Triple-helix potential of the mouse genome

Certain DNA sequences, including mirror-symmetric polypyrimidine/polypurine runs, are capable of folding into a triple-helix-containing non-B-form DNA structure called H-DNA. Such H-DNA-forming sequences are frequent in many eukaryotic genomes, including in mammals, and multiple lines of evidence indicate that these motifs are mutagenic and can impinge on DNA replication, transcription, and other aspects of genome function. In this study, we show that the triplex-forming potential of H-DNA motifs in the mouse genome can be evaluated using S1-sequencing (S1-seq), which uses the single-stranded DNA (ssDNA)-specific nuclease S1 to generate deep-sequencing libraries that report on the position of ssDNA throughout the genome. When S1-seq was applied to genomic DNA isolated from mouse testis cells and splenic B cells, we observed prominent clusters of S1-seq reads that appeared to be independent of endogenous double-strand breaks, that coincided with H-DNA motifs, and that correlated strongly with the triplex-forming potential of the motifs. Fine-scale patterns of S1-seq reads, including a pronounced strand asymmetry in favor of centrally-positioned reads on the pyrimidine-containing strand, suggested that this S1-seq signal is specific for one of the four possible isomers of H-DNA (H-y5). By leveraging the abundance and complexity of naturally occurring H-DNA motifs across the mouse genome, we further defined how polypyrimidine repeat length and the presence of repeat-interrupting substitutions modify the structure of H-DNA. This study provides a new approach for studying DNA secondary structure genome wide at high spatial resolution.

Microarray testing in patients with systemic lupus erythematosus identifies a high prevalence of CpG DNA-binding antibodies

ObjectiveMany autoantibodies are known to be associated with SLE, although their role in clinical practice is limited because of low sensitivity and weak associations with clinical manifestations. There has been great interest in the discovery of new autoantibodies to use in clinical practice. In this study, we investigated 57 new and known antibodies and their potential for diagnostics or risk stratification.MethodsBetween 2014 and 2017, residual sera of all anti-dsDNA tests in the UMC Utrecht were stored in a biobank. This included sera of patients with SLE, patients with a diagnosis of another immune-mediated inflammatory disease (IMID), patients with low (non-IMID) or medium levels of clinical suspicion of SLE but no IMID diagnosis (Rest), and self-reported healthy blood bank donors. Diagnosis and (presence of) symptoms at each blood draw were retrospectively assessed in the patient records with the Utrecht Patient-Oriented Database using a newly developed text mining algorithm. Sera of patients were analysed for the presence of 57 autoantibodies with a custom-made immunofluorescent microarray. Signal intensity cut-offs for all antigens on the microarray were set to the 95th percentile of the non-IMID control group. Differences in prevalence of autoantibodies between patients with SLE and control groups were assessed.ResultsAutoantibody profiles of 483 patients with SLE were compared with autoantibody profiles of 1397 patients from 4 different control groups. Anti-dsDNA was the most distinguishing feature between patients with SLE and other patients, followed by antibodies against Cytosine-phosphate-Guanine (anti-CpG) DNA motifs (p<0.0001). Antibodies against CMV (cytomegalovirus) and ASCA (anti-Saccharomyces cerevisiae antibodies) were more prevalent in patients with SLE with (a history of) lupus nephritis than patients with SLE without nephritis.ConclusionAntibodies against CpG DNA motifs are prevalent in patients with SLE. Anti-CMV antibodies are associated with lupus nephritis.

The rs5918 polymorphism in the ITGB3 gene increases the risk for preeclampsia in pregnant women with fetal growth retardation

Aim. To assess the relationship of rs5918 ITGB3, rs1126643 ITGA2 and rs5985 F13A1 polymorphic loci with the risk for preeclampsia (PE) in pregnant women with fetal growth retardation (FGR). Materials and methods. The study included 272 pregnant women, of which 76 had a combination of PE and FGR and 196 had FGR. In the studied groups, genetic testing was carried out for three polymorphic loci of candidate genes for hereditary thrombophilia (rs5918 ITGB3, rs1126643 ITGA2, and rs5985 F13A1). Results. The rs5918 genetic variant in the ITGB3 gene is associated with the development of PE in pregnant women with FGR: C allele of rs5918 ITGB3 increases the risk for this complication of pregnancy by 1,8 times (OR 1.761.77, p0.036, pperm0.038). The rs5918 polymorphism determines an increase in the affinity of DNA motifs for seven transcription factors (BDP1, ELF1, IRF, NRSF, Pax-5, Sp1, and Zfx), is a missense mutation and causes the Leu59Pro amino acid substitution in the 3 subunit of integrin, is multidirectionally associated with the expression of five genes (EFCAB13, TBKBP1, NPEPPS, MRPL45P2, THCAT158) and alternative splicing of two genes (EFCAB13, MRPL45P2), is located in the region of functionally important DNA regions (promoters and enhancers) in cell cultures and organs which are pathogenetically important for the formation of PE and FGR. Conclusion. The rs5918 polymorphism in the ITGB3 gene increases the risk for PE in pregnant women with FGR.

pH-Responsive DNA Motif: From Rational Design to Analytical Applications

pH-responsive DNA motifs have attracted substantial attention attributed to their high designability and versatility of DNA chemistry. Such DNA motifs typically exploit DNA secondary structures that exhibit pH response properties because of the presence of specific protonation sites. In this review, we briefly summarized second structure-based pH-responsive DNA motifs, including triplex DNA, i-motif, and A+-C mismatch base pair-based DNA devices. Finally, the challenges and prospects of pH-responsive DNA motifs are also discussed.

Aqueous Triple-Phase System in Microwell Array for Generating Uniform-Sized DNA Hydrogel Particles

DNA hydrogels are notable for their biocompatibility and ability to incorporate DNA information and computing properties into self-assembled micrometric structures. These hydrogels are assembled by the thermal gelation of DNA motifs, a process which requires a high salt concentration and yields polydisperse hydrogel particles, thereby limiting their application and physicochemical characterization. In this study, we demonstrate that single, uniform DNA hydrogel particles can form inside aqueous/aqueous two-phase systems (ATPSs) assembled in a microwell array. In this process, uniform dextran droplets are formed in a microwell array inside a microfluidic device. The dextran droplets, which contain DNA motifs, are isolated from each other by an immiscible PEG solution containing magnesium ions and spermine, which enables the DNA hydrogel to undergo gelation. Upon thermal annealing of the device, we observed the formation of an aqueous triple-phase system in which uniform DNA hydrogel particles (the innermost aqueous phase) resided at the interface of the aqueous two-phase system of dextran and PEG. We expect ATPS microdroplet arrays to be used to manufacture other hydrogel microparticles and DNA/dextran/PEG aqueous triple-phase systems to serve as a highly parallel model for artificial cells and membraneless organelles.

Z-DNA as a Touchstone for Additive Empirical Force Fields and a Refinement of the Alpha/Gamma DNA torsions for AMBER

Although current AMBER force fields are relatively accurate for canonical B-DNA, many non-canonical structures are still described incorrectly. As non-canonical motifs are attracting increasing attention due to the role they play in living organisms, further improvement is desirable. Here, we have chosen Z-DNA molecule, can be considered a touchstone of the universality of empirical force fields, since the non-canonical α and γ backbone conformations native to Z-DNA are also found in protein-DNA complexes, i-motif DNA and other non-canonical DNAs. We show that spurious α/γ conformations occurring in simulations with current AMBER force fields, OL15 and bsc1, are largely due to inaccurate α/γ parameterization. Moreover, stabilization of native Z-DNA substates involving γ = trans conformations appears to be in conflict with the correct description of the canonical B-DNA structure. Because the balance of the native and spurious conformations is influenced by non-additive effects, this is a difficult case for an additive dihedral energy scheme such as AMBER. We propose new α/γ parameters, denoted OL21, and show that they improve the stability of native α/γ Z-DNA substates while keeping the canonical DNA description virtually unchanged, and thus represent a reasonable compromise within the additive force field framework. Although further extensive testing is needed, the new modification appears to be a promising step towards a more reliable description of non-canonical DNA motifs and provides the best performance for Z-DNA molecules among current AMBER force fields.

CpGmotifs: a tool to discover DNA motifs associated to CpG methylation events

Background The investigation of molecular alterations associated with the conservation and variation of DNA methylation in eukaryotes is gaining interest in the biomedical research community. Among the different determinants of methylation stability, the DNA composition of the CpG surrounding regions has been shown to have a crucial role in the maintenance and establishment of methylation statuses. This aspect has been previously characterized in a quantitative manner by inspecting the nucleotidic composition in the region. Research in this field still lacks a qualitative perspective, linked to the identification of certain sequences (or DNA motifs) related to particular DNA methylation phenomena. Results Here we present a novel computational strategy based on short DNA motif discovery in order to characterize sequence patterns related to aberrant CpG methylation events. We provide our framework as a user-friendly, shiny-based application, CpGmotifs, to easily retrieve and characterize DNA patterns related to CpG methylation in the human genome. Our tool supports the functional interpretation of deregulated methylation events by predicting transcription factors binding sites (TFBS) encompassing the identified motifs. Conclusions CpGmotifs is an open source software. Its source code is available on GitHub https://github.com/Greco-Lab/CpGmotifs and a ready-to-use docker image is provided on DockerHub at https://hub.docker.com/r/grecolab/cpgmotifs.

Molecular Composition of Heterochromatin and Its Contribution to Chromosome Variation in the Microtus thomasi/Microtus atticus Species Complex

The voles of the Microtus thomasi/M. atticus species complex demonstrate a remarkable variability in diploid chromosomal number (2n = 38–44 chromosomes) and sex chromosome morphology. In the current study, we examined by in situ hybridization the topology of four satellite DNA motifs (Msat-160, Mth-Alu900, Mth-Alu2.2, TTAGGG telomeric sequences) and two transposons (LINE, SINE) on the karyotypes of nine chromosome races (i.e., populations with unique cytogenetic traits) of Microtus thomasi, and two chromosomal races of M. atticus. According to the topology of the repetitive DNA motifs, we were able to identify six types of biarmed chromosomes formed from either Robertsonian or/and tandem fusions. In addition, we identified 14 X chromosome variants and 12 Y chromosome variants, and we were able to reconstruct their evolutionary relations, caused mainly by distinct mechanisms of amplification of repetitive DNA elements, including the telomeric sequences. Our study used the model of the Microtus thomasi/M. atticus species complex to explore how repetitive centromeric content can alter from chromosomal rearrangements and can shape the morphology of sex chromosomes, resulting in extensive inter-species cytogenetic variability.

DeepMotifSyn: a deep learning approach to synthesize heterodimeric DNA motifs

MotivationThe cooperativity of transcription factors (TFs) is a widespread phenomenon in the gene regulation system. However, the interaction patterns between TF binding motifs remain elusive. The recent high-throughput assays, CAP-SELEX, have identified over 600 composite DNA sites (i.e. heterodimeric motifs) bound by cooperative TF pairs. However, there are over 25,000 inferentially effective heterodimeric TFs in human cell. It is not practically feasible to validate all heterodimeric motifs due to cost and labour. Therefore, it is highly demanding to develop a fast and accurate computational tool for heterodimeric motif synthesis.ResultsWe introduce DeepMotifSyn, a deep-learning-based tool for synthesizing heterodimeric motifs from monomeric motif pairs. Specifically, DeepMotifSyn is composed of heterodimeric motif generator and evaluator. The generator is a U-Net-based neural network that can synthesize heterodimeric motifs from aligned motif pairs. The evaluator is a machine-learning-based model that can score the generated heterodimeric motif candidates based on the motif sequence features. Systematic evaluations on CAP-SELEX data illustrates that DeepMotif-Syn significantly outperforms the current state-of-the-art predictors. In addition, DeepMotifSyn can synthesize multiple heterodimeric motifs with different orientation and spacing settings. Such a feature can address the shortcomings of previous models. We believe Deep-MotifSyn is a more practical and reliable model than current predictors on heterodimeric motif synthesis.Availability and implementationThe software is freely available at https://github.com/JasonLinjc/deepMotifSyn.