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Author(s): Warren Winick-Ng [sup.1] , Alexander Kukalev [sup.1] , Izabela Harabula [sup.1] [sup.2] , Luna Zea-Redondo [sup.1] [sup.2] , Dominik Szabó [sup.1] [sup.2] , Mandy Meijer [sup.3] , Leonid Serebreni [...], The three-dimensional (3D) structure of chromatin is intrinsically associated with gene regulation and cell function.sup.1-3. Methods based on chromatin conformation capture have mapped chromatin structures in neuronal systems such as in vitro differentiated neurons, neurons isolated through fluorescence-activated cell sorting from cortical tissues pooled from different animals and from dissociated whole hippocampi.sup.4-6. However, changes in chromatin organization captured by imaging, such as the relocation of Bdnf away from the nuclear periphery after activation.sup.7, are invisible with such approaches.sup.8. Here we developed immunoGAM, an extension of genome architecture mapping (GAM).sup.2,9, to map 3D chromatin topology genome-wide in specific brain cell types, without tissue disruption, from single animals. GAM is a ligation-free technology that maps genome topology by sequencing the DNA content from thin (about 220 nm) nuclear cryosections. Chromatin interactions are identified from the increased probability of co-segregation of contacting loci across a collection of nuclear slices. ImmunoGAM expands the scope of GAM to enable the selection of specific cell types using low cell numbers (approximately 1,000 cells) within a complex tissue and avoids tissue dissociation.sup.2,10. We report cell-type specialized 3D chromatin structures at multiple genomic scales that relate to patterns of gene expression. We discover extensive 'melting' of long genes when they are highly expressed and/or have high chromatin accessibility. The contacts most specific of neuron subtypes contain genes associated with specialized processes, such as addiction and synaptic plasticity, which harbour putative binding sites for neuronal transcription factors within accessible chromatin regions. Moreover, sensory receptor genes are preferentially found in heterochromatic compartments in brain cells, which establish strong contacts across tens of megabases. Our results demonstrate that highly specific chromatin conformations in brain cells are tightly related to gene regulation mechanisms and specialized functions. A new technique called immunoGAM, which combines genome architecture mapping (GAM) with immunoselection, enabled the discovery of specialized chromatin conformations linked to gene expression in specific cell populations from mouse brain tissues.

Muscular dystrophies (MDs) comprise a heterogeneous group of disorders that disrupt the function of different muscle groups, with manifestations including muscle weakness, muscle atrophy, spasm, muscle hypertonia, and myalgias (Figure [...], Muscular dystrophies are debilitating disorders that result in progressive weakness and degeneration of skeletal muscle. Although the genetic mutations and clinical abnormalities of a variety of neuromuscular diseases are well known, no curative therapies have been developed to date. The advent of genome editing technology provides new opportunities to correct the underlying mutations responsible for many monogenic neuromuscular diseases. For example, Duchenne muscular dystrophy, which is caused by mutations in the dystrophin gene, has been successfully corrected in mice, dogs, and human cells through CRISPR/Cas9 editing. In this Review, we focus on the potential for, and challenges of, correcting muscular dystrophies by editing disease-causing mutations at the genomic level. Ideally, because muscle tissues are extremely long-lived, CRISPR technology could offer a one-time treatment for muscular dystrophies by correcting the culprit genomic mutations and enabling normal expression of the repaired gene.

Author(s): Jeffrey M. Perkel Author Affiliations: Single-cell analysis enters the multiomics age Single-cell data (top) increasingly are being combined with spatial information (bottom). Credit: 10x Genomics Two sheets of visual [...], A rapidly growing collection of software tools is helping researchers to analyse multiple huge '-omics' data sets. A rapidly growing collection of software tools is helping researchers to analyse multiple huge '-omics' data sets.

Author(s): Joshua A. Arribere [sup.1] , Elif S. Cenik [sup.1] , Nimit Jain [sup.2] , Gaelen T. Hess [sup.3] , Cameron H. Lee [sup.3] , Michael C. Bassik [sup.3] , [...], Translation termination sequences are occasionally bypassed by the ribosome and the resulting proteins can be detrimental to the cell; here it is shown that cells can prevent such proteins from accumulating through peptides that are encoded within the 3' UTR of genes in both humans and C. elegans. Removal of proteins produced by translational readthrough Messenger RNAs contain stop sequences, but these are sometimes bypassed by the ribosome, leading to an extension at the C terminus of the protein. Such proteins are often detrimental to the cell, suggesting that there are surveillance mechanisms to detect and deal with these proteins. Using a combination of CRISPR-Cas9-mediated editing and transgenics, Andrew Fire and colleagues demonstrate that cells can prevent such proteins from accumulating, through peptides that are encoded within the 3' UTR of genes in both Caenorhabditis elegans and humans. Although the mechanism is not yet clear, these peptides are able to lower the levels of C-terminal-extended proteins, providing mitigation of unwelcome and varied translation errors. A fraction of ribosomes engaged in translation will fail to terminate when reaching a stop codon, yielding nascent proteins inappropriately extended on their C termini. Although such extended proteins can interfere with normal cellular processes, known mechanisms of translational surveillance.sup.1 are insufficient to protect cells from potential dominant consequences. Here, through a combination of transgenics and CRISPR-Cas9 gene editing in Caenorhabditis elegans, we demonstrate a consistent ability of cells to block accumulation of C-terminal-extended proteins that result from failure to terminate at stop codons. Sequences encoded by the 3' untranslated region (UTR) were sufficient to lower protein levels. Measurements of mRNA levels and translation suggested a co- or post-translational mechanism of action for these sequences in C. elegans. Similar mechanisms evidently operate in human cells, in which we observed a comparable tendency for translated human 3' UTR sequences to reduce mature protein expression in tissue culture assays, including 3' UTR sequences from the hypomorphic 'Constant Spring' haemoglobin stop codon variant. We suggest that 3' UTRs may encode peptide sequences that destabilize the attached protein, providing mitigation of unwelcome and varied translation errors.

Author(s): Florian Erhard [sup.1] , Marisa A. P. Baptista [sup.1] , Tobias Krammer [sup.2] , Thomas Hennig [sup.1] , Marius Lange [sup.3] [sup.4] , Panagiota Arampatzi [sup.5] , Christopher S. [...], Single-cell RNA sequencing (scRNA-seq) has highlighted the important role of intercellular heterogeneity in phenotype variability in both health and disease.sup.1. However, current scRNA-seq approaches provide only a snapshot of gene expression and convey little information on the true temporal dynamics and stochastic nature of transcription. A further key limitation of scRNA-seq analysis is that the RNA profile of each individual cell can be analysed only once. Here we introduce single-cell, thiol-(SH)-linked alkylation of RNA for metabolic labelling sequencing (scSLAM-seq), which integrates metabolic RNA labelling.sup.2, biochemical nucleoside conversion.sup.3 and scRNA-seq to record transcriptional activity directly by differentiating between new and old RNA for thousands of genes per single cell. We use scSLAM-seq to study the onset of infection with lytic cytomegalovirus in single mouse fibroblasts. The cell-cycle state and dose of infection deduced from old RNA enable dose-response analysis based on new RNA. scSLAM-seq thereby both visualizes and explains differences in transcriptional activity at the single-cell level. Furthermore, it depicts 'on-off' switches and transcriptional burst kinetics in host gene expression with extensive gene-specific differences that correlate with promoter-intrinsic features (TBP-TATA-box interactions and DNA methylation). Thus, gene-specific, and not cell-specific, features explain the heterogeneity in transcriptomes between individual cells and the transcriptional response to perturbations. A technique known as scSLAM-seq that combines single-cell RNA sequencing with metabolic RNA labelling and nucleoside conversion is used to study the onset of cytomegalovirus infection in single mouse fibroblasts.

Author(s): Paul Smaglik Author Affiliations: Ido Amit, a genome researcher and immunologist at the Weizmann Institute of Science in Rehovot, Israel, had a problem. When he and his colleagues were [...], Single-cell genomic sequencing is poised to revolutionize fields from cancer to immunology.

Author(s): Leonid A. Mirny Author Affiliations: Cells use loop extrusion to weave and tie the genome One of the challenges that all organisms face is how to compress long DNA [...], A study reveals that a process called loop extrusion, which is central to the folding and function of chromosomes, also seems to play a key part in the repair of double-strand DNA breaks. A role for loop extrusion in repairing double-strand DNA breaks.

Four-way DNA intermediates, also known as Holliday junctions, are formed during homologous recombination and DNA repair, and their resolution is necessary for proper chromosome segregation. Here we identify nucleases from [...]

Author(s): Anna Nowogrodzki [1] Aviv Regev likes to work at the edge of what is possible. In 2011, the computational biologist was collaborating with molecular geneticist Joshua Levin to test [...]

Author Affiliations: Genomic techniques can be used to specify the DNA sequence of cells in an entire mouse embryo (pictured). Credit: Meng Zhu, Zernicka-Goetz Laboratory, Cambridge University/SPL Eight-cell mouse embryo, [...], DNA barcodes help to detail the genomic architecture and sequence of an unbroken cell or even an entire embryo. DNA barcodes help to detail the genomic architecture and sequence of an unbroken cell or even an entire embryo.

In order to protect against a huge number of pathogens, the vertebrate immune system increases the limited antigen receptor repertoire encoded in the genome by taking advantage of somatic DNA [...], Activation-induced cytidine deaminase (AID), a putative RNA-editing enzyme, is indispensable for somatic hypermutation (SHM), class switch recombination, and gene conversion of immunoglobulin genes, which indicates a common molecular mechanism for these phenomena. Here we show that ectopic expression of AID alone can induce hypermutation in an artificial GFP substrate in NIH 3T3 murine fibroblast cells. The frequency of mutations was closely correlated with the level of transcription of the target gene, and the distribution of mutations in NIH 3T3 cells was similar to those of SHM in B lymphocytes. These results indicate that AID is sufficient for the generation of SHM in an actively transcribed gene in fibroblasts, as well as B cells, and that any of the required cofactors must be present in these fibroblasts.

Author(s): Hannah Stower [sup.1] Author Affiliations: (1) Nature Medicine, https://doi.org/10.1126/science.aav8130 https://doi.org/10.1038/s41586-019-1195-2 The advent of single-cell RNA sequencing (RNA-seq) technology has finally allowed scientists to analyze unstable RNA material in individual [...]

Transcription of eukaryotic genes requires the formation of a preinitiation complex containing one of three nuclear RNA polymerases and additional basal factors (1). Sequence-specific transcriptional activators function, in part, through [...], For unknown reasons, the eukaryotic transcription factor TFIID inefficiently recognizes promoters. Human TFIID was found to form highly specific homodimers that must dissociate before DNA binding. TFIID dimers formed through self-association of the TATA-binding polypeptide (TBP) subunit and could be immunoprecipitated with antibodies to [TAF.sub.∥]250, the core subunit of TFIID. Chemical cross-linking experiments in HeLa cells revealed the presence of TBP dimers in vivo. These findings suggest that dimerization through TBP is the physiological state of TFIID when not bound to DNA. Thus, the inefficiency of TFIID binding to a promoter may be partly attributable to the competitive effect of dimerization.

Two comments examine a study by Lynch and Conery (Reports, 10 Nov. 2000, p. 1151) that estimated the 'birth rate' and hail-life of gene duplications for several eukaryotic spedes over [...]

Inheritance of genetic information requires a faithful copying mechanism. DNA replication and repair provide high-fidelity inheritance over evolutionary periods, whereas epigenetic inheritance is less rigid. DNA methylation can mediate epigenetic [...], Every eukaryotic chromosome has a centromere, the locus responsible for poleward movement at mitosis and meiosis. Although conventional loci are specified by their DNA sequences, current evidence favors a chromatin-based inheritance mechanism for centromeres. The chromosome segregation machinery is highly conserved across all eukaryotes, but the DNA and protein components specific to centromeric chromatin are evolving rapidly. Incompatibilities between rapidly evolving centromeric components may be responsible for both the organization of centromeric regions and the reproductive isolation of emerging species.

Genomic DNA is the ultimate template of our heredity. Yet despite the justifiable excitement over the human genome, many challenges remain in understanding the regulation and transduction of genetic information [...], Chromatin, the physiological template of all eukaryotic genetic information, is subject to a diverse array of posttranslational modifications that largely impinge on histone amino termini, thereby regulating access to the underlying DNA. Distinct histone amino-terminal modifications can generate synergistic or antagonistic interaction affinities for chromatin-associated proteins, which in turn dictate dynamic transitions between transcriptionally active or transcriptionally silent chromatin states. The combinatorial nature of histone amino-terminal modifications thus reveals a 'histone code' that considerably extends the information potential of the genetic code. We propose that this epigenetic marking system represents a fundamental regulatory mechanism that has an impact on most, if not all, chromatin-templated processes, with far-reaching consequences for cell fate decisions and both normal and pathological development.

I. INTRODUCTION Mitogen-activated protein kinase (MAPK) signal transduction pathways are among the most widespread mechanisms of eukaryotic cell regulation. All eukaryotic cells possess multiple MAPK pathways, each of which is [...]

* The researchers have linked these epigenomic data to the corresponding genetic information, producing reference epigenomes for 127 tissue and cell [...]

Once upon a time, biologists drew the tree of life with two main branches. The prokaryotic kingdom, containing single-celled organisms, such as bacteria, that lack nuclei, occupied one limb; the [...]