Abstract
This is an extraordinary international book with 33 chapters contributed by 84 researchers from the United States, Canada, India, Switzerland, Japan, Czech Republic, and Singapore, from universities, medical schools, governmental agencies (US Food and Drug Administration), research institutes, commercial entities (Novartis Institutes for Biomedical Research and Amgen), academies of science, and foundations. It is edited by Dr Saura C. Sahu from the Division of Toxicology, Center for Food Safety and Applied Nutrition, Food and Drug Administration, Laurel, Maryland, USA.
In the preface, Dr Sahu boldly states that the new epigenetics revolution has transformed toxicology, “an old discipline of science” and that this book “builds a bridge between toxicology and epigenetics” at the forefront of this research area. This book is a collaborative effort by international experts to present “up-to-date, state-of-the-art” information on “toxico-epigenetics” for scientists in this field as well as toxicologists, geneticists, medical practitioners, pharmacologists, drug and food scientists, and “federal regulators and safety assessors of drugs, food, environmental, and consumer products.”
This is the first edition of a book essentially introducing toxico-epigenomics to (likely) graduate students and researchers and experts in other branches of toxicology, so a brief introduction is warranted. As Dr Sahu indicates “Genetics is defined as the study of heritable changes in gene expression, caused by modifications in the base sequence of the gene itself.” But “genetics, environmental factors, and xenobiotics (all) contribute to toxicology and human disease.” Toxicogenomics integrates “traditional” toxicology and genomics, which results in consequences from the changes in genomic DNA. However, heritable gene expression is altered by modifications to DNA, which do not directly alter the genomic DNA base sequence itself; that is, epigenetics. These heritable epigenetic modifications include DNA methylation/demethylation, histone modifications, and noncoding small RNAs, that is, toxico-epigenomics. This is not a book for neophytes but for “investigators … actively engaged in this rapidly developing emerging new field of research” (chapter 1, Introduction).
The 33 chapters are eclectic and variable, focused or general, short or long, detailed, dense, and well referenced, with varying success, definitions, known mechanisms, such as lifetime epigenetic changes, transgenerational inheritance, epigenetic reprogramming, and elucidation of the known (or suspected or proposed) mechanisms of action of metals, chemicals such as diethylstilbestrol, bisphenol A, and/or carcinogenic metals. The mechanisms of DNA/cell transformation are not fully understood but likely include DNA adducts, DNA damage, oxidative stress, hormonal imbalance, altered cell growth, and disruption of cell/tissue/organ function (chapter 2). Other chapters cover DNA methylation and toxicogenomics (chapter 3), epigenetic marks on chromatin, with roles in diagnosis, treatment, therapy, and involving great complexity and flexibility (chapter 4), molecular epigenetic changes caused by environmental pollutants (chapter 5), epigenetic mediation of environmental exposures to polycyclic aromatic hydrocarbons (PAH; chapter 6), epigenetic changes from arsenic and arsenicals (chapters 7, 8, and 17), environmental epigenetics and asthma and allergy (chapter 9), microRNAs (miRNAs) in prostate cancer (chapter 10), epigenetics in cardiovascular health (chapter 11), epigenetics in autoimmunity (chapter 12), epigenetics in lupus (chapter 13), ocular epigenomics (chapter 14), nuclear RNA silencing (chapter 15), epigenetic biomarkers in cancer detection and diagnosis (chapter 16), DES and endocrine-disrupting chemicals (chapter 18), epigenomics and drug safety (chapter 19), archival toxico-epigenetics (chapter 20), nanoparticles and toxico-epigenomics (chapter 21), global epigenomic profiling (chapter 22), transcriptomics (chapter 23), histone tail modifications (chapter 24), epigenetic effects after radiation exposure (chapter 25), developmentally regulated gene expression (chapter 26), chromatin insulators (chapter 27), bioinformatics for high-throughput studies (chapter 28), computational methods in toxico-epigenomics (chapter 29), databases (chapter 30), epigenetics and carcinogenic risk assessment (chapter 31), epigenetic modification in chemical carcinogenesis (chapter 32), and application of cancer toxico-epigenomics in identifying high-risk populations (chapter 33).
Comments by individual chapter are as follows.
Chapter 1 by S. C. Sahu, as mentioned earlier, is an excellent introduction to toxico-epigenomics. Chapter 2 by Y. A. S. Cheng and W-y Tang on environment, epigenetics, and disease provides a definition of epigenetics and mechanisms of action and then focuses on environmental epigenetics and human disease from exposure to metals (chromium, cadmium, arsenic, nickel, lead, mercury), diethylstilbestrol (DES), bisphenol A (BPA), 2, 3, 7, 8-tetrachlorodobenzo-pdioxin, phthalate esters, polychlorinated biphenyls, disinfection byproducts, PAHs, and diet or living style. Finally, the authors discuss implications and future prospects for environmental epigenetics and present key questions to be answered (excellent idea and excellent question choices!).
Chapter 3 by D. Deobagkar, entitled DNA Methylation and Toxicogenomics is difficult to read in places and has grammatical errors that may be from those with written English language difficulties, but the content is still understandable and important. Dr Deobagkar makes the case for the use of toxicogenomics in identifying and elucidating adverse biological effects from environmental stressors, toxins, drugs, and other chemicals and the use of functional genomics, which directly measures phenotype, to provide a direct link between a specific gene and its expression and modification by exposures. One comment on chapter 3 content, in the discussion of dose and exposure (section 3.2 Toxicology), Dr Deobagkar does not include the necessity for providing the specifics of sex, age (stage of development), of the exposed organism, the timing, and the duration of the exposure or even the route of exposure. A toxicologist would be disappointed and surprised.
Chapter 4 by D. Quenet et al discusses epigenetic marks on chromatin with extensive detail in section 4.1.1 on DNA methylation, including a very complex figure 4.2 (section 4.1.2) on posttranslational histone modifications; figure 4.6 is also very detailed and complex.
Chapter 5 by S.S. Lewis et al on molecular epigenetic changes caused by environmental pollution was very good, with an excellent figure (figure 5.1), tracking environmental exposure to altered gene expression and chromatin remodeling, to epigenetic changes, to the physiological and to genetic consequences of these epigenetic changes. A number of chemicals are evaluated, including BPA, DDT, dioxin, 17alpha-ethinylestradiol, hexabromocyclododecane, methoxychlor, organochlorine pesticide mixtures, polybrominated diphenyl ethers, phthalates, vinclozolin, and metals, including cadmium, chromium, lead, methyl mercury, nickel, tungsten alloy, and zinc (note redundancy with chapter 2).
Chapter 6, by B. Sadikovic and D.I. Rodenhiser, examines epigenetic effects of PAHs (see also chapter 2).
Chapter 7 on epigenetic/epigenomic effects of environmental arsenicals, by P. L Severson and B. W. Futscher, focused on urothelial effects. Arsenic-induced changes to the epigenome by K. A. Bailey and R. C. Fry (chapter 8) focused on DNA methylation patterns. Both chapters were excellent, clear, thorough, and enlightening.
Chapter 17 also focused on arsenic but on the histone changes. If the book was meant to be an overview, 3 chapters on arsenic appear to be a bit excessive, although each focused on a different aspect, and there is a large amount of information available on arsenic poisoning.
Chapters 9 to 13 focus on human epigenetics/epigenomics and asthma and allergy (chapter 9 by S. Lovinsky-Desir and R. L. Miller), prostate cancer (chapter 10, by E. K. Amankwah and J. K. Park), cardiovascular health (chapter 11 by S. Ghosh and A. Baccarelli), autoimmunity (chapter 12 by C. A. Cooney and K. M. Gilbert), and lupus (chapter 13, by D. Ray and B. C. Richardson). The clinical importance of epigenetics/epigenomics for diagnosis, tracking of outcomes of treatments, and long-term outcome for all of these adverse effects is striking. This body of work documents the movement of this new scientific area from fascinating theoretical construct to patient treatment regimens.
Chapter 14 (by K. P. Mitton) focuses on ocular epigenomics as potential sites of environmental impact on development and disease, using disruption of DNA methylation and inhibition of histone acetylation in zebrafish eye development.
Chapter 15 (by R. Malik and P. Svoboda) discusses nuclear RNA silencing and related phenomena in animals. The authors begin by describing RNA silencing; small RNAs (20-30 nucleotides long) function as specific guides for ribonucloprotein complexes to silence transcription and thereby translation. The silencing mechanism has 3 basic steps (1) production of these small RNAs that typically uses RNase III and/or RNA-dependent RNA polymerase, (2) formation of an effector complex, involving loading the small RNAs on argonaute (AGO) proteins, and (3) sequence-specific target recognition and induction of silencing. RNA silencing involves posttranscriptional mechanisms such as RNA interference (RNAi) or miRNA pathways at the level of transcriptional repression. Genes that regulate chromatin structure are directly or indirectly regulated by miRNAs. The miRNAs also mediate transcriptional regulation (activation or repression). These processes are well documented and understood in Arabidopsis and Shizosaccharomyces and are well described in this chapter, but the role of miRNA in mammals still remains enigmatic.
Chapter 16 (by A. G. Rivenbark and W. B. Coleman) discusses epigenetic markers (DNA methylation) in detection and diagnosis of various cancers in humans. Chapter 17 (by J. F. Reichard and A. Puga), already mentioned, discusses the epigenetic histone changes in the toxicologic mode (not yet mechanism) of action of arsenic.
Chapter 18, by S. Miyagawa et al, discusses the irreversible effects of DES on reproductive organs and the argument for epigenetic effects of endocrine disrupting chemicals. They also discuss the challenges in integrating epigenetic analysis into “traditional” toxicity testing.
In a shift of focus, chapter 19, by H. Kempiannan, et al., entitled Epigenomics-Impact for Drug Safety Science, examines this relatively new area from the perspective of the dynamic epigenome and perturbations of disease. The 3 tables and 2 figures are excellent, informative, detailed, clear, and thorough. The thrust and conclusion of this chapter is that epigenomic profiling technologies have great potential for providing novel mechanistic insight and candidate biomarkers for drug efficacy and safety assessment during both preclinical and clinical phases of drug development.
Chapter 20 (by B. A. Merrick) examines the molecular analysis of modified DNA from preserved tissues in toxicology studies, although fixed tissues have not been viewed as particularly useful for biochemical or molecular analysis(the preference has been to use frozen tissues for retrospective molecular analyses). The improvements in extraction of intact DNA, RNA, miRNA, and protein from paraffinized tissues and use of amplification technologies have enabled use of fixed tissue archives to detect epigenetic modifications (“toxicomethylomics”). This chapter mainly discusses the processes to extract DNA, RNA, and protein and methods of analysis, a very different chapter from the others.
Nanoparticles and toxicoepigenomics are the topics of chapter 21 (by M. P. Jain et al). These nanoparticles include naturally occurring nanoparticles, anthropogenic (man-made) ultrafine particles, and synthetic-engineered nanoparticles. These particles are ubiquitous in the environment and have always been resulting in widespread exposure and “potential ecotoxicological effects.” The chapter discusses nanoparticles with respect to their presence in the environment and their subsequent biological and pathological consequences. Nanoparticles in soil, water, and air, medicine, biomedial research, and toxicology (nanotoxicology) can affect humans and experimental animals (as well as other organisms), thereby complicating nanotoxicological studies, from molecular mechanisms of nanoparticle toxicity and cellular (intracellular) defense mechanisms. Induction of reactive oxygen species and therefore oxidative stress by nanoparticles occurs early in the exposure and can instigate normal protective cellular effects (the first tier); in the second tier, oxidative stress overwhelms the cellular antioxidant system, and the resulting oxidative injury can result in activation of proinflammatory transcription factors early in inflammation. In the third tier, oxidative stress is so great that cytotoxicity and cell death occur. A fascinating chapter!
Chapter 22 (by M. W. Y. Chan et al) examines modulation of developmentally regulated gene expression through targeting of polycomb and bithorax group proteins in Drosophila. This is a very different chapter using the fruit fly (the “workhorse” of genetics) and the recognition that all cell types (in the fruit fly and everything else) may have (must have) originated from the initial identical primary DNA sequence but gives rise to cell-specific gene expression programs and differentiated cells. These cell-specific patterns of gene expression are established through hormone-, growth factor-, and cytokine-initiated signaling cascades transduced through transcription factors to regulate the expression of specific subsets of genes within the genome, within specific times, and spaces. These changes are transmitted to daughter cells, so cellular memory is maintained epigenetically through specific chromatin modifications. The authors suggest that targeting the polycomb and bithorax genes and their protein products and developing specific inhibitors to modulate these proteins in (first normal and then) diseased cells will enable therapeutic intervention (excellent chapter!).
Chapter 23 (by P. Joseph) discusses transcriptomics and its applications in epigenetic toxicology. He notes that if the intent is to determine the expression of a single gene or a limited number of genes in a biological sample, then use quantitative reverse transcriptase–polymerase chain reaction analysis; however, if the objective is to determine the expression of “several hundreds or thousands or all of the genes” expressed in a biological sample, then the microarray analysis is the method of choice. The chapter continues with descriptions of the microarray analysis process, study designs, isolation of the RNA, synthesis of the complementary DNA, target synthesis, hybridization, washing, image acquisition, data generation and analysis, and so on. This chapter is a clear, thorough description of the process, results, various analyses, interpretations, and the translation of transcriptomics data from experimental animals to humans.
Chapter 24 (by Y. Chervona and M. Costa) discusses metal carcinogenesis and makes a strong case for the carcinogenic metals, such as arsenic, nickel, hexavalent chromium, and cadmium, acting epigenetically by altering “normal” histone tail modifications.
Chapter 25 (by Y. Hirabayashi and T. Inoue) discusses prediction of epigenetic and stochastic gene expression profiles of late effects after radiation exposure, initially by comparing pathological profiling (diagnostic endpoint) and toxicological profiling (probabilistic endpoint). This is a very complex and detailed chapter.
Chapter 26 (by M. Brand and F. J. Dilworth) returns to polycomb and trithorax (see chapter 22 on polycomb and bithorax) and examines modulations of developmentally regulated gene expression programs through targeting of the proteins from this gene group. There is some overlap between this chapter and chapter 22. This chapter, by Brand and Dilworth, is a detailed meticulous description of the development of cell-specific gene expression programs from primary DNA sequences, regulated temporally and spatially by hormone-, growth factor-, and cytokine-initiated signaling cascades that are transduced through transcription factors to regulate specific subsets of genes within the genome. These cell-specific gene programs are also transmitted to daughter cells during proliferation by cellular memory. This cellular memory is maintained epigenetically through specific chromatin modifications. This chapter provides detailed and specific mechanisms for enhancement and/or suppression of developmentally important genes in Drosophila (such as Zeste and EED [extra sex comb]) through polycomb group proteins (table 26.1) and trithorax group proteins (table 26.2). The authors also make the case that, since the polycomb and trithorax proteins are defined by mutations in Drosophila, which give rise to homeotic transformations, it is not surprising that somatic mutations in the genes that code for these proteins are implicated in a number of diseases in humans such as leukemia, solid cancers such as lymphomas, and myelodysplastic syndromes.
Chapter 27 (by J. Yang and V. G. Corces) on chromatin insulators and epigenetic inheritance in health and disease describes chromatin insulators as DNA-bound protein complexes, originally discovered in Drosophila and later in vertebrates, which can mediate intra- and inter-chromosome interactions; they act by bringing together regulatory sequences, originally located at great distances from each other, and/or on separate chromosomes, to locations very close together, and thereby affecting the expression of adjacent genes. This can result in activation or repression of single gene or large chromosome domains. The insulators “can alter” the status of the chromatin in these genes or domains and thereby also modify patterns of epigenetic inheritance. These insulator-mediated interactions are viewed as contributing to the establishment of the “3-dimensional organization” of the nuclear DNA. Since this structure is directly related to gene expression, the authors argue that “the organization of the genome in the nucleus is in part a determinant and in part a consequence of the transcriptional status of a cell.” Insulator function is itself regulated by enhancing or preventing the establishment of a functional loop between the 2 genetic sites (one termed an enhancer and the other termed a promotor). This process is affected by hormones, can regulate viral genomes, and defects in its function will “likely” lead to abnormalities in gene expression and disease, including neurological/neurodegenerative disorders and cancer. A fascinating chapter!
Chapter 28 (by M. A. Sartor et al) focuses on bioinformatics for high-throughput toxico-epigenomics studies, with excellent initial commentary on environmental genomics, genomic imprinting, and a cogent evolutionary perspective.
Chapter 29 (by J. C. Tong) describes computational methods in toxico-epigenomics. Chapter 30 (by U. Mashankar and S. Gurunathan) describes databases and tools for computational epigenomics. Chapter 31 (by P. Nioi) describes the interface of epigenetics and carcinogenic risk assessment by reviewing case studies of epigenetic changes, specifically DNA methylation, reported in chemically induced cancers in humans and rodents by agents with well-defined mechanisms and pathways. Chapter 32 (by I. P. Porgrbny et al) is a synergistic chapter (with chapter 31) on epigenetic modifications in chemical carcinogenesis.
Chapter 33 (by M. Verna and K. Banaudha), the last brief chapter, introduces the application of cancer toxico-epigenomics in identifying high-risk population. The authors cogently argue that epigenetic marks are tissue specific and that “aberrant DNA methylation is a hallmark of cancer.”
Minor comments by this reviewer include the chapters are very variable in size (small to huge) and scope (some are “how to” while others are more broad, explanatory, and encompassing). Not all areas of epigenetics/epigenomics are discussed, while other areas are discussed in more than 1 chapter. There are also numerous grammatical errors likely from the international authors all writing in English. In addition, some abbreviations are not defined when first used or ever (e.g., “APL” on page 137, “DC” on page 199, etc.), and some figures are very (too) complex and/or poorly explained.
After reading this book, this reviewer came cross an article by MemczaK et al 1 in Nature, March 21, 2013, introducing circular RNA species in plants and animals, which act as posttranslational regulators. They are well-expressed, stable transcripts with robust expression, which function as miRNA’s sponges (letter from Hansen et al 2 ); they are circularized using splice genes. These circular RNAs can counteract the function of regulatory miRNAs and counteract the actions of competing endogenous RNAs. These circular RNAs can act as miRNA mimics, competing with other RNAs for bonding with the RNA-binding proteins or to miRNAs. The miRNAs are approximately 21 nucleotides long, noncoding RNAs that guide the effector protein AGO to messenger RNAs (mRNAs) of coding genes to repress their protein synthesis. In humans, miRNAs directly regulate expression of most mRNAs in a large range of biological functions. This newly discovered regulatory RNA adds another complex aspect to the RNA world (in News & Views from K.S. Kosik 3 ).
In the end, what is clearly emerging, based on this book (and other articles), is the varied and complex ways a given fixed genome in a given species is manipulated to result in very different outcomes at different times under different circumstances, including the first moves toward a new and different species, by various ingenious mechanisms.
All-in-all, this is an ambitious book covering a number of important aspects of a very new and very exciting area in toxicology, epigenetics, and epigenomics. I highly recommend it (warts and all) and look forward to the second edition.