Smc5/6: a link between DNA repair and unidirectional replication?

Of the three structural maintenance of chromosome (SMC) complexes, two directly regulate chromosome dynamics. The third, Smc5/6, functions mainly in homologous recombination and in completing DNA replication. The literature suggests that Smc5/6 coordinates DNA repair, in part through post-translational modification of uncharacterized target proteins that can dictate their subcellular localization, and that Smc5/6 also functions to establish DNA-damage-dependent cohesion. A nucleolar-specific Smc5/6 function has been proposed because Smc5/6 yeast mutants display penetrant phenotypes of ribosomal DNA (rDNA) instability. rDNA repeats are replicated unidirectionally. Here, we propose that unidirectional replication, combined with global Smc5/6 functions, can explain the apparent rDNA specificity

Multiscale photosynthetic exciton transfer

Photosynthetic light harvesting provides a natural blueprint for bioengineered and biomimetic solar energy and light detection technologies. Recent evidence suggests some individual light harvesting protein complexes (LHCs) and LHC subunits efficiently transfer excitons towards chemical reaction centers (RCs) via an interplay between excitonic quantum coherence, resonant protein vibrations, and thermal decoherence. The role of coherence in vivo is unclear however, where excitons are transferred through multi-LHC/RC aggregates over distances typically large compared with intra-LHC scales. Here we assess the possibility of long-range coherent transfer in a simple chromophore network with disordered site and transfer coupling energies. Through renormalization we find that, surprisingly, decoherence is diminished at larger scales, and long-range coherence is facilitated by chromophoric clustering. Conversely, static disorder in the site energies grows with length scale, forcing localization. Our results suggest sustained coherent exciton transfer may be possible over distances large compared with nearest-neighbour (n-n) chromophore separations, at physiological temperatures, in a clustered network with small static disorder. This may support findings suggesting long-range coherence in algal chloroplasts, and provides a framework for engineering large chromophore or quantum dot high-temperature exciton transfer networks.Comment: 9 pages, 6 figures. A significantly updated version is now published online by Nature Physics (2012

Identification of disease-causing genes using microarray data mining and gene ontology

Background: One of the best and most accurate methods for identifying disease-causing genes is monitoring gene expression values in different samples using microarray technology. One of the shortcomings of microarray data is that they provide a small quantity of samples with respect to the number of genes. This problem reduces the classification accuracy of the methods, so gene selection is essential to improve the predictive accuracy and to identify potential marker genes for a disease. Among numerous existing methods for gene selection, support vector machine-based recursive feature elimination (SVMRFE) has become one of the leading methods, but its performance can be reduced because of the small sample size, noisy data and the fact that the method does not remove redundant genes. Methods: We propose a novel framework for gene selection which uses the advantageous features of conventional methods and addresses their weaknesses. In fact, we have combined the Fisher method and SVMRFE to utilize the advantages of a filtering method as well as an embedded method. Furthermore, we have added a redundancy reduction stage to address the weakness of the Fisher method and SVMRFE. In addition to gene expression values, the proposed method uses Gene Ontology which is a reliable source of information on genes. The use of Gene Ontology can compensate, in part, for the limitations of microarrays, such as having a small number of samples and erroneous measurement results. Results: The proposed method has been applied to colon, Diffuse Large B-Cell Lymphoma (DLBCL) and prostate cancer datasets. The empirical results show that our method has improved classification performance in terms of accuracy, sensitivity and specificity. In addition, the study of the molecular function of selected genes strengthened the hypothesis that these genes are involved in the process of cancer growth. Conclusions: The proposed method addresses the weakness of conventional methods by adding a redundancy reduction stage and utilizing Gene Ontology information. It predicts marker genes for colon, DLBCL and prostate cancer with a high accuracy. The predictions made in this study can serve as a list of candidates for subsequent wet-lab verification and might help in the search for a cure for cancers

Ovine pedomics : the first study of the ovine foot 16S rRNA-based microbiome

We report the first study of the bacterial microbiome of ovine interdigital skin based on 16S rRNA by pyrosequencing and conventional cloning with Sanger-sequencing. Three flocks were selected, one a flock with no signs of footrot or interdigital dermatitis, a second flock with interdigital dermatitis alone and a third flock with both interdigital dermatitis and footrot. The sheep were classified as having either healthy interdigital skin (H), interdigital dermatitis (ID) or virulent footrot (VFR). The ovine interdigital skin bacterial community varied significantly by flock and clinical condition. The diversity and richness of operational taxonomic units was greater in tissue from sheep with ID than H or VFR affected sheep. Actinobacteria, Bacteriodetes, Firmicutes and Proteobacteria were the most abundant phyla comprising 25 genera. Peptostreptococcus, Corynebacterium and Staphylococcus were associated with H, ID and VFR respectively. Sequences of Dichelobacter nodosus, the causal agent of ovine footrot, were not amplified due to mismatches in the 16S rRNA universal forward primer (27F). A specific real time PCR assay was used to demonstrate the presence of D. nodosus which was detected in all samples including the flock with no signs of ID or VFR. Sheep with ID had significantly higher numbers of D. nodosus (104-109 cells/g tissue) than those with H or VFR feet

Multiple ITS Copies Reveal Extensive Hybridization within Rheum (Polygonaceae), a Genus That Has Undergone Rapid Radiation

During adaptive radiation events, characters can arise multiple times due to parallel evolution, but transfer of traits through hybridization provides an alternative explanation for the same character appearing in apparently non-sister lineages. The signature of hybridization can be detected in incongruence between phylogenies derived from different markers, or from the presence of two divergent versions of a nuclear marker such as ITS within one individual.In this study, we cloned and sequenced ITS regions for 30 species of the genus Rheum, and compared them with a cpDNA phylogeny. Seven species contained two divergent copies of ITS that resolved in different clades from one another in each case, indicating hybridization events too recent for concerted evolution to have homogenised the ITS sequences. Hybridization was also indicated in at least two further species via incongruence in their position between ITS and cpDNA phylogenies. None of the ITS sequences present in these nine species matched those detected in any other species, which provides tentative evidence against recent introgression as an explanation. Rheum globulosum, previously indicated by cpDNA to represent an independent origin of decumbent habit, is indicated by ITS to be part of clade of decumbent species, which acquired cpDNA of another clade via hybridization. However decumbent and glasshouse morphology are confirmed to have arisen three and two times, respectively.These findings suggested that hybridization among QTP species of Rheum has been extensive, and that a role of hybridization in diversification of Rheum requires investigation

A Minimal Threshold of c-di-GMP Is Essential for Fruiting Body Formation and Sporulation in Myxococcus xanthus

Generally, the second messenger bis-(3’-5’)-cyclic dimeric GMP (c-di-GMP) regulates the switch between motile and sessile lifestyles in bacteria. Here, we show that c-di-GMP is an essential regulator of multicellular development in the social bacterium Myxococcus xanthus. In response to starvation, M. xanthus initiates a developmental program that culminates in formation of spore-filled fruiting bodies. We show that c-di-GMP accumulates at elevated levels during development and that this increase is essential for completion of development whereas excess c-di-GMP does not interfere with development. MXAN3735 (renamed DmxB) is identified as a diguanylate cyclase that only functions during development and is responsible for this increased c-di-GMP accumulation. DmxB synthesis is induced in response to starvation, thereby restricting DmxB activity to development. DmxB is essential for development and functions downstream of the Dif chemosensory system to stimulate exopolysaccharide accumulation by inducing transcription of a subset of the genes encoding proteins involved in exopolysaccharide synthesis. The developmental defects in the dmxB mutant are non-cell autonomous and rescued by co-development with a strain proficient in exopolysaccharide synthesis, suggesting reduced exopolysaccharide accumulation as the causative defect in this mutant. The NtrC-like transcriptional regulator EpsI/Nla24, which is required for exopolysaccharide accumulation, is identified as a c-diGMP receptor, and thus a putative target for DmxB generated c-di-GMP. Because DmxB can be—at least partially—functionally replaced by a heterologous diguanylate cyclase, these results altogether suggest a model in which a minimum threshold level of c-di-GMP is essential for the successful completion of multicellular development in M. xanthus

Microfluidic Perfusion for Regulating Diffusible Signaling in Stem Cells

Background Autocrine & paracrine signaling are widespread both in vivo and in vitro, and are particularly important in embryonic stem cell (ESC) pluripotency and lineage commitment. Although autocrine signaling via fibroblast growth factor-4 (FGF4) is known to be required in mouse ESC (mESC) neuroectodermal specification, the question of whether FGF4 autocrine signaling is sufficient, or whether other soluble ligands are also involved in fate specification, is unknown. The spatially confined and closed-loop nature of diffusible signaling makes its experimental control challenging; current experimental approaches typically require prior knowledge of the factor/receptor in order to modulate the loop. A new approach explored in this work is to leverage transport phenomena at cellular resolution to downregulate overall diffusible signaling through the physical removal of cell-secreted ligands. Methodology/Principal Findings We develop a multiplex microfluidic platform to continuously remove cell-secreted (autocrine\paracrine) factors to downregulate diffusible signaling. By comparing cell growth and differentiation in side-by-side chambers with or without added cell-secreted factors, we isolate the effects of diffusible signaling from artifacts such as shear, nutrient depletion, and microsystem effects, and find that cell-secreted growth factor(s) are required during neuroectodermal specification. Then we induce FGF4 signaling in minimal chemically defined medium (N2B27) and inhibit FGF signaling in fully supplemented differentiation medium with cell-secreted factors to determine that the non-FGF cell-secreted factors are required to promote growth of differentiating mESCs. Conclusions/Significance Our results demonstrate for the first time that flow can downregulate autocrine\paracrine signaling and examine sufficiency of extracellular factors. We show that autocrine\paracrine signaling drives neuroectodermal commitment of mESCs through both FGF4-dependent and -independent pathways. Overall, by uncovering autocrine\paracrine processes previously hidden in conventional culture systems, our results establish microfluidic perfusion as a technique to study and manipulate diffusible signaling in cell systems.National Institutes of Health (U.S.) (NIH grant No. EB007278)Swiss National Science FoundationSwiss National Science Foundatio

The Complete Genome Sequence of Escherichia coli EC958: A High Quality Reference Sequence for the Globally Disseminated Multidrug Resistant E. coli O25b:H4-ST131 Clone

Escherichia coli ST131 is now recognised as a leading contributor to urinary tract and bloodstream infections in both community and clinical settings. Here we present the complete, annotated genome of E. coli EC958, which was isolated from the urine of a patient presenting with a urinary tract infection in the Northwest region of England and represents the most well characterised ST131 strain. Sequencing was carried out using the Pacific Biosciences platform, which provided sufficient depth and read-length to produce a complete genome without the need for other technologies. The discovery of spurious contigs within the assembly that correspond to site-specific inversions in the tail fibre regions of prophages demonstrates the potential for this technology to reveal dynamic evolutionary mechanisms. E. coli EC958 belongs to the major subgroup of ST131 strains that produce the CTX-M-15 extended spectrum β-lactamase, are fluoroquinolone resistant and encode the fimH30 type 1 fimbrial adhesin. This subgroup includes the Indian strain NA114 and the North American strain JJ1886. A comparison of the genomes of EC958, JJ1886 and NA114 revealed that differences in the arrangement of genomic islands, prophages and other repetitive elements in the NA114 genome are not biologically relevant and are due to misassembly. The availability of a high quality uropathogenic E. coli ST131 genome provides a reference for understanding this multidrug resistant pathogen and will facilitate novel functional, comparative and clinical studies of the E. coli ST131 clonal lineage

Controlled Orientation of Active Sites in a Nanostructured Multienzyme Complex

Multistep cascade reactions in nature maximize reaction efficiency by co-assembling related enzymes. Such organization facilitates the processing of intermediates by downstream enzymes. Previously, the studies on multienzyme nanocomplexes assembled on DNA scaffolds demonstrated that closer interenzyme distance enhances the overall reaction efficiency. However, it remains unknown how the active site orientation controlled at nanoscale can have an effect on multienzyme reaction. Here, we show that controlled alignment of active sites promotes the multienzyme reaction efficiency. By genetic incorporation of a non-natural amino acid and two compatible bioorthogonal chemistries, we conjugated mannitol dehydrogenase to formate dehydrogenase with the defined active site arrangement with the residue-level accuracy. The study revealed that the multienzyme complex with the active sites directed towards each other exhibits four-fold higher relative efficiency enhancement in the cascade reaction and produces 60% more D-mannitol than the other complex with active sites directed away from each other.ope

Undernourishment and Yersinia enterocolitica

Previous studies have demonstrated that the longitudinal smooth muscle of rabbits infected with Yersinia enterocolitica and undernourished because of reduced food intake exhibit a significantly reduced ability to develop tension in response to carbachol compared with pair-fed animals, which are uninfected but equivalently undernourished. To determine whether the alteration in smooth muscle contractility results from changes in cell number (hypo- or hyperplasia), or in contractile protein content or isoform distribution, New Zealand White rabbits (600 to 1000 g) were randomly assigned to one of three treatment groups: infected, pair-fed or control. Tissue contractility was measured, morphometric studies were performed and immunoassays were developed for the measurement of total actin, gamma-enteric and alpha-vascular isoactins, and myosin heavy chain. Consistent with what was found in previous reports, the contractility of longitudinal smooth muscle in response to carbachol was found to increase in pair-fed animals and to decrease in Y enterocolitica-infected animals. There was no significant change in the proportional thickness of the ileal longitudinal smooth muscle coat, and the number of cross-sectioned longitudinal smooth muscle cells/mm2 was not significantly different in infected, pair-fed or control tissues. Immunoassay indicated that the proportion of each specific contractile protein, relative to total protein content in the muscularis propria, was unaffected by Y enterocolitica infection or by pair-feeding. Thus, the alterations in intestinal longitudinal smooth muscle function observed after Y enterocolitica infection were concluded not to be associated with tissue hypo- or hyperplasia, or changes in the total content or isoform distribution of contractile proteins in the muscularis propria