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DNA and RNA can adopt various secondary structures. Four-stranded G-quadruplex (G4) structures form through self-recognition of guanines into stacked tetrads, and considerable biophysical and structural evidence exists for G4 formation in vitro. Computational studies and sequencing methods have revealed the prevalence of G4 sequence motifs at gene regulatory regions in various genomes, including in humans. Experiments using chemical, molecular and cell biology methods have demonstrated that G4s exist in chromatin DNA and in RNA, and have linked G4 formation with key biological processes ranging from transcription and translation to genome instability and cancer. In this Review, we first discuss the identification of G4s and evidence for their formation in cells using chemical biology, imaging and genomic technologies. We then discuss possible functions of DNA G4s and their interacting proteins, particularly in transcription, telomere biology and genome instability. Roles of RNA G4s in RNA biology, especially in translation, are also discussed. Furthermore, we consider the emerging relationships of G4s with chromatin and with RNA modifications. Finally, we discuss the connection between G4 formation and synthetic lethality in cancer cells, and recent progress towards considering G4s as therapeutic targets in human diseases.G-quadruplexes (G4s) are structures formed in guanine-rich DNA or RNA, which are linked to transcription, translation, chromatin biology, genome instability and RNA modifications. Recent studies connect G4 formation with cancer-cell lethality and indicate that G4s could be therapeutic targets.

PurposeThe present study considers disruption in the buyer–supplier–supplier triad. This triad has a common second-tier supplier as the disruption source, which gives us the tetradic context. The goal is to advance the knowledge on how a first-tier supplier's resilience against lower-tier disruptive events can be developed through horizontally connecting with the other first-tier supplier and how the buyer can benefit from its first-tier suppliers' resilience capability.Design/methodology/approachData from 33 triads was collected and analyzed.FindingsAs predicted, coopetition between two first-tier suppliers increases the first-tier supplier's capability to be resilient to disruptive events emanating from a lower tier source. However, contrary to initial theorization, the first-tier supplier's resilience capability affects the buyer's performance during disruptive events negatively. With increasing buyer–supplier social bonds, this negative relationship can partly be alleviated.Research limitations/implicationsAnalyzing resilience within a triad to a disruption in the tetradic context reveals unexpected dynamics. Individual supplier's resilience may have a negative impact on the buyer's resilience in certain disruption events.Practical implicationsThe buyer can increase collective suppliers' resilience through establishing horizontal links. To prevent becoming a victim of the supplier's resilience in the event of a second-tier disruption, a buyer needs to become a member of the supplier's relational network.Originality/valueWe propose that resilience can rest with the suppliers. This observation has implications for the buyer when selecting and coordinating suppliers. Further, it considers a context beyond a triad by venturing into the tetradic context. We anticipate more studies in tetrads in future and this study can serve as a bridge.

KRAS is one of the most highly mutated oncoproteins, which is overexpressed in various human cancers and implicated in poor survival. The G-quadruplex formed in KRAS oncogene promoter ( KRAS -G4) is a transcriptional modulator and amenable to small molecule targeting. However, no available KRAS -G4-ligand complex structure has yet been determined, which seriously hinders the structure-based rational design of KRAS -G4 targeting drugs. In this study, we report the NMR solution structures of a bulge-containing KRAS -G4 bound to berberine and coptisine, respectively. The determined complex structure shows a 2:1 binding stoichiometry with each compound recruiting the adjacent flacking adenine residue to form a “quasi-triad plane” that stacks over the two external G-tetrads. The binding involves both π -stacking and electrostatic interactions. Moreover, berberine and coptisine significantly lowered the KRAS mRNA levels in cancer cells. Our study thus provides molecular details of ligand interactions with KRAS -G4 and is beneficial for the design of specific KRAS -G4-interactive drugs. The G-quadruplex formed in KRAS oncogene promoter (KRAS-G4) is a transcriptional modulator and amenable to small molecule targeting. Herein, the authors report the NMR solution structures of a bulge-containing KRAS-G4 that bound to two small molecules. The study provides molecular details of ligand interactions with KRAS-G4 and contributes insight into the design of specific KRAS-G4-interactive drugs.

Background It has been proposed that more than 450 million years ago, two successive whole genome duplications took place in a marine chordate lineage before leading to the common ancestor of vertebrates. A precise reconstruction of these founding events would provide a framework to better understand the impact of these early whole genome duplications on extant vertebrates. Results We reconstruct the evolution of chromosomes at the beginning of vertebrate evolution. We first compare 61 extant animal genomes to reconstruct the highly contiguous order of genes in a 326-million-year-old ancestral Amniota genome. In this genome, we establish a well-supported list of duplicated genes originating from the two whole genome duplications to identify tetrads of duplicated chromosomes. From this, we reconstruct a chronology in which a pre-vertebrate genome composed of 17 chromosomes duplicated to 34 chromosomes and was subject to seven chromosome fusions before duplicating again into 54 chromosomes. After the separation of the lineage of Gnathostomata (jawed vertebrates) from Cyclostomata (extant jawless fish), four more fusions took place to form the ancestral Euteleostomi (bony vertebrates) genome of 50 chromosomes. Conclusions These results firmly establish the occurrence of two whole genome duplications in the lineage that precedes the ancestor of vertebrates, resolving in particular the ambiguity raised by the analysis of the lamprey genome. This work provides a foundation for studying the evolution of vertebrate chromosomes from the standpoint of a common ancestor and particularly the pattern of duplicate gene retention and loss that resulted in the gene composition of extant vertebrate genomes.

Perfect matching of an assembled physical sequence to a specified designed sequence is crucial to verify design principles in genome synthesis. We designed and de novo synthesized 536,024–base pair chromosome synV in the “Build-A-Genome China” course. We corrected an initial isolate of synV to perfectly match the designed sequence using integrative cotransformation and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)–mediated editing in 22 steps; synV strains exhibit high fitness under a variety of culture conditions, compared with that of wild-type V strains. A ring synV derivative was constructed, which is fully functional in Saccharomyces cerevisiae under all conditions tested and exhibits lower spore viability during meiosis. Ring synV chromosome can extends Sc2.0 design principles and provides a model with which to study genomic rearrangement, ring chromosome evolution, and human ring chromosome disorders.

Knowledge of the exact distribution of meiotic crossovers (COs) and gene conversions (GCs) is essential for understanding many aspects of population genetics and evolution, from haplotype structure and long-distance genetic linkage to the generation of new allelic variants of genes. To this end, we resequenced the four products of 13 meiotic tetrads along with 10 doubled haploids derived from Arabidopsis thaliana hybrids. GC detection through short reads has previously been confounded by genomic rearrangements. Rigid filtering for misaligned reads allowed GC identification at high accuracy and revealed an ∼80-kb transposition, which undergoes copy-number changes mediated by meiotic recombination. Non-crossover associated GCs were extremely rare most likely due to their short average length of ∼25–50 bp, which is significantly shorter than the length of CO-associated GCs. Overall, recombination preferentially targeted non-methylated nucleosome-free regions at gene promoters, which showed significant enrichment of two sequence motifs. Most living organisms package their DNA into bundles called chromosomes. These chromosomes generally form pairs, with each chromosome in the pair containing the same number of genes. The genes also come in the same order, but the exact sequence of DNA bases within the genes can be different. When sex cells—such as egg, sperm or pollen cells—are made, each pair of chromosomes is separated so that the each sex cell contains only half the normal number of chromosomes. However, before they are separated, the pairs swap lengths of DNA via recombination events. These can involve exchanging large chunks of the chromosomes: this is called a ‘crossover’. Alternatively, short stretches of one chromosome can be replaced by the corresponding region from the other in the pair. When these ‘non-crossovers’ cause a change in the DNA sequence they are known as gene conversions. Long-standing questions in the field of plant biology include: how common are gene conversions? How much DNA is typically exchanged? And where in the chromosomes do these events happen most? Now, Wijnker et al. have addressed these questions by focusing on the accurate detection of recombination events, with a special emphasis on gene conversions, in the plant biologist’s favourite species: Arabidopsis. Searching for recombination events is a challenge because, when piecing together an entire genome from lots of shorter stretches of DNA—typically called ‘reads’, it is easy to misplace some of the pieces. However, meticulous examination of these short DNA reads allowed Wijnker et al. to reliably identify gene conversions on a genome-wide scale. In Arabidopsis, gene conversion appears to be unexpectedly rare—with approximately one gene conversion detected per 140–240 non-crossovers. Recombination tends to occur in regions of the chromosomes where the DNA is only loosely packaged, is not heavily modified by the process of ‘DNA methylation’, and also near the start of genes. Furthermore, two specific sequences of DNA bases were identified that marked ‘hot spots’ in the chromosomes, where recombination happens more frequently. Wijnker et al. suggest that the low number of gene conversions detected indicates that non-crossovers tend to exchange very short stretches of DNA. However, future research may point to additional mechanisms that explain the low incidence of gene conversion in Arabidopsis.

The skeletal muscle dihydropyridine receptor (DHPR) β1a subunit is indispensable for full trafficking of DHPRs into triadic junctions (i.e., the close apposition of transverse tubules and sarcoplasmic reticulum [SR]), facilitation of DHPRα1S voltage sensing, and arrangement of DHPRs into tetrads as a consequence of their interaction with ryanodine receptor (RyR1) homotetramers. These three features are obligatory for skeletal muscle excitation–contraction (EC) coupling. Previously, we showed that all four vertebrate β isoforms (β₁–β₄) facilitate α1S triad targeting and, except for β₃, fully enable DHPRα1S voltage sensing [Dayal et al., Proc. Natl. Acad. Sci. U.S.A. 110, 7488–7493 (2013)]. Consequently, β₃ failed to restore EC coupling despite the fact that both β₃ and β1a restore tetrads. Thus, all β-subunits are able to restore triad targeting, but only β1a restores both tetrads and proper DHPR–RyR1 coupling [Dayal et al., Proc. Natl. Acad. Sci. U.S.A. 110, 7488–7493 (2013)]. To investigate the molecular region(s) of β1a responsible for the tetradic arrangement of DHPRs and thus DHPR–RyR1 coupling, we expressed loss- and gain-of-function chimeras between β1a and β₄, with systematically swapped domains in zebrafish strain relaxed (β₁-null) for patch clamp, cytoplasmic Ca2+ transients, motility, and freeze-fracture electron microscopy. β1a/β₄ chimeras with either N terminus, SH3, HOOK, or GK domain derived from β₄ showed complete restoration of SR Ca2+ release. However, chimera β1a/β₄(C) with β₄ C terminus produced significantly reduced cytoplasmic Ca2+ transients. Conversely, gain-of-function chimera β₄/β1a(C) with β1a C terminus completely restored cytoplasmic Ca2+ transients, DHPR tetrads, and motility. Furthermore, we found that the nonconserved, distal C terminus of β1a plays a pivotal role in reconstitution of DHPR tetrads and thus allosteric DHPR–RyR1 interaction, essential for skeletal muscle EC coupling.

Annonaceae, comprising approximately 107 genera and 2400 species, is the largest family among early-divergent Magnoliales. Previous studies have concentrated on the binding mechanism that holds together the four members of tetrads in Annonaceae. However, the development mechanisms of different tetrad types remain largely unknown. Mitrephora tomentosa was found to exhibit five permanent tetrad types, with two or three of them existing in the same microsporangium, which is ideal for studying the formation mechanisms of different permanent tetrad pollens in a single microsporangium and explaining the relationship between cytokinesis and pollen tetrad types. The ontogenetic development of the different tetrads in M. tomentosa was investigated using electron microscopy technologies, histochemical staining, and immunocytochemistry. During meiosis, pollen mother cells produce decussate and tetragonal tetrads by successive cytokinesis and produce tetrahedral and rhomboidal tetrads by simultaneous cytokinesis. Bidirectional callose deposition was observed in tetrahedral, tetragonal, rhomboidal, and decussate tetrads. The variations in the process of microsporogenesis randomly accumulate and manifest as different combinations of cytokinesis and callose deposition, leading to the formation of differently shaped tetrads. In mature permanent tetrad pollens, four microspores are connected by both simple cohesion and cytoplasmic channels, which also play an important role in maintaining the synchronization of the tetrad members.

During meiosis, interhomolog recombination produces crossovers and noncrossovers to create genetic diversity. Meiotic recombination frequency varies at multiple scales, with high subtelomeric recombination and suppressed centromeric recombination typical in many eukaryotes. During recombination, sister chromatids are tethered as loops to a polymerized chromosome axis, which, in plants, includes the ASY1 HORMA domain protein and REC8–cohesin complexes. Using chromatin immunoprecipitation, we show an ascending telomere-to-centromere gradient of ASY1 enrichment, which correlates strongly with REC8–cohesin ChIP-seq data. We mapped crossovers genome-wide in the absence of ASY1 and observe that telomere-led recombination becomes dominant. Surprisingly, asy1/+ heterozygotes also remodel crossovers toward subtelomeric regions at the expense of the pericentromeres. Telomeric recombination increases in asy1/+ occur in distal regions where ASY1 and REC8 ChIP enrichment are lowest in wild type. In wild type, the majority of crossovers show interference, meaning that they are more widely spaced along the chromosomes than expected by chance. To measure interference, we analyzed double crossover distances, MLH1 foci, and fluorescent pollen tetrads. Interestingly, while crossover interference is normal in asy1/+, it is undetectable in asy1 mutants, indicating that ASY1 is required to mediate crossover interference. Together, this is consistent with ASY1 antagonizing telomere-led recombination and promoting spaced crossover formation along the chromosomes via interference. These findings provide insight into the role of the meiotic axis in patterning recombination frequency within plant genomes.

Guanine quadruplexes (G4s) are important targets for cancer treatments as their stabilization has been associated with a reduction of telomere ends or a lower oncogene expression. Although less abundant than purely organic ligands, metal complexes have shown remarkable abilities to stabilize G4s, and a wide variety of techniques have been used to characterize the interaction between ligands and G4s. However, improper alignment between the large variety of experimental techniques and biological activities can lead to improper identification of top candidates, which hampers progress of this important class of G4 stabilizers. To address this, we first review the different techniques for their strengths and weaknesses to determine the interaction of the complexes with G4s, and provide a checklist to guide future developments towards comparable data. Then, we surveyed 74 metal-based ligands for G4s that have been characterized to the in vitro level. Of these complexes, we assessed which methods were used to characterize their G4-stabilizing capacity, their selectivity for G4s over double-stranded DNA (dsDNA), and how this correlated to bioactivity data. For the biological activity data, we compared activities of the G4-stabilizing metal complexes with that of cisplatin. Lastly, we formulated guidelines for future studies on G4-stabilizing metal complexes to further enable maturation of this field. Graphical abstract