understanding of how they are involved in transcription. The success of silencing therapeutically relevant genes in cancer or the reactivation of genes in developmental syndromes involved in human mental disease is molecularly connected by virtue of DNA methylation and chromatin modifications. One of our goals is to exploit this information to resolve how genes are regulated in heart disease, cancer and the developmental syndromes such as Fragile X disease. Our laboratory has a focus on gene regulation. We have generally relied on genetic and biochemical systems to study transcription. Genetics excels in identifying transcription factors and regulatory complexes required for expression, whereas biochemical testing transcend what they are capable of in vitro. However, neither discipline can tell us definitely the order of molecular events that naturally occur on genes. It is the very understanding of these mechanisms of transcriptional regulation that allows the biologist to manipulate a molecular system often seen as difficult to control. Research on these epigenetic events provides an exciting opportunity to control therapeutically relevant genes that have historically eluded translational science.

Sample Research Topics:

1. Fragile X gene rescue by knockout of key transcriptional regulators
   
   Fragile X syndrome (FRAXA, OMIM 309550 Online Mendelian Inheritance in Man) is one of the most common forms of inherited mental impairment in humans. There is currently no cure for fragile X individuals. The disease is linked with perhaps the most pertinent of all epigenetic modifications, DNA methylation. This modification to DNA switches the gene off. One of our goals in the lab is to rescue the fragile X phenotype by manipulating transcriptional regulation. We have identified a number of molecular determinants involved in the binding and repression of the fragile X gene. This project involves somatic knockout of these regulators. Our desire to dissect the molecular details of fragile X transcription allows us to determine the role of the chromatin regulators in our attempts to resolve fragile X disease.
2. Gene Therapy Projects
   
   Until now, almost all activity in the field of gene control focussed on the role of transcription factors as the sole participants that regulate gene behaviour. Despite the fact that we have known for almost 30 years that DNA is organized into chromosomal domains that controls gene expression. These chromosome domains are referred to as "chromatin" and play a central role in all aspects of DNA biology and are typified by the spectacular molecular events of DNA replication, chromosomal packaging and exquisite transcriptional control. The mechanisms underlying the control of gene expression are dependent on modifications to chromatin and are relevant to human development and disease and have come to prominence with the growing interest of gene therapy. Of particular interest to the lab and therapeutic relevance is exploiting the knowledge of chromatin components to wilfully regulate gene expression. We have three disease models and research projects that require further investigation; (Project I) understanding transcriptional competence and the relevance of the histone code in heart disease (Project II) wilfully regulating cancer related genes using engineered transcription factors (Project III) switching on the mental retardation gene in fragile X individuals. We have the molecular tools at hand to study chromatin modifications and candidates will be taught basic and advanced molecular techniques to kick-start each project.
3. When activators bind transcriptional repressors
   
   DNA methylation is a major determinant in the epigenetic silencing of genes. The mechanisms underlying its targeting and subsequent repression of transcription are relevant to human development and disease. The primary aims of this project are to analyse the functional overlap between the methylation dependent transcriptional repressor, MeCP2 and the ATP-dependent chromatin remodeler and transcriptional activator, SWI/SNF. Among the activities that are dependent on methylation and histone deacetylation are the ATPase remodeling complexes that alter chromatin accessibility and transcriptional competence. We have studied chromatin activities that rely on methylation and deacetylation and have profiled their relevance by chromatin immunoprecipitation technique (ChIP) and in vivo reconstitution studies. The mechanisms underlying the correlation between DNA methylation and histone deacetylation have recently been elucidated for MeCP2 (the best characterized member of the methyl-CpG binding domain (MBD) proteins). Evidence suggests that MeCP2/MBD proteins are involved in the recruitment of co-repressor complexes and the assembly of chromatin on methylated DNA. However, the precise mechanism of MeCP2 repression on methylated DNA in the context of chromatin has been a hotly debated topic. This project investigates the role of DNA methylation and SWI/SNF repression using a variety of molecular and biochemical approaches to determine their cooperativity. As a result of these studies we expect to learn more about chromatin remodelling and the pathways of transcriptional regulation.

Keywords and abbreviations

ChIP; Chromatin ImmunoPrecipitation platform demonstrates definitive protein-DNA association in vivo Epigenetic; Modification of DNA (such as methylation) in the absence of genetic mutations.
FMR1; Fragile X Mental Retardation 1 gene responsible for the disease and is repressed by methylation.
FMRP; Fragile X Mental Retardation Protein is an RNA binding protein necessary for neuronal function.
FRAXA; Fragile X syndrome.
MBD; Family of Methyl-CpG Binding Domain proteins that bind methylated DNA and repress transcription.
MeCP2; Member of the MBD family and a powerful methylation specific repressor.
Methylation; Modification to DNA by the methylation of the cytosine residue in the CpG dinucleotide and is correlated with transcriptional repression.
Remodeling; Chromatin structure and function can be regulated by at least three strategies, (i) modification of (acetylation/deacetylation) of core histone proteins, (ii) differential association of non-histone proteins such as such as MeCP2/MBD and (iii) ATP-dependent movement of nucleosomes by determinants such as SWI/SNF.
siRNA ; Small interfering RNA (RNAi) a sensitive and useful tool used to knockdown gene expression.
SWI/SNF; ATP-dependent chromatin remodeling machine generally involved in transcriptional activation.

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Assessment Breakdown

Research Component: 100 points
Research Report (up to 10,000 words): 80%
Supervisor's Evaluation: 12.5%
Oral presentation: 7.5%

Coursework Component: 0 points
N/A

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Student Numbers

Number of places available:
3