Nigel P. Dyer

Conserved Noncoding Sequences Highlight Shared Components of Regulatory Networks in Dicotyledonous Plants

This study identifies regions of noncoding DNA in dicot plants that are likely to facilitate complex regulation of genes by binding multiple transcription factors. Regulatory mechanisms that the model organism Arabidopsis is likely to share with crop plants provide a focus for research that has real-world applications. Conserved noncoding sequences (CNSs) in DNA are reliable pointers to regulatory elements controlling gene expression. Using a comparative genomics approach with four dicotyledonous plant species (Arabidopsis thaliana, papaya [Carica papaya], poplar [Populus trichocarpa], and grape [Vitis vinifera]), we detected hundreds of CNSs upstream of Arabidopsis genes. Distinct positioning, length, and enrichment for transcription factor binding sites suggest these CNSs play a functional role in transcriptional regulation. The enrichment of transcription factors within the set of genes associated with CNS is consistent with the hypothesis that together they form part of a conserved transcriptional network whose function is to regulate other transcription factors and control development. We identified a set of promoters where regulatory mechanisms are likely to be shared between the model organism Arabidopsis and other dicots, providing areas of focus for further research.

Dynamic Distribution of SeqA Protein across the Chromosome of Escherichia coli K-12

ABSTRACT The bacterial SeqA protein binds to hemi-methylated GATC sequences that arise in newly synthesized DNA upon passage of the replication machinery. In Escherichia coli K-12, the single replication origin oriC is a well-characterized target for SeqA, which binds to multiple hemi-methylated GATC sequences immediately after replication has initiated. This sequesters oriC, thereby preventing reinitiation of replication. However, the genome-wide DNA binding properties of SeqA are unknown, and hence, here, we describe a study of the binding of SeqA across the entire Escherichia coli K-12 chromosome, using chromatin immunoprecipitation in combination with DNA microarrays. Our data show that SeqA binding correlates with the frequency and spacing of GATC sequences across the entire genome. Less SeqA is found in highly transcribed regions, as well as in the ter macrodomain. Using synchronized cultures, we show that SeqA distribution differs with the cell cycle. SeqA remains bound to some targets after replication has ceased, and these targets locate to genes encoding factors involved in nucleotide metabolism, chromosome replication, and methyl transfer. IMPORTANCE DNA replication in bacteria is a highly regulated process. In many bacteria, a protein called SeqA plays a key role by binding to newly replicated DNA. Thus, at the origin of DNA replication, SeqA binding blocks premature reinitiation of replication rounds. Although most investigators have focused on the role of SeqA at replication origins, it has long been suspected that SeqA has a more pervasive role. In this study, we describe how we have been able to identify scores of targets, across the entire Escherichia coli chromosome, to which SeqA binds. Using synchronously growing cells, we show that the distribution of SeqA between these targets alters as replication of the chromosome progresses. This suggests that sequential changes in SeqA distribution orchestrate a program of gene expression that ensures coordinated DNA replication and cell division. DNA replication in bacteria is a highly regulated process. In many bacteria, a protein called SeqA plays a key role by binding to newly replicated DNA. Thus, at the origin of DNA replication, SeqA binding blocks premature reinitiation of replication rounds. Although most investigators have focused on the role of SeqA at replication origins, it has long been suspected that SeqA has a more pervasive role. In this study, we describe how we have been able to identify scores of targets, across the entire Escherichia coli chromosome, to which SeqA binds. Using synchronously growing cells, we show that the distribution of SeqA between these targets alters as replication of the chromosome progresses. This suggests that sequential changes in SeqA distribution orchestrate a program of gene expression that ensures coordinated DNA replication and cell division.

Loss of endometrial plasticity in recurrent pregnancy loss

Menstruation drives cyclic activation of endometrial progenitor cells, tissue regeneration, and maturation of stromal cells, which differentiate into specialized decidual cells prior to and during pregnancy. Aberrant responsiveness of human endometrial stromal cells (HESCs) to deciduogenic cues is strongly associated with recurrent pregnancy loss (RPL), suggesting a defect in cellular maturation. MeDIP‐seq analysis of HESCs did not reveal gross perturbations in CpG methylation in RPL cultures, although quantitative differences were observed in or near genes that are frequently deregulated in vivo. However, RPL was associated with a marked reduction in methylation of defined CA‐rich motifs located throughout the genome but enriched near telomeres. Non‐CpG methylation is a hallmark of cellular multipotency. Congruently, we demonstrate that RPL is associated with a deficiency in endometrial clonogenic cell populations. Loss of epigenetic stemness features also correlated with intragenic CpG hypomethylation and reduced expression of HMGB2, coding high mobility group protein 2. We show that knockdown of this sequence‐independent chromatin protein in HESCs promotes senescence and impairs decidualization, exemplified by blunted time‐dependent secretome changes. Our findings indicate that stem cell deficiency and accelerated stromal senescence limit the differentiation capacity of the endometrium and predispose for pregnancy failure. Stem Cells 2016;34:346–356

CisGenome Browser

SUMMARY We present an open source, platform independent tool, called CisGenome Browser, which can work together with any other data analysis program to serve as a flexible component for genomic data visualization. It can also work by itself as a standalone genome browser. By working as a light-weight web server, CisGenome Browser is a convenient tool for data sharing between labs. It has features that are specifically designed for ultra high-throughput sequencing data visualization. AVAILABILITY http://biogibbs.stanford.edu/ approximately jiangh/browser/

A CMOS subscriber line audio processing circuit including adaptive balance

The design of a CMOS subscriber-line audio processing circuit including self-adaptive trans-hybrid balance has been described. Its transmission performance has been shown to exceed CCITT specifications with up to 12 dB of transmit gain and receive loss. This has been achieved using a single 5-V power supply while maintaining simple analog interfaces to a subscriber line interface circuit. This circuit will enable the design of programmable and flexible line cards, superior transmission performance over those realized with more conventional components.<<ETX>>

Variable structure motifs for transcription factor binding sites

BackgroundClassically, models of DNA-transcription factor binding sites (TFBSs) have been based on relatively few known instances and have treated them as sites of fixed length using position weight matrices (PWMs). Various extensions to this model have been proposed, most of which take account of dependencies between the bases in the binding sites. However, some transcription factors are known to exhibit some flexibility and bind to DNA in more than one possible physical configuration. In some cases this variation is known to affect the function of binding sites. With the increasing volume of ChIP-seq data available it is now possible to investigate models that incorporate this flexibility. Previous work on variable length models has been constrained by: a focus on specific zinc finger proteins in yeast using restrictive models; a reliance on hand-crafted models for just one transcription factor at a time; and a lack of evaluation on realistically sized data sets.ResultsWe re-analysed binding sites from the TRANSFAC database and found motivating examples where our new variable length model provides a better fit. We analysed several ChIP-seq data sets with a novel motif search algorithm and compared the results to one of the best standard PWM finders and a recently developed alternative method for finding motifs of variable structure. All the methods performed comparably in held-out cross validation tests. Known motifs of variable structure were recovered for p53, Stat5a and Stat5b. In addition our method recovered a novel generalised version of an existing PWM for Sp1 that allows for variable length binding. This motif improved classification performance.ConclusionsWe have presented a new gapped PWM model for variable length DNA binding sites that is not too restrictive nor over-parameterised. Our comparison with existing tools shows that on average it does not have better predictive accuracy than existing methods. However, it does provide more interpretable models of motifs of variable structure that are suitable for follow-up structural studies. To our knowledge, we are the first to apply variable length motif models to eukaryotic ChIP-seq data sets and consequently the first to show their value in this domain. The results include a novel motif for the ubiquitous transcription factor Sp1.

An alignment-free model for comparison of regulatory sequences

MOTIVATION Some recent comparative studies have revealed that regulatory regions can retain function over large evolutionary distances, even though the DNA sequences are divergent and difficult to align. It is also known that such enhancers can drive very similar expression patterns. This poses a challenge for the in silico detection of biologically related sequences, as they can only be discovered using alignment-free methods. RESULTS Here, we present a new computational framework called Regulatory Region Scoring (RRS) model for the detection of functional conservation of regulatory sequences using predicted occupancy levels of transcription factors of interest. We demonstrate that our model can detect the functional and/or evolutionary links between some non-alignable enhancers with a strong statistical significance. We also identify groups of enhancers that are likely to be similarly regulated. Our model is motivated by previous work on prediction of expression patterns and it can capture similarity by strong binding sites, weak binding sites and even the statistically significant absence of sites. Our results support the hypothesis that weak binding sites contribute to the functional similarity of sequences. Our model fills a gap between two families of models: detailed, data-intensive models for the prediction of precise spatio-temporal expression patterns on the one side, and crude, generally applicable models on the other side. Our model borrows some of the strengths of each group and addresses their drawbacks. AVAILABILITY The RRS source code is freely available upon publication of this manuscript: http://www2.warwick.ac.uk/fac/sci/systemsbiology/staff/ott/tools_and_software/rrs.

Success after failure: the role of endometrial stem cells in recurrent miscarriage

Endometrial stem-like cells, including mesenchymal stem cells (MSCs) and epithelial progenitor cells, are essential for cyclic regeneration of the endometrium following menstrual shedding. Emerging evidence indicates that endometrial MSCs (eMSCs) constitute a dynamic population of cells that enables the endometrium to adapt in response to a failed pregnancy. Recurrent miscarriage is associated with relative depletion of endometrial eMSCs, which not only curtails the intrinsic ability of the endometrium to adapt to reproductive failure but also compromises endometrial decidualization, an obligatory transformation process for embryo implantation. These novel findings should pave the way for more effective screening of women at risk of pregnancy failure before conception.

Tubulin and its prokaryotic homologue FtsZ: a structural and functional comparison.

Microtubules are one of the three primary constituents of the eukaryotic cytoskeleton and are constructed from the protein tubulin. FtsZ is a close structural homologue of tubulin within prokaryotes, and plays an important structural role during cell division. This article compares what is known about the structures that these two homologues are able to form in vivo and in vitro and examines the evidence that the water in the immediate vicinity of the structures, particularly in microtubules, may play an important role in their formation and stability. The article then examines evidence that this hydration layer might help our understanding of how the structures formed by tubulin and FtsZ are stabilised by associated proteins and selected cations. The article then considers recent studies of the charge distribution and dipole moments of tubulin and extends this work to include the electrostatic characteristics of FtsZ. There is then an examination of the ways in which the electrostatic similarities and differences between the two proteins might be related to the similarities and differences in the filamentary structures that they form.

Artifacts in the data of Hu et al.

To the Editor: The recent study by Hu et al. reports 3,667 nuclear integration events of human papillomavirus (HPV) in 135 cervical cancer samples identified on the basis of hybrid human-viral DNA fragments1. Widespread microhomology at integration sites is reported, suggesting a microhomologybased mechanism as the main driver of viral insertions. We identify 87% of the reported integration sites as likely experimental and computational artifacts. As most results were based on the identified insertions, our finding of artifacts draws most of the conclusions of this publication into question. We exemplify this by showing that sites of artifacts more frequently feature microhomology than genuine sites, with the majority of genuine sites not featuring microhomology. We established a data analysis pipeline that achieved good agreement with the originally reported results. Data for all tumor samples were analyzed, whereas data for cell lines were not considered here (excluding 121 reported insertion sites from the analysis). As DNA fragments are PCR amplified before sequencing, one frequently obtains multiple reads corresponding to the same original fragment (Fig. 1). Hu et al. tackled this problem by filtering out duplicate reads and implementing a threshold based on read number. However, as this filter ignores the occurrence of sequencing and PCR errors, it is insufficient to completely exclude amplification bias. For example, filtering out the identical reads among the reads shown in Figure 1 still leaves multiple reads, as some contain mismatching bases. We filtered reads on the basis of their mapped genomic position and found 72% of sites to be supported by single original fragments and, therefore, indistinguishable from ligation events of unrelated fragments during sample processing. As a computational control, we repeated the analysis swapping the viral genome with the mitochondrial genome, which is physically separate from the nuclear genome. We found that 0.024% (median across samples) of mitochondrial reads corresponded to nuclear-mitochondrial hybrid fragments, closely matching the rate of nuclear-HPV hybrids (median of 0.017%) mapping to single-fragment loci. Therefore, if single-fragment loci were accepted as evidence, one would need to conclude that mitochondrial genomes integrate at a similar rate to viruses. Removing single-fragment loci left 28% of the reported integration sites for further consideration. Within a hybrid viral-human DNA fragment, the point of transition from one genomic sequence to the other identifies the exact insertion position. Where both ends of a fragment are sequenced without covering the transition point, the insertion position cannot be resolved exactly. Applying Occam’s razor, we explained the data using a minimal number of insertions and found that only 50% of the insertion positions reported for multi-fragment loci could be justified. Where insertions were detected at exactly the same genomic position in multiple samples, cross-contamination seemed a more likely explanation than extremely precise viral targeting. In 55% of the cases where samples shared a reported insertion site, these samples had consecutive or nearly consecutive IDs, which is highly improbable if the process of taking and analyzing samples is unrelated to Artifacts in the data of Hu et al.