Epigenetic changes can switch genes on or off and determine which proteins are transcribed. Epigenetics involves genetic control by factors other than an individual's DNAsequence. Changes in epigenetic factors can play a critical role in disease.
Epigenetic changes can switch genes on or off and determine which proteins are transcribed. Epigenetics involves genetic control by factors other than an individual's DNAsequence. Changes in epigenetic factors can play a critical role in disease.
Direitos autorais:
Attribution Non-Commercial (BY-NC)
Formatos disponíveis
Baixe no formato DOC, PDF, TXT ou leia online no Scribd
Epigenetic changes can switch genes on or off and determine which proteins are transcribed. Epigenetics involves genetic control by factors other than an individual's DNAsequence. Changes in epigenetic factors can play a critical role in disease.
Direitos autorais:
Attribution Non-Commercial (BY-NC)
Formatos disponíveis
Baixe no formato DOC, PDF, TXT ou leia online no Scribd
Education Citation: Simmons, D. (2008) Epigenetic influence and disease. Nature Education 1(1) The behavior of a person's genes doesn't just depend on the genes' DNA sequence--it's also affected by so-called epigenetic factors. Changes in these factors can play a critical role in disease. The external environment's effects upon genes can influence disease, and some of these effects can be inherited in humans. Studies investigating how environmental factors impact the genetics of an individual's offspring are difficult to design. However, in certain parts of the world in which social systems are highly centralized, environmental information that might have influenced families can be obtained. For example, Swedish scientists recently conducted investigations examining whether nutrition affected the death rate associated with cardiovascular disease and diabetes and whether these effects were passed from parents to their children and grandchildren (Kaati et al., 2002). These researchers estimated how much access individuals had to food by examining records of annual harvests and food prices in Sweden across three generations of families, starting as far back as the 1890s. These researchers found that if a father did not have enough food available to him during a critical period in his development just before puberty, his sons were less likely to die from cardiovascular disease. Remarkably, death related to diabetes increased for children if food was plentiful during thiscritical period for the paternal grandfather, but it decreased when excess food was available to the father. These findings suggest that diet can cause changes to genes that are passed down though generations by the males in a family, and that these alterations can affect susceptibility to certain diseases. But what are these changes, and how are they remembered? The answers to questions such as these lie in the concept of epigenetics. What Is Epigenetics? How Do Epigenetic Changes Affect Genes?
Figure 1: Interaction between RNA, histone modification and DNA
methylation in heritable gene silencing. Epigenetics involves genetic control by factors other than an individual's DNAsequence. Epigenetic changes can switch genes on or off and determine which proteins are transcribed. Epigenetics is involved in many normal cellular processes. Consider the fact that our cells all have the same DNA, but our bodies contain many different types of cells: neurons, liver cells, pancreatic cells, inflammatory cells, and others. How can this be? In short, cells, tissues, and organs differ because they have certain sets of genes that are "turned on" or expressed, as well as other sets that are "turned off" or inhibited. Epigenetic silencing is one way to turn genes off, and it can contribute to differential expression. Silencing might also explain, in part, why genetic twins are not phenotypically identical. In addition, epigenetics is important for X-chromosome inactivation in female mammals, which is necessary so that females do not have twice the number of X-chromosome gene products as males (Egger et al., 2004). Thus, the significance of turning genes off via epigenetic changes is readily apparent. Within cells, there are three systems that can interact with each other to silence genes: DNA methylation, histone modifications, and RNA-associatedsilencing (Figure 1; Egger et al., 2004). DNA Methylation DNA methylation is a chemical process that adds a methyl group to DNA. It is highly specific and always happens in a region in which a cytosinenucleotide is located next to a guanine nucleotide that is linked by a phosphate; this is called a CpG site (Egger et al., 2004; Jones & Baylin, 2002; Robertson, 2002). CpG sites are methylated by one of three enzymes called DNA methyltransferases (DNMTs) (Egger et al., 2004; Robertson, 2002). Inserting methyl groups changes the appearance and structure of DNA, modifying a gene's interactions with the machinery within a cell's nucleus that is needed for transcription. DNA methylation is used in some genes to differentiate which gene copy is inherited from the father and which gene copy is inherited from the mother, a phenomenon known as imprinting. Histone Modifications Histones are proteins that are the primary components of chromatin, which is the complex of DNA and proteins that makes up chromosomes. Histonesact as a spool around which DNA can wind. When histones are modified after they are translated into protein (i.e., post-translation modification), they can influence how chromatin is arranged, which, in turn, can determine whether the associated chromosomal DNA will be transcribed. If chromatin is not in a compact form, it is active, and the associated DNA can be transcribed. Conversely, if chromatin is condensed (creating a complex calledheterochromatin), then it is inactive, and DNA transcription does not occur. There are two main ways histones can be modified: acetylation and methylation. These are chemical processes that add either an acetyl or methyl group, respectively, to the amino acid lysine that is located in the histone. Acetylation is usually associated with active chromatin, while deacetylation is generally associated with heterochromatin. On the other hand, histone methylation can be a marker for both active and inactive regions of chromatin. For example, methylation of a particular lysine (K9) on a specific histone (H3) that marks silent DNA is widely distributed throughout heterochromatin. This is the type of epigenetic change that is responsible for the inactivated X chromosome of females. In contrast, methylation of a different lysine (K4) on the same histone (H3) is a marker for active genes (Egger et al., 2004). RNA-Associated Silencing Genes can also be turned off by RNA when it is in the form of antisense transcripts, noncoding RNAs, or RNA interference. RNA might affect geneexpression by causing heterochromatin to form, or by triggering histone modifications and DNA methylation (Egger et al., 2004). Epigenetics and Disease: Some Examples While epigenetic changes are required for normal development and health, they can also be responsible for some disease states. Disrupting any of the three systems that contribute to epigenetic alterations can cause abnormal activation or silencing of genes. Such disruptions have been associated withcancer, syndromes involving chromosomal instabilities, and mental retardation (Table 1).
Epigenetics and Cancer
The first human disease to be linked to epigenetics was cancer, in 1983. Researchers found that diseased tissue from patients with colorectal cancerhad less DNA methylation than normal tissue from the same patients (Feinberg & Vogelstein, 1983). Because methylated genes are typically turned off, loss of DNA methylation can cause abnormally high gene activation by altering the arrangement of chromatin. On the other hand, too much methylationcan undo the work of protective tumor suppressor genes. As previously mentioned, DNA methylation occurs at CpG sites, and a majority of CpG cytosines are methylated in mammals. However, there are stretches of DNA near promoter regions that have higher concentrations of CpG sites (known as CpG islands) that are free of methylation in normal cells. These CpG islands become excessively methylated in cancer cells, thereby causing genes that should not be silenced to turn off. This abnormality is the trademark epigenetic change that occurs in tumors and happens early in the development of cancer (Egger et al., 2004; Robertson, 2002; Jones & Baylin, 2002). Hypermethylation of CpG islands can cause tumors by shutting off tumor-suppressor genes. In fact, these types of changes may be more common in human cancer than DNA sequence mutations (Figure 2). Furthermore, although epigenetic changes do not alter the sequence of DNA, they can cause mutations. About half of the genes that cause familial or inherited forms of cancer are turned off by methylation. Most of these genes normally suppress tumor formation and help repair DNA, including O6-methylguanine-DNA methyltransferase (MGMT), MLH1 cyclin-dependent kinase inhibitor 2B (CDKN2B), and RASSF1A. For example, hypermethylation of the promoter of MGMT causes the number of G-to-A mutations to increase (Figure 2). Hypermethylation can also lead to instability of microsatellites, which are repeated sequences of DNA. Microsatellites are common in normal individuals, and they usually consist of repeats of the dinucleotide CA. Too much methylation of the promoter of the DNA repair gene MLH1 can make a microsatelliteunstable and lengthen or shorten it (Figure 2). Microsatellite instability has been linked to many cancers, including colorectal, endometrial, ovarian, and gastric cancers (Jones & Baylin, 2002).
Figure 2: Mechanism of action of nucleoside analogue inhibitors.
Metaphase chromosomes showing the peculiar constriction at the end of the long arm of chromosome X that is characteristic in fragile X (FX) individuals. The black arrowhead marks the marker X chromosome in the upper right hand quadrant. Fragile X syndrome is the most frequently inherited mental disability, particularly in males. Both sexes can be affected by this condition, but because males only have one X chromosome, one fragile X will impact them more severely. Indeed, fragile X syndrome occurs in approximately 1 in 4,000 males and 1 in 8,000 females. People with this syndrome have severe intellectual disabilities, delayed verbal development, and "autistic-like" behavior (Penagarikano et al., 2007). Fragile X syndrome gets its name from the way the part of the X chromosomethat contains the gene abnormality looks under a microscope; it usually appears as if it is hanging by a thread and easily breakable (Figure 3). Thesyndrome is caused by an abnormality in the FMR1 (fragile X mental retardation 1) gene. People who do not have fragile X syndrome have 6 to 50 repeats of the trinucleotide CGG in their FMR1 gene. However, individuals with over 200 repeats have a full mutation, and they usually show symptoms of the syndrome. Too many CGGs cause the CpG islands at the promoterregion of the FMR1 gene to become methylated; normally, they are not. Thismethylation turns the gene off, stopping the FMR1 gene from producing an important protein called fragile X mental retardation protein. Loss of this specific protein causes fragile X syndrome. Although a lot of attention has been given to the CGG expansion mutation as the cause of fragile X, the epigenetic change associated with FMR1 methylation is the real syndromeculprit. Fragile X syndrome is not the only disorder associated with mental retardation that involves epigenetic changes. Other such conditions include Rubenstein-Taybi, Coffin-Lowry, Prader- Willi, Angelman, Beckwith-Wiedemann, ATR-X, and Rett syndromes (Table 1). Combating Diseases with Epigenetic Therapy Because so many diseases, such as cancer, involve epigenetic changes, it seems reasonable to try to counteract these modifications with epigenetic treatments. These changes seem an ideal target because they are by nature reversible, unlike DNA sequence mutations. The most popular of these treatments aim to alter either DNA methylation or histone acetylation. Inhibitors of DNA methylation can reactivate genes that have been silenced. Two examples of these types of drugs are 5-azacytidine and 5-aza-2′-deoxycytidine (Egger et al., 2004). These medications work by acting like the nucleotide cytosine and incorporating themselves into DNA while it is replicating. After they are incorporated into DNA, the drugs block DNMT enzymes from acting, which inhibits DNA methylation. Drugs aimed at histone modifications are called histone deacetylase (HDAC) inhibitors. HDACs are enzymes that remove the acetyl groups from DNA, which condenses chromatin and stops transcription. Blocking this process with HDAC inhibitors turns on gene expression. The most common HDAC inhibitors include phenylbutyric acid, SAHA, depsipeptide, and valproic acid (Egger et al., 2004). Caution in using epigenetic therapy is necessary because epigenetic processes and changes are so widespread. To be successful, epigenetic treatments must be selective to irregular cells; otherwise, activating gene transcription in normal cells could make them cancerous, so the treatments could cause the very disorders they are trying to counteract. Despite this possible drawback, researchers are finding ways to specifically target abnormal cells with minimal damage to normal cells, and epigenetic therapy is beginning to look increasingly promising..
Epistasis: Gene Interaction and the Phenotypic Expression of
cerevisiae of cell processes (e.g., mitosis) anddiseases (e.g., cancer Used by Gregor Mendel to describe Pisum sativum Pea plant patterns of inheritance
Employed in a wide variety of studies
ranging from early genemapping Drosophila Fruit fly via linkage and recombination studies, to melanogaster large scalemutant screens to identify genes related to specific biological functions
Valuable for studying the development
Caenorhabditis Roundworm simple nervous systems and elegans (nematode) the aging process
Used for mapping and identifying genes
Danio rerio Zebra fish involved in organdevelopment
Gene-transfer approaches are particularly useful when a disease-
associated mutation encodes a protein with decreasedfunction, called a loss-of-function mutation; in this case, normal cellular function is restored when a wild-type copy of thegene is introduced, because loss-of-function mutations are usually recessive. However, human disease can also be associated with dominant mutations in genes that encode hyperactive proteins, called gain-of-function mutations. Furthermore, human disease can be associated with dominant mutations in which the mutant proteins interfere with thefunction of wild- type proteins, called dominant negative mutations. In the case of a dominant mutation, introduction of the corresponding wild-type gene is usually not sufficient to rescue the disease-associated phenotype(s); rather, researchers would prefer to "turn off" expression of the mutant gene, or to inhibit the function of the mutant protein it encodes. To achieve this goal, researchers have turned their attention to RNA-based approaches, which target the RNA(either pre-mRNA or mRNA) transcribed from the dominant negative gene and effectively inhibit expression of the mutant protein. Figure 2 shows examples of RNA-based strategies for the treatment of disease. Five approaches have been used experimentally to modify RNA levels: antisense oligonucleotides (ASO), RNA interference (RNAi), trans-splicing, segmental trans-splicing, and ribozymes. ASO strategies use short single-stranded DNA (ssDNA) molecules, usually between 18 and 30 bases long, which are complementary to the mRNA to be targeted (Figure 2a). The ssDNA binds to the target mRNA, and the resulting DNA- RNA hybrid molecule is then degraded by the intracellular enzymeribonuclease H (RNase H). RNAi involves the use of double-stranded RNA molecules (dsRNA), typically 22 base pairs long, corresponding to a region of the target gene (Figure 2b). The dsRNA is processed within the cell in such a way that it becomes part of an RNA- induced silencing complex (RISC) that recognizes and degrades the corresponding target mRNA. Trans-splicing is a gene-transfer approach that targets a pre-mRNA containing a disease-associated mutation within one of its exons (Figure 2c). In this case, the transgene is used to replace the exon carrying the disease-associated mutation (exon C* in Figure 2c) with a wild-type copy of the exon. Thetransgene contains a hybridization domain, which is complementary to a region of the 5′ flanking intron between the donor and branch-point sites for RNAsplicing, followed by the splicing branch point, the splice acceptor site, the wild- type exon sequence, and the rest of the gene. Trans-splicing leads to the production of a wild-type copy of the mature mRNA and thus a corresponding wild-type protein. Segmental trans-splicing is an approach used to get around the size limitations associated with gene-transfer methods that involve vectors. (Sometimes, a given cDNA is too large to be carried within a single viral vector.) In this case, the gene is divided into two smaller pieces, which are delivered together using two separate gene-transfer vectors (Figure 2d). The vector carrying the second half of the gene includes a hybridization domain complementary to anintron located at the 3′ end of the first half of the gene, similar to that described for trans-splicing. In this case, trans-splicing leads to the production of a mature mRNA encoding the full length of the wild- type protein of interest. Ribozymes are RNA molecules with inherent catalytic activity that recognize a particular mRNA and cleave it (Figure 2e). Ribozymes containing ahybridization domain followed by a ribozyme nucleolytic motif that recognizes a target mRNA and the corresponding wild-type gene sequence can be used to selectively cleave a target mRNA that contains a mutation after the ribozyme cleavage site. Once the target gene is cleaved, the ribozyme-derivedhybridization motif binds, and RNA splicing leads to the formation of a wild-type copy of the mature mRNA. Many of these RNA-based strategies have been developed in recent years, and numerous questions remain regarding the cellular mechanisms involved inmRNA targeting. Furthermore, researchers must exercise caution with respect to the specificity of any given mRNA-targeting approach, whether ASO, RNAi, trans-splicing, or ribozyme based, to ensure that only the mRNA of interest is targeted. Stem Cell Therapy On the genetic horizon, the modern-day equivalent of organ transplantation is likely to be the use of stem cell therapy. Unlike organs, which are built of specialized mature cells with tissue- specific characteristics and a very limited ability to divide, stem cells are immature cells that have not yet specialized and that have the capacity to divide and mature into a wide variety of tissue types. Stem cells naturally occur in two forms: embryonic stem cells and somatic stem cells. Embryonic stem cells are derived from a specific group of cells within an embryo. They are capable of unrestricted cell divisions (i.e., they are immortal) and are pluripotent, which means that they are able to become nearly anycell type imaginable as long as they are provided with the appropriate environment. Ethical concerns regarding the use of human embryos as a source ofstem cells, as well as technical difficulties in obtaining and culturing these cells, have hampered the clinical use of embryonic stem cells. Somatic stem cells are derived from a specific group of cells within an adult tissue that serve to renew the tissue cell populations over time. Unlike pluripotent cells, somatic stem cells are more restricted in terms of the type of cells they can become after they divide; their fate depends upon the tissue type from which they are derived. Figure 3 shows the differences between embryonic stem cells and somatic stem cells in terms of how they are derived and their ability to differentiate.
One of the most exciting breakthroughs in stem cell research
occurred recently and has been reproduced in labs across the world: the ability to convert somatic cells into pluripotent stem cells (Takahashi et al., 2007; Yu et al., 2007; Lowry et al., 2008; Park et al., 2008b). Scientists have identified a set of four genes (OCT4/POU5F1, SOX2, KLF4, and c-MYC/MYC) that encode transcription factors that, when expressed at the same time, can convert skin cells (dermal fibroblasts) taken from an adult human into induced pluripotent stem cells (iPS cells) that are phenotypically indistinguishable from human embryonic stem cells in terms of their gene expression, cell surface markers, and cellular morphology. Like human embryonic stem cells, the iPS cells are immortal, are pluripotent, and express genes characteristic of all three embryonic germ cell layers (endoderm, ectoderm, and mesoderm) when induced to differentiate. Both opponents and proponents of human stem cell research have warmly welcomed the promise of somatic cell–derived iPS cell lines. The ability to produce iPS cell lines from somatic cells was heralded as an invaluable tool for understanding the underlying mechanisms associated withdisease and for developing novel approaches to the treatment of human disease. Recently, a team of researchers at Harvard University applied this technique and established a panel of human disease–specific iPS cell lines (Park et al., 2008a); in this case, the investigators expressed three (OCT4/POU5F1, SOX2, and KLF4), four (OCT4/POU5F1, SOX2, KLF4, and c-MYC/MYC), or five (OCT4/POU5F1, SOX2, KLF4, c- MYC/MYC andNANOG) transcription factor genes. To generate disease-specific iPS cell lines, researchers collected skin cells and bone marrow–derived mesenchymal cells from patients with one of ten different diseases, including adenosine deaminase deficiency-related severe combined immunodeficiency (ADA-SCID), Shwachman-Bodian-Diamondsyndrome (SBDS), Gaucher's disease (GD) type III, Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD), Parkinson's disease (PD), Huntington's disease (HD), juvenile- onset type 1 diabetes mellitus (JDM), Down syndrome/trisomy 21 (DS), and Lesch-Nyhan syndrome (carrier). Similar to the original iPS cell lines, the disease-specific iPS cell lines were immortal, pluripotent, and capable of expressing genes corresponding to all three embryonic cell layers when induced to differentiate (Park et al., 2008a). Table 2 summarizes the panel of disease-specific iPS cell lines and provides details about the corresponding mutated somatic cell types from which they were derived, as well as the age and sex of the somatic cell donor (Park et al., 2008a). These cell lines are freely accessible to researchers worldwide, and they will serve as a strong foundation for future studies aimed at the eradication of these devastating diseases. Due to the viral-based methods used to deliver the transcription factor genes into the somatic cells, the resulting iPS cell lines cannot currently be used to treat human patients. Nevertheless, thesecell lines are an invaluable resource for future investigation. With these disease-specific iPS cell lines in hand, researchers will be able to carry out experiments to better understand disease pathology and to develop effective gene-transfer techniques, RNA-based therapies, and drug-screening approaches to target disease phenotypes. Table 2: iPS Cells Derived from Somatic Cells of Patients with Genetic Disease.Reproduced from Park et al., 2008a Looking Ahead: Gene-Inspired Drug Design and Multimodal Therapies Researchers' ever-increasing knowledge of human genes and their disease-associated mutations has inspired new approaches to drug design and discovery. By understanding the underlying molecular mechanisms linked to disease, investigators can better target the activities of the enzymes, cellsurface receptors, secreted proteins, intracellular signaling proteins, and transcription factors that regulate disease-associated phenotypes. As gene-based therapeutics continue to evolve, multimodal approaches to human disease will emerge. The future of genetic medicine will require collaboration and multidisciplinary approaches, which will most certainly be accompanied by unexpected, life-changing discoveries.