The role of clathrin in post-golgi trafficking in toxoplasma gondii

Apicomplexan parasites are single eukaryotic cells with a highly polarised secretory system that contains unique secretory organelles (micronemes and rhoptries) that are required for host cell invasion. In contrast, the role of the endosomal system is poorly understood in these parasites. With many typical endocytic factors missing, we speculated that endocytosis depends exclusively on a clathrin-mediated mechanism. Intriguingly, in Toxoplasma gondii we were only able to observe the endogenous clathrin heavy chain 1 (CHC1) at the Golgi, but not at the parasite surface. For the functional characterisation of Toxoplasma gondii CHC1 we generated parasite mutants conditionally expressing the dominant negative clathrin Hub fragment and demonstrate that CHC1 is essential for vesicle formation at the trans-Golgi network. Consequently, the functional ablation of CHC1 results in Golgi aberrations, a block in the biogenesis of the unique secretory microneme and rhoptry organelles, and of the pellicle. However, we found no morphological evidence for clathrin mediating endocytosis in these parasites and speculate that they remodelled their vesicular trafficking system to adapt to an intracellular lifestyle

Retinoic acid elicits a coordinated expression of gut homing markers on T lymphocytes of Zambian men receiving oral Vivotif, but not Rotarix, Dukoral or OPVERO vaccines

This work was supported by the Bill and Melinda Gates Foundation through a Grand Challenges Explorations grant (OPP1043696)

Structural Disorder Provides Increased Adaptability for Vesicle Trafficking Pathways

Vesicle trafficking systems play essential roles in the communication between the organelles of eukaryotic cells and also between cells and their environment. Endocytosis and the late secretory route are mediated by clathrin-coated vesicles, while the COat Protein I and II (COPI and COPII) routes stand for the bidirectional traffic between the ER and the Golgi apparatus. Despite similar fundamental organizations, the molecular machinery, functions, and evolutionary characteristics of the three systems are very different. In this work, we compiled the basic functional protein groups of the three main routes for human and yeast and analyzed them from the structural disorder perspective. We found similar overall disorder content in yeast and human proteins, confirming the well-conserved nature of these systems. Most functional groups contain highly disordered proteins, supporting the general importance of structural disorder in these routes, although some of them seem to heavily rely on disorder, while others do not. Interestingly, the clathrin system is significantly more disordered (,23%) than the other two, COPI (,9%) and COPII (,8%). We show that this structural phenomenon enhances the inherent plasticity and increased evolutionary adaptability of the clathrin system, which distinguishes it from the other two routes. Since multi-functionality (moonlighting) is indicative of both plasticity and adaptability, we studied its prevalence in vesicle trafficking proteins and correlated it with structural disorder. Clathrin adaptors have the highest capability for moonlighting while also comprising the most highly disordered members. The ability to acquire tissue specific functions was also used to approach adaptability: clathrin route genes have the most tissue specific exons encoding for protein segments enriched in structural disorder and interaction sites. Overall, our results confirm the general importance of structural disorder in vesicle trafficking and suggest major roles for this structural property in shaping the differences of evolutionary adaptability in the three routes

A Didactic Model of Macromolecular Crowding Effects on Protein Folding

A didactic model is presented to illustrate how the effect of macromolecular crowding on protein folding and association is modeled using current analytical theory and discrete molecular dynamics. While analytical treatments of crowding may consider the effect as a potential of average force acting to compress a polypeptide chain into a compact state, the use of simulations enables the presence of crowding reagents to be treated explicitly. Using an analytically solvable toy model for protein folding, an approximate statistical thermodynamic method is directly compared to simulation in order to gauge the effectiveness of current analytical crowding descriptions. Both methodologies are in quantitative agreement under most conditions, indication that both current theory and simulation methods are capable of recapitulating aspects of protein folding even by utilizing a simplistic protein model

Cognitive effects of transcranial direct current stimulation combined with working memory training in fibromyalgia: a randomized clinical trial

Cognitive dysfunction in fibromyalgia has been reported, especially memory. Anodal transcranial direct current stimulation (tDCS) over the dorsolateral prefrontal cortex (DLPFC) has been effective in enhancing this function. We tested the effects of eight sessions of tDCS and cognitive training on immediate and delayed memory, verbal fluency and working memory and its association with brain-derived neurotrophic factor (BDNF) levels. Forty females with fibromyalgia were randomized to receive eight sessions of active or sham tDCS. Anodal stimulation (2 mA) was applied over the DLPFC and online combined with a working memory training (WMT) for 20 minutes. Pre and post-treatment neurocognitive tests were administered. Data analysis on deltas considering years of education and BDNF as covariates, indicated active-tDCS + WMT significantly increased immediate memory indexed by Rey Auditory Verbal Learning Test score when compared to sham. This effect was dependent on basal BDNF levels. In addition, the model showed active stimulation increased orthographic and semantic verbal fluency scores (Controlled Oral Word Association Test) and short-term memory (Forward Digit Span). The combination of both techniques seemed to produce effects on specific cognitive functions related to short-term and long-term episodic memory and executive functions, which has clinical relevance for top-down treatment approaches in FM.financiamento: This research was supported by grants and material support from the following Brazilian agencies: Committee for the Development of Higher Education Personnel - CAPES - PNPD/CAPES and material support. National Council for Scientific and Technological Development - CNPq (grants to Dr. I.L.S. Torres, Dr. W. Caumo). Postgraduate Program in Medical Sciences at the School of Medicine of the Federal University of Rio Grande do Sul (material support). Postgraduate Research Group at the Hospital de Clinicas de Porto Alegre - FIPE HCPA (material support). Foundations for Support of Research at Rio Grande do Sul (FAPERGS) (material support)

Curvature of Double-Membrane Organelles Generated by Changes in Membrane Size and Composition

Transient double-membrane organelles are key players in cellular processes such as autophagy, reproduction, and viral infection. These organelles are formed by the bending and closure of flat, double-membrane sheets. Proteins are believed to be important in these morphological transitions but the underlying mechanism of curvature generation is poorly understood. Here, we describe a novel mechanism for this curvature generation which depends primarily on three membrane properties: the lateral size of the double-membrane sheets, the molecular composition of their highly curved rims, and a possible asymmetry between the two flat faces of the sheets. This mechanism is evolutionary advantageous since it does not require active processes and is readily available even when resources within the cell are restricted as during starvation, which can induce autophagy and sporulation. We identify pathways for protein-assisted regulation of curvature generation, organelle size, direction of bending, and morphology. Our theory also provides a mechanism for the stabilization of large double-membrane sheet-like structures found in the endoplasmic reticulum and in the Golgi cisternae

The Effect of Macromolecular Crowding, Ionic Strength and Calcium Binding on Calmodulin Dynamics

The flexibility in the structure of calmodulin (CaM) allows its binding to over 300 target proteins in the cell. To investigate the structure-function relationship of CaM, we combined methods of computer simulation and experiments based on circular dichroism (CD) to investigate the structural characteristics of CaM that influence its target recognition in crowded cell-like conditions. We developed a unique multiscale solution of charges computed from quantum chemistry, together with protein reconstruction, coarse-grained molecular simulations, and statistical physics, to represent the charge distribution in the transition from apoCaM to holoCaM upon calcium binding. Computationally, we found that increased levels of macromolecular crowding, in addition to calcium binding and ionic strength typical of that found inside cells, can impact the conformation, helicity and the EF hand orientation of CaM. Because EF hand orientation impacts the affinity of calcium binding and the specificity of CaM's target selection, our results may provide unique insight into understanding the promiscuous behavior of calmodulin in target selection inside cells.Comment: Accepted to PLoS Comp Biol, 201

Peer Support Workers in Health:A Qualitative Metasynthesis of Their Experiences

Peer support models, where an individual has a specific illness or lifestyle experience and supports others experiencing similar challenges, have frequently been used in different fields of healthcare to successfully engage hard-to-reach groups. Despite recognition of their value, the impact of these roles on the peer has not been systematically assessed. By synthesising the qualitative literature we sought to review such an impact, providing a foundation for designing future clinical peer models.Systematic review and qualitative metasynthesis of studies found in Medline, CINAHL or Scopus documenting peer worker experiences.1,528 papers were found, with 34 meeting the criteria of this study. Findings were synthesised to reveal core constructs of reframing identity through reciprocal relations and the therapeutic use of self, enhancing responsibility.The ability of the Peer Support Worker to actively engage with other marginalised or excluded individuals based on their unique insight into their own experience supports a therapeutic model of care based on appropriately sharing their story. Our findings have key implications for maximising the effectiveness of Peer Support Workers and in contributing their perspective to the development of a therapeutic model of care

Management of latent Mycobacterium tuberculosis infection: WHO guidelines for low tuberculosis burden countries

Latent tuberculosis infection (LTBI) is characterised by the presence of immune responses to previously acquired Mycobacterium tuberculosis infection without clinical evidence of active tuberculosis (TB). Here we report evidence-based guidelines from the World Health Organization for a public health approach to the management of LTBI in high risk individuals in countries with high or middle upper income and TB incidence of <100 per 100 000 per year. The guidelines strongly recommend systematic testing and treatment of LTBI in people living with HIV, adult and child contacts of pulmonary TB cases, patients initiating anti-tumour necrosis factor treatment, patients receiving dialysis, patients preparing for organ or haematological transplantation, and patients with silicosis. In prisoners, healthcare workers, immigrants from high TB burden countries, homeless persons and illicit drug users, systematic testing and treatment of LTBI is conditionally recommended, according to TB epidemiology and resource availability. Either commercial interferon-gamma release assays or Mantoux tuberculin skin testing could be used to test for LTBI. Chest radiography should be performed before LTBI treatment to rule out active TB disease. Recommended treatment regimens for LTBI include: 6 or 9 month isoniazid; 12 week rifapentine plus isoniazid; 3–4 month isoniazid plus rifampicin; or 3–4 month rifampicin alone