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Immunodeficiency, centromeric instability and facial anomalies (ICF) syndrome is in most cases caused by mutations in either DNA methyltransferase (DNMT)3B, zinc finger and BTB domain containing 24, cell division cycle associated 7 or helicase lymphoid-specific. However, the causative genes of a few ICF patients remain unknown. We, herein, identified ubiquitin-like with plant homeodomain and really interesting new gene finger domains 1 (UHRF1) as a novel causative gene of one such patient with atypical symptoms. This patient is a compound heterozygote for two previously unreported mutations in UHRF1: c.886C > T (p.R296W) and c.1852C > T (p.R618X). The R618X mutation plausibly caused nonsense-mediated decay, while the R296W mutation changed the higher order structure of UHRF1, which is indispensable for the maintenance of CG methylation along with DNMT1. Genome-wide methylation analysis revealed that the patient had a centromeric/pericentromeric hypomethylation, which is the main ICF signature, but also had a distinctive hypomethylation pattern compared to patients with the other ICF syndrome subtypes. Structural and biochemical analyses revealed that the R296W mutation disrupted the protein conformation and strengthened the binding affinity of UHRF1 with its partner LIG1 and reduced ubiquitylation activity of UHRF1 towards its ubiquitylation substrates, histone H3 and proliferating cell nuclear antigen -associated factor 15 (PAF15). We confirmed that the R296W mutation causes hypomethylation at pericentromeric repeats by generating the HEK293 cell lines that mimic the patient's UHRF1 molecular context. Since proper interactions of the UHRF1 with LIG1, PAF15 and histone H3 are essential for the maintenance of CG methylation, the mutation could disturb the maintenance process. Evidence for the importance of the UHRF1 conformation for CG methylation in humans is, herein, provided for the first time and deepens our understanding of its role in regulation of CG methylation., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Recent studies have pointed out the essential role of genetic ancestry in population pharmacogenetics. In this study, we analyzed the whole-genome sequencing data from The 1000 Genomes Project (Phase 3) and the pharmacogenetic information from Drug Bank, PharmGKB, PharmaADME, and Biotransformation. Here we show that ancestry-informative markers are enriched in pharmacogenetic loci, suggesting that trans-ancestry differentiation must be carefully considered in population pharmacogenetics studies. Ancestry-informative pharmacogenetic loci are located in both protein-coding and non-protein-coding regions, illustrating that a whole-genome analysis is necessary for an unbiased examination over pharmacogenetic loci. Finally, those ancestry-informative pharmacogenetic loci that target multiple drugs are often a functional variant, which reflects their importance in biological functions and pathways. In summary, we develop an efficient algorithm for an ultrahigh-dimensional principal component analysis. We create genetic catalogs of ancestry-informative markers and genes. We explore pharmacogenetic patterns and establish a high-accuracy prediction panel of genetic ancestry. Moreover, we construct a genetic ancestry pharmacogenomic database Genetic Ancestry PhD ( http://hcyang.stat.sinica.edu.tw/databases/genetic_ancestry_phd/ ).

Chromatin Conformation Capture techniques have unveiled several layers of chromosome organization such as the segregation in compartments, the folding in topologically associating domains (TADs), and site-specific looping interactions. The discovery of this genome hierarchical organization emerged from the computational analysis of chromatin capture data. With the increasing availability of such data, automatic pipelines for the robust comparison, grouping, and classification of multiple experiments are needed. Here we present a pipeline based on the TADbit framework that emphasizes reproducibility, automation, quality check, and statistical robustness. This comprehensive modular pipeline covers all the steps from the sequencing products to the visualization of reconstructed 3D models of the chromatin.

The nucleolus is the largest compartment of the eukaryotic cell's nucleus. It acts as a ribosome factory, thereby sustaining the translation machinery. The nucleolus is also the subnuclear compartment with the highest transcriptional activity in the cell, where hundreds of ribosomal RNA (rRNA) genes transcribe the overwhelming majority of RNAs. The structure and composition of the nucleolus change according to the developmental state. For instance, in embryonic stem cells (ESCs), rRNA genes display a hyperactive transcriptional state and open chromatin structure compared with differentiated cells. Increasing evidence indicates that the role of the nucleolus and rRNA genes might go beyond the control of ribosome biogenesis. One such role is linked to the genome architecture, since repressive domains are often located close to the nucleolus. This review highlights recent findings describing how the nucleolus is regulated in ESCs and its role in regulating ribosome biogenesis and genome organization for the maintenance of stem cell identity., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Large-scale epigenomic projects have mapped hundreds of thousands of potential regulatory sites in the human genome, but only a small proportion of these elements are proximal to transcription start sites. It is believed that the majority of these sequences are remote promoter-activating genomic sites scattered within several hundreds of kilobases from their cognate promoters and referred to as enhancers. It is still unclear what principles, aside from relative closeness in the linear genome, determine which promoter(s) is controlled by a given enhancer; however, this understanding is of great fundamental and clinical relevance. In recent years, C-methods (chromosome conformation capture-based methods) have become a powerful tool for the identification of enhancer-promoter spatial contacts that, in most cases, reflect their functional link. Here, we describe a new hybridisation-based promoter Capture-C protocol that makes use of biotinylated dsDNA probes generated by PCR from a custom pool of long oligonucleotides. The described protocol allows high-resolution promoter interactome description, providing a flexible and cost-effective alternative to the existing promoter Capture-C modifications. Based on the obtained data, we propose several tips on probe design that could potentially improve the results of future experiments.

Nucleoli, the sites of ribosome biogenesis and the largest structures in human nuclei, form around nucleolar organizer regions (NORs) comprising ribosomal DNA (rDNA) arrays. NORs are located on the p-arms of the five human acrocentric chromosomes. Defining the rules of engagement between these p-arms and nucleoli takes on added significance as describing the three-dimensional organization of the human genome represents a major research goal. Here we used fluorescent in situ hybridization (FISH) and immuno-FISH on metaphase chromosomes from karyotypically normal primary and hTERT-immortalized human cell lines to catalog NORs in terms of their relative rDNA content and activity status. We demonstrate that a proportion of acrocentric p-arms in cell lines and from normal human donors have no detectable rDNA. Surprisingly, we found that all NORs with detectable rDNA are active, as defined by upstream binding factor loading. We determined the nucleolar association status of all NORs during interphase, and found that nucleolar association of acrocentric p-arms can occur independently of rDNA content, suggesting that sequences elsewhere on these chromosome arms drive nucleolar association. In established cancer lines, we characterize a variety of chromosomal rearrangements involving acrocentric p-arms and observe silent, rDNA-containing NORs that are dissociated from nucleoli. In conclusion, our findings indicate that within human nuclei, positioning of all 10 acrocentric chromosomes is dictated by nucleolar association. Furthermore, these nucleolar associations are buffered against interindividual variation in the distribution of rDNA., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Molecular endocrinology of vitamin D is based on the activation of the transcription factor vitamin D receptor (VDR) by the vitamin D metabolite 1α,25-dihydroxyvitamin D3. This nuclear vitamin D-sensing process causes epigenome-wide effects, such as changes in chromatin accessibility as well as in the contact of VDR and its supporting pioneer factors with thousands of genomic binding sites, referred to as vitamin D response elements. VDR binding enhancer regions loop to transcription start sites of hundreds of vitamin D target genes resulting in changes of their expression. Thus, vitamin D signaling is based on epigenome- and transcriptome-wide shifts in VDR-expressing tissues. Monocytes are the most responsive cell type of the immune system and serve as a paradigm for uncovering the chromatin model of vitamin D signaling. In this review, an alternative approach for selecting vitamin D target genes is presented, which are most relevant for understanding the impact of vitamin D endocrinology on innate immunity. Different scenarios of the regulation of primary upregulated vitamin D target genes are presented, in which vitamin D-driven super-enhancers comprise a cluster of persistent (constant) and/or inducible (transient) VDR-binding sites. In conclusion, the spatio-temporal VDR binding in the context of chromatin is most critical for the regulation of vitamin D target genes.

Our genome is a historic record of successive invasions of mobile genetic elements. Like other eukaryotes, we have evolved mechanisms to limit their propagation and minimize the functional impact of new insertions. Although these mechanisms are vitally important, they are imperfect, and a handful of retroelement families remain active in modern humans. This review introduces the intrinsic functions of transposons, the tactics employed in their restraint, and the relevance of this conflict to human pathology. The most straightforward examples of disease-causing transposable elements are germline insertions that disrupt a gene and result in a monogenic disease allele. More enigmatic are the abnormal patterns of transposable element expression in disease states. Changes in transposon regulation and cellular responses to their expression have implicated these sequences in diseases as diverse as cancer, autoimmunity, and neurodegeneration. Distinguishing their epiphenomenal from their pathogenic effects may provide wholly new perspectives on our understanding of disease.

Chronic lymphocytic leukemia is a common disease in Western countries and has heterogeneous clinical behavior. The relevance of the genetic basis of the disease has come to the forefront recently, with genome-wide studies that have provided a comprehensive view of structural variants, somatic mutations, and different layers of epigenetic changes. The mutational landscape is characterized by relatively common copy number alterations, a few mutated genes occurring in 10-15% of cases, and a large number of genes mutated in a small number of cases. The epigenomic profile has revealed a marked reprogramming of regulatory regions in tumor cells compared with normal B cells. All of these alterations are differentially distributed in clinical and biological subsets of the disease, indicating that they may underlie the heterogeneous evolution of the disease. These global studies are revealing the molecular complexity of chronic lymphocytic leukemia and provide new perspectives that have helped to understand its pathogenic mechanisms and improve the clinical management of patients.

The 3D organization of mammalian chromatin was described more than 30 years ago by visualizing sites of DNA synthesis at different times during the S phase of the cell cycle. These early cytogenetic studies revealed structurally stable chromosome domains organized into subnuclear compartments. Active-gene-rich domains in the nuclear interior replicate early, whereas more condensed chromatin domains that are largely at the nuclear and nucleolar periphery replicate later. During the past decade, this spatiotemporal DNA replication programme has been mapped along the genome and found to correlate with epigenetic marks, transcriptional activity and features of 3D genome architecture such as chromosome compartments and topologically associated domains. But the causal relationship between these features and DNA replication timing and the regulatory mechanisms involved have remained an enigma. The recent identification of cis-acting elements regulating the replication time and 3D architecture of individual replication domains and of long non-coding RNAs that coordinate whole chromosome replication provide insights into such mechanisms.

The programmes that direct an organism's development and maintenance are encoded in its genome. Decoding of this information begins with regulated transcription of genomic DNA into RNA. Although transcription and its control can be tracked indirectly by measuring stable RNAs, it is only by directly measuring nascent RNAs that the immediate regulatory changes in response to developmental, environmental, disease and metabolic signals are revealed. Multiple complementary methods have been developed to quantitatively track nascent transcription genome-wide at nucleotide resolution, all of which have contributed novel insights into the mechanisms of gene regulation and transcription-coupled RNA processing. Here we critically evaluate the array of strategies used for investigating nascent transcription and discuss the recent conceptual advances they have provided.

MicroRNAs form a class of short, non-coding RNA molecules which are essential for proper development in tissue-based plants and animals. To help explain their role in gene regulation, a number of mathematical and computational studies have demonstrated the potential canalizing effects of microRNAs. However, such studies have typically focused on the effects of microRNAs on only one or a few target genes. Consequently, it remains unclear how these small-scale effects add up to the experimentally observed developmental outcomes resulting from microRNA perturbation at the whole-genome level. To answer this question, we built a general computational model of cell differentiation to study the effect of microRNAs in genome-scale gene regulatory networks. Our experiments show that in large gene regulatory networks, microRNAs can control differentiation time without significantly changing steady-state gene expression profiles. This temporal regulatory role cannot be naturally replicated using protein-based transcription factors alone. While several microRNAs have been shown to regulate differentiation time in vivo , our findings provide a new explanation of how the cumulative molecular actions of individual microRNAs influence genome-scale cellular dynamics. Taken together, these results may help explain why tissue-based organisms exclusively depend on miRNA-mediated regulation, while their more primitive counterparts do not.

Differentiation between phenotypically neutral and disease-causing genetic variation remains an open and relevant problem. Among different types of variation, non-frameshifting insertions and deletions (indels) represent an understudied group with widespread phenotypic consequences. To address this challenge, we present a machine learning method, MutPred-Indel, that predicts pathogenicity and identifies types of functional residues impacted by non-frameshifting insertion/deletion variation. The model shows good predictive performance as well as the ability to identify impacted structural and functional residues including secondary structure, intrinsic disorder, metal and macromolecular binding, post-translational modifications, allosteric sites, and catalytic residues. We identify structural and functional mechanisms impacted preferentially by germline variation from the Human Gene Mutation Database, recurrent somatic variation from COSMIC in the context of different cancers, as well as de novo variants from families with autism spectrum disorder. Further, the distributions of pathogenicity prediction scores generated by MutPred-Indel are shown to differentiate highly recurrent from non-recurrent somatic variation. Collectively, we present a framework to facilitate the interrogation of both pathogenicity and the functional effects of non-frameshifting insertion/deletion variants. The MutPred-Indel webserver is available at http://mutpred.mutdb.org/., Competing Interests: The authors have declared that no competing interests exist.

Physical access to DNA is a highly dynamic property of chromatin that plays an essential role in establishing and maintaining cellular identity. The organization of accessible chromatin across the genome reflects a network of permissible physical interactions through which enhancers, promoters, insulators and chromatin-binding factors cooperatively regulate gene expression. This landscape of accessibility changes dynamically in response to both external stimuli and developmental cues, and emerging evidence suggests that homeostatic maintenance of accessibility is itself dynamically regulated through a competitive interplay between chromatin-binding factors and nucleosomes. In this Review, we examine how the accessible genome is measured and explore the role of transcription factors in initiating accessibility remodelling; our goal is to illustrate how chromatin accessibility defines regulatory elements within the genome and how these epigenetic features are dynamically established to control gene expression.

Systemic lupus erythematosus (SLE) is a systemic inflammatory autoimmune disease characterized by a broad spectrum of clinical and serological manifestations. This may reflect a complex and multifactorial etiology involving several identified genetic and environmental factors, though not explaining the full risk of SLE. Established SLE risk genotypes are either very rare or with modest effect sizes and twin studies indicate that other factors besides genetics must be operative in SLE etiology. The exposome comprises the cumulative environmental influences on an individual and associated biological responses through the lifespan. It has been demonstrated that exposure to silica, smoking and exogenous hormones candidate as environmental risk factors in SLE, while alcohol consumption seems to be protective. Very few studies have investigated potential gene-environment interactions to determine if some of the unexplained SLE risk is attributable hereto. Even less have focused on interactions between specific risk genotypes and environmental exposures relevant to SLE pathogenesis. Cohort and case-control studies may provide data to suggest such biological interactions and various statistical measures of interaction can indicate the magnitude of such. However, such studies do often have very large sample-size requirements and we suggest that the rarity of SLE to some extent can be compensated by increasing the ratio of controls. This review summarizes the current body of knowledge on gene-environment interactions in SLE. We argue for the prioritization of studies that comprise the increasing details available of the genome and exposome relevant to SLE as they have the potential to disclose new aspects of SLE pathogenesis including phenotype heterogeneity., (Copyright © 2019 The Author(s). Published by Elsevier B.V. All rights reserved.)

Progression through the cell cycle is driven by the activities of the cyclin-dependent kinase (CDK) family of enzymes, which establish an ordered passage through the cell cycle phases. CDK activity is crucial for the cellular transitions from G1 to S and G2 to M, which are highly controlled to promote the faithful duplication of the genetic material and the transmission of the genome into daughter cells, respectively. While oscillations in CDK activity are essential for cell division, how its specific dynamics may shape cellular processes remains an open question. Recently, we have investigated the potential role of CDK in establishing the profile of replication initiation along the chromosomes, also referred to as the replication program. Our results demonstrated that the timing and level of CDK activity at G1/S provide two critical and independent inputs that modulate the pattern of origin usage. In this review, we will present the conclusions of our study and discuss the implications of our findings for cellular function and physiology.

The genome is packaged and organized nonrandomly within the 3D space of the nucleus to promote efficient gene expression and to faithfully maintain silencing of heterochromatin. The genome is enclosed within the nucleus by the nuclear envelope membrane, which contains a set of proteins that actively participate in chromatin organization and gene regulation. Technological advances are providing views of genome organization at unprecedented resolution and are beginning to reveal the ways that cells co-opt the structures of the nuclear periphery for nuclear organization and gene regulation. These genome regulatory roles of proteins of the nuclear periphery have important influences on development, disease and ageing.

Inherited mitochondrial DNA (mtDNA) diseases were discovered 30 years ago, and their characterization has provided a new perspective on the etiology of the common metabolic and degenerative diseases, cancer, and aging. The maternally inherited mtDNA contains 37 critical bioenergetic genes that are present in hundreds of copies per cell, but the 'mitochondrial genome' encompasses an additional 1,000-2,000 nuclear DNA (nDNA) mitochondrial genes. The interaction between these two mitochondrial genetic systems provides explanations for phenomena such as the non-Mendelian transmission of the common 'complex' diseases, age-related disease risk and progression, variable penetrance and expressivity, and gene-environment interactions. Thus, mtDNA genetics contributes to the quantitative and environmental components of human genetics that cannot be explained by Mendelian genetics. Because mtDNA is maternally inherited and cytoplasmic, it has fostered the first germline gene therapy, nuclear transplantation. However, effective interventions are still lacking for existing patients with mitochondrial dysfunction.

Studies of 3D chromatin organization have suggested that chromosomes are hierarchically organized into large compartments composed of smaller domains called topologically associating domains (TADs). Recent evidence suggests that compartments are smaller than previously thought and that the transcriptional or chromatin state is responsible for interactions leading to the formation of small compartmental domains in all organisms. In vertebrates, CTCF forms loop domains, probably via an extrusion process involving cohesin. CTCF loops cooperate with compartmental domains to establish the 3D organization of the genome. The continuous extrusion of the chromatin fibre by cohesin may also be responsible for the establishment of enhancer-promoter interactions and stochastic aspects of the transcription process. These observations suggest that the 3D organization of the genome is an emergent property of chromatin and its components, and thus may not be only a determinant but also a consequence of its function.

Repetitive DNA, formerly referred to by the misnomer "junk DNA," comprises a majority of the human genome. One class of this DNA, alpha satellite, comprises up to 10% of the genome. Alpha satellite is enriched at all human centromere regions and is competent for de novo centromere assembly. Because of the highly repetitive nature of alpha satellite, it has been difficult to achieve genome assemblies at centromeres using traditional next-generation sequencing approaches, and thus, centromeres represent gaps in the current human genome assembly. Moreover, alpha satellite DNA is transcribed into repetitive noncoding RNA and contributes to a large portion of the transcriptome. Recent efforts to characterize these transcripts and their function have uncovered pivotal roles for satellite RNA in genome stability, including silencing "selfish" DNA elements and recruiting centromere and kinetochore proteins. This review will describe the genomic and epigenetic features of alpha satellite DNA, discuss recent findings of noncoding transcripts produced from distinct alpha satellite arrays, and address current progress in the functional understanding of this oft-neglected repetitive sequence. We will discuss unique challenges of studying human satellite DNAs and RNAs and point toward new technologies that will continue to advance our understanding of this largely untapped portion of the genome.

The HGP-write project, announced in 2016 but not really implemented yet, comes back as a project aimed at constructing an "ultra-safe" human cell line fully resistant to virus infection and with other desirable characteristics. This involves introducing 400,000 changes in the genome and raises a number of technical and financial issues, but may become realistic in mid-term., (© 2018 médecine/sciences – Inserm.)

In recent years, protocols have been established to differentiate stem and precursor cells into more mature cell types. However, progress in this field has been hampered by difficulties to assess the differentiation status of stem cell-derived cells in an unbiased manner. Here, we present an analysis pipeline based on published data and methods to quantify the degree of differentiation and to identify transcriptional control factors explaining differences from the intended target cells or tissues. The pipeline requires RNA-Seq or gene array data of the stem cell starting population, derived 'mature' cells and primary target cells or tissue. It consists of a principal component analysis to represent global expression changes and to identify possible problems of the dataset that require special attention, such as: batch effects; clustering techniques to identify gene groups with similar features; over-representation analysis to characterize biological motifs and transcriptional control factors of the identified gene clusters; and metagenes as well as gene regulatory networks for quantitative cell-type assessment and identification of influential transcription factors. Possibilities and limitations of the analysis pipeline are illustrated using the example of human embryonic stem cell and human induced pluripotent cells to generate 'hepatocyte-like cells'. The pipeline quantifies the degree of incomplete differentiation as well as remaining stemness and identifies unwanted features, such as colon- and fibroblast-associated gene clusters that are absent in real hepatocytes but typically induced by currently available differentiation protocols. Finally, transcription factors responsible for incomplete and unwanted differentiation are identified. The proposed method is widely applicable and allows an unbiased and quantitative assessment of stem cell-derived cells.This article is part of the theme issue 'Designer human tissue: coming to a lab near you'., (© 2018 The Author(s).)

Structural and quantitative chromosomal rearrangements, collectively referred to as structural variation (SV), contribute to a large extent to the genetic diversity of the human genome and thus are of high relevance for cancer genetics, rare diseases and evolutionary genetics. Recent studies have shown that SVs can not only affect gene dosage but also modulate basic mechanisms of gene regulation. SVs can alter the copy number of regulatory elements or modify the 3D genome by disrupting higher-order chromatin organization such as topologically associating domains. As a result of these position effects, SVs can influence the expression of genes distant from the SV breakpoints, thereby causing disease. The impact of SVs on the 3D genome and on gene expression regulation has to be considered when interpreting the pathogenic potential of these variant types.

The enhancer-promoter interactions (EPIs) with strong tissue-specificity play an important role in cis-regulatory mechanism of human cell lines. However, it still remains a challenging work to predict these interactions so far. Due to that these interactions are regulated by the cooperativeness of diverse functional genomic signatures, DNA spatial structure and DNA sequence elements. In this paper, by adding DNA structure properties and transcription factor binding motifs, we presented an improved computational method to predict EPIs in human cell lines. In comparison with the results of other group on the same datasets, our best accuracies by cross-validation test were about 15%-24% higher in the same cell lines, and the accuracies by independent test were about 11%-15% higher in new cell lines. Meanwhile, we found that transcription factor binding motifs and DNA structure properties have important information that would largely determine long range EPIs prediction. From the distribution comparisons, we also found their distinct differences between interacting and non-interacting sets in each cell line. Then, the correlation analysis and network models for relationships among top-ranked functional genomic signatures indicated that diverse genomic signatures would cooperatively establish a complex regulatory network to facilitate long range EPIs. The experimental results provided additional insights about the roles of DNA intrinsic properties and functional genomic signatures in EPIs prediction., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Chromosome conformation capture (3C) and its derivatives (e.g., 4C, 5C and Hi-C) are used to analyze the 3D organization of genomes. We recently developed targeted chromatin capture (T2C), an inexpensive method for studying the 3D organization of genomes, interactomes and structural changes associated with gene regulation, the cell cycle, and cell survival and development. Here, we present the protocol for T2C based on capture, describing all experimental steps and bio-informatic tools in full detail. T2C offers high resolution, a large dynamic interaction frequency range and a high signal-to-noise ratio. Its resolution is determined by the resulting fragment size of the chosen restriction enzyme, which can lead to sub-kilobase-pair resolution. T2C's high coverage allows the identification of the interactome of each individual DNA fragment, which makes binning of reads (often used in other methods) basically unnecessary. Notably, T2C requires low sequencing efforts. T2C also allows multiplexing of samples for the direct comparison of multiple samples. It can be used to study topologically associating domains (TADs), determining their position, shape, boundaries, and intra- and inter-domain interactions, as well as the composition of aggregated loops, interactions between nucleosomes, individual transcription factor binding sites, and promoters and enhancers. T2C can be performed by any investigator with basic skills in molecular biology techniques in ∼7-8 d. Data analysis requires basic expertise in bioinformatics and in Linux and Python environments.

Histones are punctuated with small chemical modifications that alter their interaction with DNA. One attractive hypothesis stipulates that certain combinations of these histone modifications may function, alone or together, as a part of a predictive histone code to provide ground rules for chromatin folding. We consider four features that relate histone modifications to chromatin folding: charge neutralisation, molecular specificity, robustness and evolvability. Next, we present evidence for the association among different histone modifications at various levels of chromatin organisation and show how these relationships relate to function such as transcription, replication and cell division. Finally, we propose a model where the histone code can set critical checkpoints for chromatin to fold reversibly between different orders of the organisation in response to a biological stimulus., (Copyright © 2017 Elsevier B.V. All rights reserved.)

It is well established that cells sense chemical signals from their local microenvironment and transduce them to the nucleus to regulate gene expression programmes. Although a number of experiments have shown that mechanical cues can also modulate gene expression, the underlying mechanisms are far from clear. Nevertheless, we are now beginning to understand how mechanical cues are transduced to the nucleus and how they influence nuclear mechanics, genome organization and transcription. In particular, recent progress in super-resolution imaging, in genome-wide application of RNA sequencing, chromatin immunoprecipitation and chromosome conformation capture and in theoretical modelling of 3D genome organization enables the exploration of the relationship between cell mechanics, 3D chromatin configurations and transcription, thereby shedding new light on how mechanical forces regulate gene expression.

Alternative splicing plays important roles in generating different transcripts from one gene, and consequently various protein isoforms. However, there has been no systematic approach that facilitates characterizing functional roles of protein isoforms in the context of the entire human metabolism. Here, we present a systematic framework for the generation of gene-transcript-protein-reaction associations (GeTPRA) in the human metabolism. The framework in this study generated 11,415 GeTPRA corresponding to 1,106 metabolic genes for both principal and nonprincipal transcripts (PTs and NPTs) of metabolic genes. The framework further evaluates GeTPRA, using a human genome-scale metabolic model (GEM) that is biochemically consistent and transcript-level data compatible, and subsequently updates the human GEM. A generic human GEM, Recon 2M.1, was developed for this purpose, and subsequently updated to Recon 2M.2 through the framework. Both PTs and NPTs of metabolic genes were considered in the framework based on prior analyses of 446 personal RNA-Seq data and 1,784 personal GEMs reconstructed using Recon 2M.1. The framework and the GeTPRA will contribute to better understanding human metabolism at the systems level and enable further medical applications., Competing Interests: Conflict of interest statement: S.Y.L. and H.U.K. have coauthored publications with J.N. and N.D.P. These were Commentary articles and did not involve any research collaboration., (Published under the PNAS license.)

The AT-rich DNA is mostly associated with condensed chromatin, whereas the GC-rich sequence is preferably located in the dispersed chromatin. The AT-rich genes are prone to be tissue-specific (silenced in most tissues), while the GC-rich genes tend to be housekeeping (expressed in many tissues). This paper reports another important property of DNA base composition, which can affect repertoire of genes with high AT content. The GC-rich sequence is more liable to mutation. We found that Spearman correlation between human gene GC content and mutation probability is above 0.9. The change of base composition even in synonymous sites affects mutation probability of nonsynonymous sites and thus of encoded proteins. There is a unique type of housekeeping genes, which are especially unsafe when prone to mutation. Natural selection which usually removes deleterious mutations, in the case of these genes only increases the hazard because it can descend to suborganismal (cellular) level. These are cell cycle-related genes. In accordance with the proposed concept, they have low GC content of synonymous sites (despite them being housekeeping). The gene-centred protein interaction enrichment analysis (PIEA) showed the core clusters of genes whose interactants are modularly enriched in genes with AT-rich synonymous codons. This interconnected network is involved in double-strand break repair, DNA integrity checkpoints and chromosome pairing at mitosis. The damage of these genes results in genome and chromosome instability leading to cancer and other 'error catastrophes'. Reducing the nonsynonymous mutations, the usage of AT-rich synonymous codons can decrease probability of cancer by above 20-fold.

Translation of mRNAs through Internal Ribosome Entry Sites (IRESs) has emerged as a prominent mechanism of cellular and viral initiation. It supports cap-independent translation of select cellular genes under normal conditions, and in conditions when cap-dependent translation is inhibited. IRES structure and sequence are believed to be involved in this process. However due to the small number of IRESs known, there have been no systematic investigations of the determinants of IRES activity. With the recent discovery of thousands of novel IRESs in human and viruses, the next challenge is to decipher the sequence determinants of IRES activity. We present the first in-depth computational analysis of a large body of IRESs, exploring RNA sequence features predictive of IRES activity. We identified predictive k-mer features resembling IRES trans-acting factor (ITAF) binding motifs across human and viral IRESs, and found that their effect on expression depends on their sequence, number and position. Our results also suggest that the architecture of retroviral IRESs differs from that of other viruses, presumably due to their exposure to the nuclear environment. Finally, we measured IRES activity of synthetically designed sequences to confirm our prediction of increasing activity as a function of the number of short IRES elements.

During the cell cycle, the genome must undergo dramatic changes in structure, from a decondensed, yet highly organized interphase structure to a condensed, generic mitotic chromosome and then back again. For faithful cell division, the genome must be replicated and chromosomes and sister chromatids physically segregated from one another. Throughout these processes, there is feedback and tension between the information-storing role and the physical properties of chromosomes. With a combination of recent techniques in fluorescence microscopy, chromosome conformation capture (Hi-C), biophysical experiments, and computational modeling, we can now attribute mechanisms to many long-observed features of chromosome structure changes during cell division. Apparent conflicts that arise when integrating the concepts from these different proposed mechanisms emphasize that orchestrating chromosome organization during cell division requires a complex system of factors rather than a simple pathway. Cell division is both essential for and threatening to proper genome organization. As interphase three-dimensional (3D) genome structure is quite static at a global level, cell division provides an important window of opportunity to make substantial changes in 3D genome organization in daughter cells, allowing for proper differentiation and development. Mistakes in the process of chromosome condensation or rebuilding the structure after mitosis can lead to diseases such as cancer, premature aging, and neurodegeneration. WIREs Syst Biol Med 2017, 9:e1389. doi: 10.1002/wsbm.1389 For further resources related to this article, please visit the WIREs website., (© 2017 Wiley Periodicals, Inc.)

Remarkable progress has been made in understanding chromosome structures inside the cell nucleus. Recent advances in Hi-C technologies enable the detection of genome-wide chromatin interactions, providing insight into three-dimensional (3D) genome organization. Advancements in the spatial and temporal resolutions of imaging as well as in molecular biological techniques allow the tracking of specific chromosomal loci, improving our understanding of chromosome movements. From these data, we are beginning to understand how the intra-nuclear locations of chromatin loci and the 3D genome structure change during development and differentiation. This emerging field of genome structure and dynamics research requires an interdisciplinary approach including efficient collaborations between experimental biologists and physicists, informaticians, or engineers. Quantitative and mathematical analyses based on polymer physics are becoming increasingly important for processing and interpreting experimental data on 3D chromosome structures and dynamics. In this review, we aim to provide an overview of recent research on the physical aspects of chromosome structure and dynamics oriented for biologists. These studies have mainly focused on chromosomes at the cellular level, using unicellular organisms and cultured cells. However, physical parameters that change during development, such as nuclear size, may impact genome structure and dynamics. Here, we discuss how chromatin dynamics and genome structures in early embryos change during development, which we expect will be a hot topic in the field of chromatin dynamics in the near future. We hope this review helps developmental biologists to quantitatively investigate the physical natures of chromosomes in developmental biology research., (© 2017 Japanese Society of Developmental Biologists.)

Here we describe AGCGA-quadruplexes, an unexpected addition to the well-known tetrahelical families, G-quadruplexes and i-motifs, that have been a focus of intense research due to their potential biological impact in G- and C-rich DNA regions, respectively. High-resolution structures determined by solution-state nuclear magnetic resonance (NMR) spectroscopy demonstrate that AGCGA-quadruplexes comprise four 5'-AGCGA-3' tracts and are stabilized by G-A and G-C base pairs forming GAGA- and GCGC-quartets, respectively. Residues in the core of the structure are connected with edge-type loops. Sequences of alternating 5'-AGCGA-3' and 5'-GGG-3' repeats could be expected to form G-quadruplexes, but are shown herein to form AGCGA-quadruplexes instead. Unique structural features of AGCGA-quadruplexes together with lower sensitivity to cation and pH variation imply their potential biological relevance in regulatory regions of genes responsible for basic cellular processes that are related to neurological disorders, cancer and abnormalities in bone and cartilage development.

Cell division, cell differentiation and cell death are the three principal physiological processes that regulate tissue homoeostasis in multicellular organisms. The growth and survival of cells as well as the integrity of the genome are regulated by a complex network of pathways, in which cell cycle checkpoints, DNA repair and programmed cell death have critical roles. Disruption of genomic integrity and impaired regulation of cell death may both lead to uncontrolled cell growth. Compromised cell death can also favour genomic instability. It is becoming increasingly clear that dysregulation of cell cycle and cell death processes plays an important role in the development of major disorders such as cancer, cardiovascular disease, infection, inflammation and neurodegenerative diseases. Research achievements in these fields have led to the development of novel approaches for treatment of various conditions associated with abnormalities in the regulation of cell cycle progression or cell death. A better understanding of how cellular life-and-death processes are regulated is essential for this development. To highlight these important advances, the Third Nobel Conference entitled 'The Cell Cycle and Cell Death in Disease' was organized at Karolinska Institutet in 2016. In this review we will summarize current understanding of cell cycle progression and cell death and discuss some of the recent advances in therapeutic applications in pathological conditions such as cancer, neurological disorders and inflammation., (© 2017 The Association for the Publication of the Journal of Internal Medicine.)

The variability potential of a phenotype, on the background of the same genotype, is termed "phenotypic plasticity". This is considered by some scientists as a more important evolutionary procedure than mutation based natural selection. In this review, I discuss the fact that microRNAs have the potential to determine significantly the spectrum of phenotypic plasticity by regulating translation rate. MicroRNAs constitute a complicate network that can adjust at a variety of external environmental stimuli. They can also rebalance genome's function after genomic duplication events. Despite this, phenotypic plasticity is not limitless. Modern way of human life is pushing our physiology at its limits. On the other site, old people have dysregulated microRNA levels, this reducing their potential for plastic phenotypes. Can we intervene correcting this dysregulation? The system is very complicate and our knowledge is incomplete. More research is needed in order to understand deeper the exact relation of microRNAs with plasticity and aging, by taking advantage on the newly developed high-throughput methods., (Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.)

Human copy number variants (CNVs) account for genome variation an order of magnitude larger than single-nucleotide polymorphisms. Although much of this variation has no phenotypic consequences, some variants have been associated with disease, in particular neurodevelopmental disorders. Pathogenic CNVs are typically very large and contain multiple genes, and understanding the cause of the pathogenicity remains a major challenge. Here we show that pathogenic CNVs are significantly enriched for genes involved in development and genes that have greater evolutionary copy number conservation across mammals, indicative of functional constraints. Conversely, genes found in benign CNV regions have more variable copy number. These evolutionary constraints are characteristic of genes in pathogenic CNVs and can only be explained by dosage sensitivity of those genes. These results implicate dosage sensitivity of individual genes as a common cause of CNV pathogenicity. These evolutionary metrics suggest a path to identifying disease genes in pathogenic CNVs.

It is estimated that more than 90% of the mammalian genome is transcribed as non-coding RNAs. Recent evidences have established that these non-coding transcripts are not junk or just transcriptional noise, but they do serve important biological purpose. One of the rapidly expanding fields of this class of transcripts is the regulatory lncRNAs, which had been a major challenge in terms of their molecular functions and mechanisms of action. The emergence of high-throughput technologies and the development in various conventional approaches have led to the expansion of the lncRNA world. The combination of multidisciplinary approaches has proven to be essential to unravel the complexity of their regulatory networks and helped establish the importance of their existence. Here, we review the current methodologies available for discovering and investigating functions of long non-coding RNAs (lncRNAs) and focus on the powerful technological advancement available to specifically address their functional importance.

For the last four decades, we have known that noncoding RNAs maintain critical housekeeping functions such as transcription, RNA processing, and translation. However, in the late 1990s and early 2000s, the advent of high-throughput sequencing technologies and computational tools to analyze these large sequencing datasets facilitated the discovery of thousands of small and long noncoding RNAs (lncRNAs) and their functional role in diverse biological functions. For example, lncRNAs have been shown to regulate dosage compensation, genomic imprinting, pluripotency, cell differentiation and development, immune response, etc. Here we review how lncRNAs bring about such copious functions by employing diverse mechanisms such as translational inhibition, mRNA degradation, RNA decoys, facilitating recruitment of chromatin modifiers, regulation of protein activity, regulating the availability of miRNAs by sponging mechanism, etc. In addition, we provide a detailed account of different mechanisms as well as general principles by which lncRNAs organize functionally different nuclear sub-compartments and their impact on nuclear architecture.

Background: A fundamental issue in bioscience is to understand the mechanism that underlies the dynamic control of genome-wide expression through the complex temporal-spatial self-organization of the genome to regulate the change in cell fate. We address this issue by elucidating a physically motivated mechanism of self-organization., Principal Findings: Building upon transcriptome experimental data for seven distinct cell fates, including early embryonic development, we demonstrate that self-organized criticality (SOC) plays an essential role in the dynamic control of global gene expression regulation at both the population and single-cell levels. The novel findings are as follows: i) Mechanism of cell-fate changes: A sandpile-type critical transition self-organizes overall expression into a few transcription response domains (critical states). A cell-fate change occurs by means of a dissipative pulse-like global perturbation in self-organization through the erasure of initial-state critical behaviors (criticality). Most notably, the reprogramming of early embryo cells destroys the zygote SOC control to initiate self-organization in the new embryonal genome, which passes through a stochastic overall expression pattern. ii) Mechanism of perturbation of SOC controls: Global perturbations in self-organization involve the temporal regulation of critical states. Quantitative evaluation of this perturbation in terminal cell fates reveals that dynamic interactions between critical states determine the critical-state coherent regulation. The occurrence of a temporal change in criticality perturbs this between-states interaction, which directly affects the entire genomic system. Surprisingly, a sub-critical state, corresponding to an ensemble of genes that shows only marginal changes in expression and consequently are considered to be devoid of any interest, plays an essential role in generating a global perturbation in self-organization directed toward the cell-fate change., Conclusion and Significance: 'Whole-genome' regulation of gene expression through self-regulatory SOC control complements gene-by-gene fine tuning and represents a still largely unexplored non-equilibrium statistical mechanism that is responsible for the massive reprogramming of genome expression., Competing Interests: The authors have declared that no competing interests exist.

High altitude, defined as elevations lying above 2500m sea level, challenges human survival and reproduction. This environment provides a natural experimental design wherein specific populations, Andeans, Ethiopians, and Tibetans, have lived in a chronic hypoxia state for millennia. These human groups have overcome the low ambient oxygen tension of high elevation via unique physiologic and genetic adaptations. Genomic studies have identified several genes that underlie high-altitude adaptive phenotypes, many of which are central components of the Hypoxia Inducible Factor (HIF) pathway. Further study of mechanisms governing the adaptive changes responsible for high-altitude adaptation will contribute to our understanding of the molecular basis of evolutionary change and assist in the functional annotation of the human genome., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Aquaporin-2 (AQP2) is regulated in part via vasopressin-mediated changes in protein half-life that are in turn dependent on AQP2 ubiquitination. Here we addressed the question, "What E3 ubiquitin ligase is most likely to be responsible for AQP2 ubiquitination?" using large-scale data integration based on Bayes' rule. The first step was to bioinformatically identify all E3 ligase genes coded by the human genome. The 377 E3 ubiquitin ligases identified in the human genome, consisting predominant of HECT, RING, and U-box proteins, have been used to create a publically accessible and downloadable online database (https://hpcwebapps.cit.nih.gov/ESBL/Database/E3-ligases/). We also curated a second database of E3 ligase accessory proteins that included BTB domain proteins, cullins, SOCS-box proteins, and F-box proteins. Using Bayes' theorem to integrate information from multiple large-scale proteomic and transcriptomic datasets, we ranked these 377 E3 ligases with respect to their probability of interaction with AQP2. Application of Bayes' rule identified the E3 ligases most likely to interact with AQP2 as (in order of probability): NEDD4 and NEDD4L (tied for first), AMFR, STUB1, ITCH, ZFPL1. Significantly, the two E3 ligases tied for top rank have also been studied extensively in the reductionist literature as regulatory proteins in renal tubule epithelia. The concordance of conclusions from reductionist and systems-level data provides strong motivation for further studies of the roles of NEDD4 and NEDD4L in the regulation of AQP2 protein turnover.

The regions of the genome that interact frequently with the nucleolus have been termed nucleolar-associated domains (NADs). Deep sequencing and DNA-fluorescence in situ hybridization (FISH) experiments have revealed that these domains are enriched for repetitive elements, regions of the inactive X chromosome (Xi), and several RNA polymerase III-transcribed genes. NADs are often marked by chromatin modifications characteristic of heterochromatin, including H3K27me3, H3K9me3, and H4K20me3, and artificial targeting of genes to this area is correlated with reduced expression. It has therefore been hypothesized that NAD localization to the nucleolar periphery contributes to the establishment and/or maintenance of heterochromatic silencing. Recently published studies from several multicellular eukaryotes have begun to reveal the trans-acting factors involved in NAD localization, including the insulator protein CCCTC-binding factor (CTCF), chromatin assembly factor (CAF)-1 subunit p150, several nucleolar proteins, and two long non-coding RNAs (lncRNAs). The mechanisms by which these factors coordinate with one another in regulating NAD localization and/or silencing are still unknown. This review will summarize recently published studies, discuss where additional research is required, and speculate about the mechanistic and functional implications of genome organization around the nucleolus.

The maintenance of genome stability is critical for proper cell function, and loss of this stability contributes to many human diseases and developmental disorders. Therefore, cells have evolved partially redundant mechanisms to monitor and protect the genome. One subcellular organelle implicated in the maintenance of genome stability is the centrosome, best known as the primary microtubule organizing center of most animal cells. Centrosomes serve many different roles throughout the cell cycle, and many of those roles, including mitotic spindle assembly, nucleation of the interphase microtubule array, DNA damage response, and efficient cell cycle progression, have been proposed to help maintain genome stability. As a result, the centrosome is itself a highly regulated entity. Here, we review evidence concerning the significance of the centrosome in promoting genome integrity. Recent advances permitting acute and persistent centrosome removal suggest we still have much to learn regarding the specific function and actual importance of centrosomes in different contexts, as well as how cells may compensate for centrosome dysfunction to maintain the integrity of the genome. Although many animal cells survive and proliferate in the absence of centrosomes, they do so aberrantly. Based on these and other studies, we conclude that centrosomes serve as critical, multifunctional organelles that promote genome stability.

Transposable elements (TEs) have the capacity to replicate and insert into new genomic locations. This contributs significantly to evolution of genomes, but can also result in DNA breaks and illegitimate recombination, and therefore poses a significant threat to genomic integrity. Excess damage to the germ cell genome results in sterility. A specific RNA silencing pathway, termed the piRNA pathway operates in germ cells of animals to control TE activity. At the core of the piRNA pathway is a ribonucleoprotein complex consisting of a small RNA, called piRNA, and a protein from the PIWI subfamily of Argonaute nucleases. The piRNA pathway relies on the specificity provided by the piRNA sequence to recognize complementary TE targets, while effector functions are provided by the PIWI protein. PIWI-piRNA complexes silence TEs both at the transcriptional level - by attracting repressive chromatin modifications to genomic targets - and at the posttranscriptional level - by cleaving TE transcripts in the cytoplasm. Impairment of the piRNA pathway leads to overexpression of TEs, significantly compromised genome structure and, invariably, germ cell death and sterility.The piRNA pathway is best understood in the fruit fly, Drosophila melanogaster, and in mouse. This Chapter gives an overview of current knowledge on piRNA biogenesis, and mechanistic details of both transcriptional and posttranscriptional TE silencing by the piRNA pathway. It further focuses on the importance of post-translational modifications and subcellular localization of the piRNA machinery. Finally, it provides a brief description of analogous pathways in other systems.

A complex interplay between multiple biological effects shapes the aging process. The advent of genome-wide quantitative approaches in the epigenetic field has highlighted the effective impact of epigenetic deregulation, particularly of DNA methylation, on aging. Age-associated alterations in DNA methylation are commonly grouped in the phenomenon known as "epigenetic drift" which is characterized by gradual extensive demethylation of genome and hypermethylation of a number of promoter-associated CpG islands. Surprisingly, specific DNA regions show directional epigenetic changes in aged individuals suggesting the importance of these events for the aging process. However, the epigenetic information obtained until now in aging needs a re-consideration due to the recent discovery of 5-hydroxymethylcytosine, a new DNA epigenetic mark present on genome. A recapitulation of the factors involved in the regulation of DNA methylation and the changes occurring in aging will be described in this review also considering the data available on 5 hmC., (Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.)

Genome instability represents a primary hallmark of aging and cancer. RecQL helicases (i.e., RECQL1, WRN, BLM, RECQL4, RECQL5) as well as poly(ADP-ribose) polymerases (PARPs, in particular PARP1) represent two central quality control systems to preserve genome integrity in mammalian cells. Consistently, both enzymatic families have been linked to mechanisms of aging and carcinogenesis in mice and humans. This is in accordance with clinical and epidemiological findings demonstrating that defects in three RecQL helicases, i.e., WRN, BLM, RECQL4, are related to human progeroid and cancer predisposition syndromes, i.e., Werner, Bloom, and Rothmund Thomson syndrome, respectively. Moreover, PARP1 hypomorphy is associated with a higher risk for certain types of cancer. On a molecular level, RecQL helicases and PARP1 are involved in the control of DNA repair, telomere maintenance, and replicative stress. Notably, over the last decade, it became apparent that all five RecQL helicases physically or functionally interact with PARP1 and/or its enzymatic product poly(ADP-ribose) (PAR). Furthermore, a profound body of evidence revealed that the cooperative function of RECQLs and PARP1 represents an important factor for maintaining genome integrity. In this review, we summarize the status quo of this molecular cooperation and discuss open questions that provide a basis for future studies to dissect the cooperative functions of RecQL helicases and PARP1 in aging and carcinogenesis., (Copyright © 2015 Elsevier B.V. All rights reserved.)