Highlights • Impressive advances in NGS have enabled an immense diversity of novel applications. • The barrier of the $1000 genome has recently been broken. • Important novel tools for clinical diagnostics based on NGS are appearing. • Third-generation technologies may further revolutionize genomics research. • Significant challenges for NGS remain, in particular data storage and processing.
Highlights • ncRNAs constitute the majority of transcriptional output of the human genome. • Lack of functional knowledge of most of ncRNAs has led to classification with a number of issues. • A new classification is emerging rooted in unbiased whole-genome surveys of RNA. • The new classification should allow for easy data integration across multiple experiments.
Highlights • Most human genes are expressed in all tissues to some extent • Housekeeping genes may be defined by constant level of expression across tissues • We use RNA-seq data to provide a list of 3804 human housekeeping genes • Several exceptionally uniform genes are suggested as control genes for RT-PCR.
The emerging field of speciation genomics is advancing our understanding of the evolution of reproductive isolation from the individual gene to a whole-genome perspective. In this new view it is important to understand the conditions under which ‘divergence hitchhiking’ associated with the physical linkage of gene regions, versus ‘genome hitchhiking’ associated with reductions in genome-wide rates of gene flow caused by selection, can enhance speciation-with-gene-flow. We describe here a theory predicting four phases of speciation, defined by changes in the relative effectiveness of divergence and genome hitchhiking, and review empirical data in light of the theory. We outline future directions, emphasizing the need to couple next-generation sequencing with selection, transplant, functional genomics, and mapping studies. This will permit a natural history of speciation genomics that will help to elucidate the factors responsible for population divergence and the roles that genome structure and different forms of hitchhiking play in facilitating the genesis of new biodiversity.
In 2012, a new feature of eukaryotic gene expression emerged: ubiquitous expression of circular RNA (circRNA) from genes traditionally thought to express messenger or linear noncoding (nc)RNA only. CircRNAs are covalently closed, circular RNA molecules that typically comprise exonic sequences and are spliced at canonical splice sites. This feature of gene expression was first recognized in humans and mouse, but it quickly emerged that it was common across essentially all eukaryotes studied by molecular biologists. CircRNA abundance, and even which alternatively spliced circRNA isoforms are expressed, varies by cell type and can exceed the abundance of the traditional linear mRNA or ncRNA transcript. CircRNAs are enriched in the brain and increase in abundance during fetal development. Together, these features raise fundamental questions regarding the regulation of circRNA in cis and in trans , and its function.
Highlights • Nuclear localised lncRNAs regulate the expression of both local and distal genes. • lncRNAs can function locally to regulate enhancer–promoter interactions. • lncRNAs can interact with chromatin at many different locations genome wide. • RNA–protein–DNA and RNA–DNA interactions guide lncRNAs to their target sites.
If one accepts that the fundamental pursuit of genetics is to determine the genotypes that explain phenotypes, the meteoric increase of DNA sequence information applied toward that pursuit has nowhere to go but up. The recent introduction of instruments capable of producing millions of DNA sequence reads in a single run is rapidly changing the landscape of genetics, providing the ability to answer questions with heretofore unimaginable speed. These technologies will provide an inexpensive, genome-wide sequence readout as an endpoint to applications ranging from chromatin immunoprecipitation, mutation mapping and polymorphism discovery to noncoding RNA discovery. Here I survey next-generation sequencing technologies and consider how they can provide a more complete picture of how the genome shapes the organism.
Highlights • The zebrafish is a key genetic model system for vertebrate regeneration research. • Toolsets continue to evolve for studies of zebrafish appendage, heart, and neural regeneration. • Regeneration concepts and mechanisms in zebrafish have implications for mammals.
Highlights • DNA methylation regulates alternative splicing of mRNA precursors. • Exons have higher levels of DNA methylation compared to flanking introns. • DNA methylation can influence alternative splicing by two different mechanisms.
The DNA of each cell is wrapped around histone octamers, forming so-called ‘nucleosomal core particles’. These histone proteins have tails that project from the nucleosome and many residues in these tails can be post-translationally modified, influencing all DNA-based processes, including chromatin compaction, nucleosome dynamics, and transcription. In contrast to those present in histone tails, modifications in the core regions of the histones had remained largely uncharacterised until recently, when some of these modifications began to be analysed in detail. Overall, recent work has shown that histone core modifications can not only directly regulate transcription, but also influence processes such as DNA repair, replication, stemness, and changes in cell state. In this review, we focus on the most recent developments in our understanding of histone modifications, particularly those on the lateral surface of the nucleosome. This region is in direct contact with the DNA and is formed by the histone cores. We suggest that these lateral surface modifications represent a key insight into chromatin regulation in the cell. Therefore, lateral surface modifications form a key area of interest and a focal point of ongoing study in epigenetics.
Understanding the mechanisms of evolution requires information on the rate of appearance of new mutations and their effects at the molecular and phenotypic levels. Although procuring such data has been technically challenging, high-throughput genome sequencing is rapidly expanding knowledge in this area. With information on spontaneous mutations now available in a variety of organisms, general patterns have emerged for the scaling of mutation rate with genome size and for the likely mechanisms that drive this pattern. Support is presented for the hypothesis that natural selection pushes mutation rates down to a lower limit set by the power of random genetic drift rather than by intrinsic physiological limitations, and that this has resulted in reduced levels of replication, transcription, and translation fidelity in eukaryotes relative to prokaryotes.
Nuclear bodies including nucleoli, Cajal bodies, nuclear speckles, Polycomb bodies, and paraspeckles are membraneless subnuclear organelles. They are present at steady-state and dynamically respond to basic physiological processes as well as to various forms of stress, altered metabolic conditions and alterations in cellular signaling. The formation of a specific nuclear body has been suggested to follow a stochastic or ordered assembly model. In addition, a seeding mechanism has been proposed to assemble, maintain, and regulate particular nuclear bodies. In coordination with noncoding RNAs, chromatin modifiers and other machineries, various nuclear bodies have been shown to sequester and modify proteins, process RNAs and assemble ribonucleoprotein complexes, as well as epigenetically regulate gene expression. Understanding the functional relationships between the 3D organization of the genome and nuclear bodies is essential to fully uncover the regulation of gene expression and its implications for human disease.
The development of deep sequencing has enabled the identification of novel microRNAs (miRNAs), leading to a growing appreciation for the fact that individual miRNAs can be heterogeneous in length and/or sequence. These variants, termed isomiRs, can be expressed in a cell-specific manner, and numerous recent studies suggest that at least some isomiRs may affect target selection, miRNA stability, or loading into the RNA-induced silencing complex (RISC). Reports indicating differential functionality for isomiRs are currently confined to several specific variants, and although isomiRs are common, their broader biological significance is yet to be fully resolved. Here we review the growing body of evidence suggesting that isomiRs have functional differences, of which at least some appear biologically relevant, and caution researchers to take miRNA isoforms into consideration in their experiments.
Single-cell sequencing provides information that is not confounded by genotypic or phenotypic heterogeneity of bulk samples. Sequencing of one molecular type (RNA, methylated DNA or open chromatin) in a single cell, furthermore, provides insights into the cell's phenotype and links to its genotype. Nevertheless, only by taking measurements of these phenotypes and genotypes from the same single cells can such inferences be made unambiguously. In this review, we survey the first experimental approaches that assay, in parallel, multiple molecular types from the same single cell, before considering the challenges and opportunities afforded by these and future technologies.
Genome instability contributes to cancer development and accelerates age-related pathologies as evidenced by a variety of congenital cancer susceptibility and progeroid syndromes that are caused by defects in genome maintenance mechanisms. DNA damage response (DDR) pathways that are mediated through the tumor suppressor p53 play an important role in the cell-intrinsic responses to genome instability, including a transient cell cycle arrest, senescence and apoptosis. Both senescence and apoptosis are powerful tumor-suppressive pathways preventing the uncontrolled proliferation of transformed cells. However, both pathways can potentially deplete stem and progenitor cell pools, thus promoting tissue degeneration and organ failure, which are both hallmarks of aging. p53 signaling is also involved in mediating non-cell-autonomous interactions with the innate immune system and in the systemic adjustments during the aging process. The network of p53 target genes thus functions as an important regulator of cancer prevention and aging.
Histone modifications are key components of chromatin packaging but whether they constitute a ‘code’ has been contested. We believe that the central issue is causality: are histone modifications responsible for differences between chromatin states, or are differences in modifications mostly consequences of dynamic processes, such as transcription and nucleosome remodeling? We find that inferences of causality are often based on correlation and that patterns of some key histone modifications are more easily explained as consequences of nucleosome disruption in the presence of histone modifying enzymes. We suggest that the 35-year-old DNA accessibility paradigm provides a mechanistically sound basis for understanding the role of nucleosomes in gene regulation and epigenetic inheritance. Based on this view, histone modifications and variants contribute to diversification of a chromatin landscape shaped by dynamic processes that are driven primarily by transcription and nucleosome remodeling.
Highlights • Several weeds worldwide have evolved resistance in response to intense herbicide use. • Single-locus mutations and complex multigenic stress responses underlie resistance. • Large population sizes and standing genetic variation likely cause rapid evolution. • Future research should integrate genomic data, evolutionary ecology, and modeling.
Highlights • RNA sequencing uncovers mechanisms regulating gene expression. • Use of alternative TSSs, PASs, and exons is the rule. • Alternative translation initiation at 5′-UTRs and downstream codons is widespread. • Transcription, RNA processing, and translation are often interdependent processes.
DNA double-strand breaks are normal consequences of cell division and differentiation and must be repaired faithfully to maintain genome stability. Two mechanistically distinct pathways are known to efficiently repair double-strand breaks: homologous recombination and Ku-dependent non-homologous end joining. Recently, a third, less characterized repair mechanism, named microhomology-mediated end joining (MMEJ), has received increasing attention. MMEJ repairs DNA breaks via the use of substantial microhomology and always results in deletions. Furthermore, it probably contributes to oncogenic chromosome rearrangements and genetic variation in humans. Here, we summarize the genetic attributes of MMEJ from several model systems and discuss the relationship between MMEJ and ‘alternative end joining’. We propose a mechanistic model for MMEJ and highlight important questions for future research.
Spatial organization is an inherent property of the vertebrate genome to accommodate the roughly 2 m of DNA in the nucleus of a cell. In this nonrandom organization, topologically associating domains (TADs) emerge as a fundamental structural unit that is thought to guide regulatory elements to their cognate promoters. In this review we summarize the most recent findings about TADs and the boundary regions separating them. We discuss how the disruption of these structures by genomic rearrangements can result in gene misexpression and disease.