Holliday junction resolvase in Schizosaccharomyces pombe has identical endonuclease activity to the CCE1 homologue YDC2

A novel Holliday junction resolving activity has been identified in fractionated cell extracts of the fission yeast Schizosaccharomyces pombe . The enzyme catalyses endonucleolytic cleavage of Holliday junction-containing chi DNA and synthetic four-way DNA junctions. The activity cuts with high specificity a synthetic four-way junction containing a 12 bp core of homologous sequences but has no activity on another four-way junction (with a fixed crossover point), a three-way junction, linear duplex DNA or duplex DNA containing six mismatched nucleotides in the centre. The major cleavage sites map as single nicks in the vicinity of the crossover point, 3' of a thymidine residue. These data indicate that the activity has a strong DNA structure selectivity as well as a limited sequence preference; features similar to the Holliday junction resolving enzymes RuvC of Escherichia coli and the mitochondrial CCE1 (cruciform-cuttingenzyme 1) of Saccharomyces cerevisiae. A putative homologue of CCE1 in S.pombe (YDC2_SCHPO) has been identified through a search of the sequence database. The open reading frame of this gene has been cloned and the encoded protein, YDC2, expressed in E.coli . The purified recombinant YDC2 exhibits Holliday junction resolvase activity and is, therefore, a functional S.pombe homologue of CCE1. The resolvase YDC2 shows the same substrate specificity and produces identical cleavage sites as the activity obtained from S. pombe cells. Both YDC2 and the cellular activity cleave Holliday junctions in both orientations to give nicks that can be ligated in vitro. The partially purified Holliday junction resolving enzyme in fission yeast is biochemically indistinguishable from recombinant YDC2 and appears to be the same protein

Biochemical characterization and DNA repair pathway interactions of Mag1-mediated base excision repair in Schizosaccharomyces pombe

The Schizosaccharomyces pombe mag1 gene encodes a DNA repair enzyme with sequence similarity to the AlkA family of DNA glycosylases, which are essential for the removal of cytotoxic alkylation products, the premutagenic deamination product hypoxanthine and certain cyclic ethenoadducts such as ethenoadenine. In this paper, we have purified the Mag1 protein and characterized its substrate specificity. It appears that the substrate range of Mag1 is limited to the major alkylation products, such as 3-mA, 3-mG and 7-mG, whereas no significant activity was found towards deamination products, ethenoadducts or oxidation products. The efficiency of 3-mA and 3-mG removal was 5–10 times slower for Mag1 than for Escherichia coli AlkA whereas the rate of 7-mG removal was similar to the two enzymes. The relatively low efficiency for the removal of cytotoxic 3-methylpurines is consistent with the moderate sensitivity of the mag1 mutant to methylating agents. Furthermore, we studied the initial steps of Mag1-dependent base excision repair (BER) and genetic interactions with other repair pathways by mutant analysis. The double mutants mag1 nth1, mag1 apn2 and mag1 rad2 displayed increased resistance to methyl methanesulfonate (MMS) compared with the single mutants nth1, apn2 and rad2, respectively, indicating that Mag1 initiates both short-patch (Nth1-dependent) and long-patch (Rad2-dependent) BER of MMS-induced damage. Spontaneous intrachromosomal recombination frequencies increased 3-fold in the mag1 mutant suggesting that Mag1 and recombinational repair (RR) are both involved in repair of alkylated bases. Finally, we show that the deletion of mag1 in the background of rad16, nth1 and rad2 single mutants reduced the total recombination frequencies of all three double mutants, indicating that abasic sites formed as a result of Mag1 removal of spontaneous base lesions are substrates for nucleotide excision repair, long- and short-patch BER and RR

A 10-YEARS LONGITUDINAL STUDY OF ADOLESCENT DIABETICS WITH INITIAL NEPHROPATHY

The data from a 10-years prospective study among 85 boys and 64 girls, mean age 14,1 ±3,8 years with mean diabetes duration 8,45 ±3,7 years are analyzed. Using a complex prospective screening we found the frequency of microalbuminuria (MAU) and estimated some parameters of progression of the renal damage. The patients were divided into 3 groups: no MAU, with MAU and with clinical signs of nephropathy. We traced the dynamics of glomerular fdtration, albumin excretion and arterial pressure. The effect of ACEIs was according to the level of proteinuria. In diabetics with macroalbuminuria a short-lasting effect run out to the end of the 2-nd year. In MAU-positive ptients ACEIs led to delay of nephropathy for 10-years in 86,48%, progression in 8,1% and regression to normoalbuminuria in 5,4%. In 15% of all the diabetics we found a significant raise of proteinuria. In 21,73% of patients with "borderline" albuminuria there was a progression after the 5-th year with a cumulative index 4,58% or 0,91%

Dynamical systems analysis of spike-adding mechanisms in transient bursts

Transient bursting behaviour of excitable cells, such as neurons, is a common feature observed experimentally, but theoretically, it is not well understood. We analyse a five-dimensional simplified model of after-depolarisation that exhibits transient bursting behaviour when perturbed with a short current injection. Using one-parameter continuation of the perturbed orbit segment formulated as a well-posed boundary value problem, we show that the spike-adding mechanism is a canard-like transition that has a different character from known mechanisms for periodic burst solutions. The biophysical basis of the model gives a natural time-scale separation, which allows us to explain the spike-adding mechanism using geometric singular perturbation theory, but it does not involve actual bifurcations as for periodic bursts. We show that unstable sheets of the critical manifold, formed by saddle equilibria of the system that only exist in a singular limit, are responsible for the spike-adding transition; the transition is organised by the slow flow on the critical manifold near folds of this manifold. Our analysis shows that the orbit segment during the spike-adding transition includes a fast transition between two unstable sheets of the slow manifold that are of saddle type. We also discuss a different parameter regime where the presence of additional saddle equilibria of the full system alters the spike-adding mechanism

Mathematical modeling of gonadotropin-releasing hormone signaling.

This is the author accepted manuscript. The final version is available from the publisher via the DOI in this record.Gonadotropin-releasing hormone (GnRH) acts via G-protein coupled receptors on pituitary gonadotropes to control of reproduction. These are Gq-coupled receptors that mediate acute effects of GnRH on the exocytotic secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH), as well as the chronic regulation of their synthesis. GnRH is secreted in short pulses and GnRH effects on its target cells are dependent upon the dynamics of these pulses. Here we overview GnRH receptors and their signaling network, placing emphasis on pulsatile signaling, and how mechanistic mathematical models and an information theoretic approach have helped further this field.This work was funded Project Grants from MRC (93447) and the BBSRC (J014699). KTA and MV gratefully acknowledge the financial support of the EPSRC via grant EP/N014391/1 and an MRC Biomedical Informatics Fellowship (MR/K021826/1), respectively

Control of clustered action potential firing in a mathematical model of entorhinal cortex stellate cells.

This is the author accepted manuscript. The final version is available from Elsevier via the DOI in this record.The entorhinal cortex is a crucial component of our memory and spatial navigation systems and is one of the first areas to be affected in dementias featuring tau pathology, such as Alzheimer's disease and frontotemporal dementia. Electrophysiological recordings from principle cells of medial entorhinal cortex (layer II stellate cells, mEC-SCs) demonstrate a number of key identifying properties including subthreshold oscillations in the theta (4-12 Hz) range and clustered action potential firing. These single cell properties are correlated with network activity such as grid firing and coupling between theta and gamma rhythms, suggesting they are important for spatial memory. As such, experimental models of dementia have revealed disruption of organised dorsoventral gradients in clustered action potential firing. To better understand the mechanisms underpinning these different dynamics, we study a conductance based model of mEC-SCs. We demonstrate that the model, driven by extrinsic noise, can capture quantitative differences in clustered action potential firing patterns recorded from experimental models of tau pathology and healthy animals. The differential equation formulation of our model allows us to perform numerical bifurcation analyses in order to uncover the dynamic mechanisms underlying these patterns. We show that clustered dynamics can be understood as subcritical Hopf/homoclinic bursting in a fast-slow system where the slow sub-system is governed by activation of the persistent sodium current and inactivation of the slow A-type potassium current. In the full system, we demonstrate that clustered firing arises via flip bifurcations as conductance parameters are varied. Our model analyses confirm the experimentally suggested hypothesis that the breakdown of clustered dynamics in disease occurs via increases in AHP conductance.The contribution of MG, KTR and JB was generously supported by a Wellcome Trust Institutional Strategic Support Award (WT105618MA). MG and KT gratefully acknowledge the financial support of the EPSRC via grant EP/N014391/1. LT’s doctoral studentship is supported by the Alzheimer’s Society in partnership with the Garfield Weston Foundation (grant reference 231). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Gonadotropin-releasing hormone signaling: An information theoretic approach

This is the author accepted manuscript. The final version is available from Elsevier via the DOI in this record.Gonadotropin-releasing hormone (GnRH) is a peptide hormone that mediates central control of reproduction, acting via G-protein coupled receptors that are primarily Gq coupled and mediate GnRH effects on the synthesis and secretion of luteinizing hormone and follicle-stimulating hormone. A great deal is known about the GnRH receptor signaling network but GnRH is secreted in short pulses and much less is known about how gonadotropes decode this pulsatile signal. Similarly, single cell measures reveal considerable cell-cell heterogeneity in responses to GnRH but the impact of this variability on signaling is largely unknown. Ordinary differential equation-based mathematical models have been used to explore the decoding of pulse dynamics and information theory-derived statistical measures are increasingly used to address the influence of cell-cell variability on the amount of information transferred by signaling pathways. Here, we describe both approaches for GnRH signaling, with emphasis on novel insights gained from the information theoretic approach and on the fundamental question of why GnRH is secreted in pulses.This work was funded Project Grants from MRC (93447) and the BBSRC (J014699). KTA and MV gratefully acknowledge the financial support of the EPSRC via grant EP/N014391/1 and an MRC Biomedical Informatics Fellowship (MR/K021826/1), respectively

Design and validation of a virtual player for studying interpersonal coordination in the mirror game

This is the author accepted manuscript. The final version is available from the publisher via the DOI in this record.The mirror game has been recently proposed as a simple, yet powerful paradigm for studying interpersonal interactions. It has been suggested that a virtual partner able to play the game with human subjects can be an effective tool to affect the underlying neural processes needed to establish the necessary connections between the players, and also to provide new clinical interventions for rehabilitation of patients suffering from social disorders. Inspired by the motor processes of the central nervous system (CNS) and the musculoskeletal system in the human body, in this paper we develop a novel interactive cognitive architecture based on nonlinear control theory to drive a virtual player (VP) to play the mirror game with a human player (HP) in different configurations. Specifically, we consider two cases: the former where the VP acts as leader and the latter where it acts as follower. The crucial problem is to design a feedback control architecture capable of imitating and following or leading a human player in a joint action task. Movement of the end-effector of the VP is modeled by means of a feedback controlled Haken-Kelso-Bunz (HKB) oscillator, which is coupled with the observed motion of the HP measured in real time. To this aim, two types of control algorithms (adaptive control and optimal control) are used and implemented on the HKB model so that the VP can generate a human-like motion while satisfying certain kinematic constraints. A proof of convergence of the control algorithms is presented in the paper together with an extensive numerical and experimental validation of their effectiveness. A comparison with other existing designs is also discussed, showing the flexibility and the advantages of our control-based approach.This work was funded by the European Project AlterEgo FP7 ICT 2.9 - Cognitive Sciences and Robotics, Grant Number 600610

Artificial intelligence, bias and clinical safety

This is the final version. Available on open access from BMJ Publishing group via the DOI in this recordEngineering and Physical Sciences Research Council (EPSRC

Critical currents in vicinal YBa2_2Cu3_3O7−δ_{7-\delta} films

Most measurements of critical current densities in YBa$_2$Cu$_3$O$_{7-\delta}$ thin films to date have been performed on films where the \textit{c}-axis is grown normal to the film surface. With such films, the analysis of the dependence of $j_c$ on the magnetic field angle is complex. The effects of extrinsic contributions to the angular field dependence of $j_c$, such as the measurement geometry and disposition of pinning centres, are convoluted with those intrinsically due to the anisotropy of the material. As a consequence of this, it is difficult to distinguish between proposed FLL structure models on the basis of angular critical current density measurements on \textit{c}-axis films. Films grown on mis-cut (vicinal) substrates have a reduced measurement symmetry and thus provide a greater insight into the critical current anisotropy. In this paper previous descriptions of the magnetic field angle dependence of $j_c$ in YBa$_2$Cu$_3$O$_{7-\delta}$ are reviewed. Measurements on YBa$_2$Cu$_3$O$_{7-\delta}$ thin films grown on a range of vicinal substrates are presented and the results interpreted in terms of the structure and dimensionality of the FLL in YBa$_2$Cu$_3$O$_{7-\delta}$. There is strong evidence for a transition in the structure of the flux line lattice depending on magnetic field magnitude, orientation and temperature. As a consequence, a simple scaling law can not, by itself, describe the observed critical current anisotropy in YBa$_2$Cu$_3$O$_{7-\delta}$. The experimentally obtained $j_c(\theta)$ behaviour of YBCO is successfully described in terms of a kinked vortex structure for fields applied near parallel to the \textit{a-b} planes.Comment: 10 pages, 12 figures, Submitted to PR