Experimental and theoretical investigation of mechanism of Kinesin motility

Thesis (S.B.)--Massachusetts Institute of Technology, Dept. of Physics, 2007.Includes bibliographical references (p. 42-53).Kinesin is a motor protein capable of utilizing chemical energy from ATP hydrolysis to generate mechanical force to power its progressive motility along a microtubule track. The mechanism of motility has been a subject of extensive study for last decade. Recently, it has been proposed that novel element-cover strand-is essential in power-stroke-like force generation. In this work we attempt an experimental verification of this hypothesis by studying the mechanical properties, such as unloaded velocity, force velocity relationship, stall forces, processivity and step size of kinesin and mutants targeting cover strand region. We show that A9G and D11G mutants move slower and have lower stall force then the wild type molecule, but the mutants are ultraprocessive, make steps of 7nm and have a higher probability of taking backward steps suggesting that, indeed, force generating mechanism might been adversely affected by this mutation but it could also affect flexibility and directionality of the molecule.by Anna Kinga Labno.S.B

La protección del equilibrio presupuestario en la Constitución de Polonia

En este artículo se estudia el tope de gastos previsto en la Constitución polaca, los debates constituyentes, las problemáticas de su aplicación y la complejidad de determinar ulteriores violaciones a la limitación constitucional. De igual modo, se estudian algunos pronunciamientos del Tribunal Constitucional polaco. Se argumenta que dicha prevención constitucional juega un papel preventivo importante.This article deals with the expenditure ceiling as established in Polish constitution, the constituent debates, the problems of its application and the complexity o determining the violation of constitutional limits. This paper analyzes as well the rulings of Poland’s Constitutional Tribunal. It argues that the constitutional limitation plays an important preventing role

Media from macrophages co-incubated with <i>Enterococcus faecalis</i> induces epithelial cell monolayer reassembly and altered cell morphology

Signal exchange between intestinal epithelial cells, microbes and local immune cells is an important mechanism of intestinal homeostasis. Given that intestinal macrophages are in close proximity to both the intestinal epithelium and the microbiota, their pathologic interactions may result in epithelial damage. The present study demonstrates that co-incubation of murine macrophages with E. faecalis strains producing gelatinase (GelE) and serine protease (SprE) leads to resultant condition media (CM) capable of inducing reassembly of primary colonic epithelial cell monolayers. Following the conditioned media (CM) exposure, some epithelial cells are shed whereas adherent cells are observed to undergo dissolution of cell-cell junctions and morphologic transformation with actin cytoskeleton reorganization resulting in flattened and elongated shapes. These cells exhibit marked filamentous filopodia and lamellipodia formation. Cellular reorganization is not observed when epithelial monolayers are exposed to: CM from macrophages co-incubated with E. faecalis GelE/SprE-deficient mutants, CM from macrophages alone, or E. faecalis (GelE/SprE) alone. Flow cytometry analysis reveals increased expression of CD24 and CD44 in cells treated with macrophage/E. faecalis CM. This finding in combination with the appearance colony formation in matrigel demonstrate that the cells treated with macrophage/E. faecalis CM contain a higher proportion progenitor cells compared to untreated control. Taken together, these findings provide evidence for a triangulated molecular dialogue between E. faecalis, macrophages and colonic epithelial cells, which may have important implications for conditions in the gut that involve inflammation, injury or tumorigenesis

Children and young people's mental health outcome measures in paediatrics

Paediatricians are increasingly likely to encounter children and young people with mental health difficulties, either as primary presentations or as comorbidities linked with chronic illnesses. However, paediatricians may have limited training or experience regarding the tools available to identify mental health needs and how to use these tools. The current paper aims to provide a go-to guide for paediatricians when considering the use of mental health and well-being outcome measures, including how to select, administer and interpret measures effectively. It also provides practical guidance on the most common mental health outcome measures used in children and young people's mental health services across the UK and elsewhere, which paediatricians are likely to encounter in their practice. Paediatricians may also find these measures useful in their own practice to screen for potential mental health difficulties, monitor the impact of chronic health conditions on a young person's mental health and well-being, or to provide evidence when referring young people to mental health services

Gender, marginalised groups, and young people’s mental health: a longitudinal analysis of trajectories

Background Individuals from marginalised groups experience higher levels of mental health difficulties and lower levels of wellbeing which may be due to the exposure to stress and adversity. This study explores trajectories of mental health over time for young women and girls and young people with other marginalised identities.Methods We conducted a secondary analysis on N=14,215 children and young people (7,501 or 52.8% female, 6,571 or 46.2% male, and 81 or 0.6% non-binary or questioning) who completed a survey at age 11 to 12 years and at least one other annual survey aged 12 to 13 years and/or aged 13 to 14 years. We used group-based trajectory models to examine mental health difficulties.Results Except for behavioural difficulties, young women’s and girls’ trajectories showed that they consistently had higher levels of mental health difficulties compared to young men and boys. A similar pattern was shown for nonbinary and questioning children and young people. Children and young people with economic disadvantage and/or special education needs, and/or for whom there were welfare concerns, were generally more likely to experience higher levels of mental health difficulties.Conclusions This information could inform public policy, guidance and interventions

Cambridge Psycholinguistic Inventory of Christian Beliefs: A registered report of construct validity, internal consistency and test-retest reliability.

While religious beliefs are typically studied using questionnaires, there are no standardized tools available for cognitive psychology and neuroscience studies of religious cognition. Here we present the first such tool-the Cambridge Psycholinguistic Inventory of Christian Beliefs (CPICB)-which consists of audio-recorded items of religious beliefs as well as items of three control conditions: moral beliefs, abstract scientific knowledge and empirical everyday life knowledge. The CPICB is designed in such a way that the ultimate meaning of each sentence is revealed only by its final critical word, which enables the precise measurement of reaction times and/or latencies of neurophysiological responses. Each statement comes in a pair of Agree/Disagree versions of critical words, which allows for experimental contrasting between belief and disbelief conditions. Psycholinguistic and psychoacoustic matching between Agree/Disagree versions of sentences, as well as across different categories of the CPICB items (Religious, Moral, Scientific, Everyday), enables rigorous control of low-level psycholinguistic and psychoacoustic features while testing higher-level beliefs. In the exploratory Study 1 (N = 20), we developed and tested a preliminary version of the CPICB that had 480 items. After selecting 400 items that yielded the most consistent responses, we carried out a confirmatory test-retest Study 2 (N = 40). Preregistered data analyses confirmed excellent construct validity, internal consistency and test-retest reliability of the CPICB religious belief statements. We conclude that the CPICB is suitable for studying Christian beliefs in an experimental setting involving behavioural and neuroimaging paradigms, and provide Open Access to the inventory items, fostering further development of the experimental research of religiosity

Regulated Capture of Vκ Gene Topologically Associating Domains by Transcription Factories

Expression of vast repertoires of antigen receptors by lymphocytes, with each cell expressing a single receptor, requires stochastic activation of individual variable (V) genes for transcription and recombination. How this occurs remains unknown. Using single-cell RNA sequencing (scRNA-seq) and allelic variation, we show that individual pre-B cells monoallelically transcribe divergent arrays of Vκ genes, thereby opening stochastic repertoires for subsequent Vκ-Jκ recombination. Transcription occurs upon translocation of Vκ genes to RNA polymerase II arrayed on the nuclear matrix in transcription factories. Transcription is anchored by CTCF-bound sites or E2A-loaded Vκ promotors and continues over large genomic distances delimited only by topological associating domains (TADs). Prior to their monoallelic activation, Vκ loci are transcriptionally repressed by cyclin D3, which prevents capture of Vκ gene containing TADs by transcription factories. Cyclin D3 also represses protocadherin, olfactory, and other monoallelically expressed genes, suggesting a widely deployed mechanism for coupling monoallelic gene activation with cell cycle exit

Altered Chromosomal Positioning, Compaction, and Gene Expression with a Lamin A/C Gene Mutation

Lamins A and C, encoded by the LMNA gene, are filamentous proteins that form the core scaffold of the nuclear lamina. Dominant LMNA gene mutations cause multiple human diseases including cardiac and skeletal myopathies. The nuclear lamina is thought to regulate gene expression by its direct interaction with chromatin. LMNA gene mutations may mediate disease by disrupting normal gene expression.To investigate the hypothesis that mutant lamin A/C changes the lamina's ability to interact with chromatin, we studied gene misexpression resulting from the cardiomyopathic LMNA E161K mutation and correlated this with changes in chromosome positioning. We identified clusters of misexpressed genes and examined the nuclear positioning of two such genomic clusters, each harboring genes relevant to striated muscle disease including LMO7 and MBNL2. Both gene clusters were found to be more centrally positioned in LMNA-mutant nuclei. Additionally, these loci were less compacted. In LMNA mutant heart and fibroblasts, we found that chromosome 13 had a disproportionately high fraction of misexpressed genes. Using three-dimensional fluorescence in situ hybridization we found that the entire territory of chromosome 13 was displaced towards the center of the nucleus in LMNA mutant fibroblasts. Additional cardiomyopathic LMNA gene mutations were also shown to have abnormal positioning of chromosome 13, although in the opposite direction.These data support a model in which LMNA mutations perturb the intranuclear positioning and compaction of chromosomal domains and provide a mechanism by which gene expression may be altered