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Key Points The carboxy-terminal tails of α- and β-tubulin are essential for microtubule function. They lie on the outer surface of the microtubule where they can influence the binding of associated proteins. With the exception of acetylation, the post-translational modifications of microtubules — that is, detyrosination/tyrosination, formation of Δ2-tubulin, polyglutamylation and polyglycylation — are all located in the carboxy-terminal tails. Acetylation of α-tubulin can be abolished without consequences in Tetrahymena , but it seems to have a function in cell motility. Two histone deacetylases, HDAC6 and SIRT2, have been shown to function as tubulin deacetylases. Genetic analysis of polyglycylation in Tetrahymena demonstrates its essential function in the organization of axonemes, cell motility and cytokinesis. Polyglutamylation can influence the binding of structural and motor microtubule-associated proteins (MAPs) to microtubules. Antibody-injection studies indicate an important role for polyglutamylation in centriole stability. The functional role of the tyrosination cycle of tubulin is still unclear; cells cultured with low activity of the tubulin tyrosine ligase (TTL) enzyme show no obvious defects. TTL-knockout mice, however, die early in development owing to an as-yet-uncharacterized defect. The αβ-tubulin heterodimer, the building block of microtubules, is subject to a large number of post-translational modifications, comparable in diversity to the intensively studied histone modifications. Although these unusual modifications are conserved throughout evolution, their functions have remained almost completely elusive. Recently, however, important advances in the understanding of how tubulin modifications regulate function and organization have been made.

Analyses of protein complexes are facilitated by methods that enable the generation of recombinant complexes via coexpression of their subunits from multigene DNA constructs. However, low experimental throughput limits the generation of such constructs in parallel. Here we describe a method that allows up to 25 cDNAs to be assembled into a single baculoviral expression vector in only two steps. This method, called biGBac, uses computationally optimized DNA linker sequences that enable the efficient assembly of linear DNA fragments, using reactions developed by Gibson for the generation of synthetic genomes. The biGBac method uses a flexible and modular “mix and match” approach and enables the generation of baculoviruses from DNA constructs at any assembly stage. Importantly, it is simple, efficient, and fast enough to allow the manual generation of many multigene expression constructs in parallel. We have used this method to generate and characterize recombinant forms of the anaphase-promoting complex/cyclosome, cohesin, and kinetochore complexes.

The Ndc80 complex is the key microtubule‐binding element of the kinetochore. In contrast to the well‐characterized interaction of Ndc80‐Nuf2 heads with microtubules, little is known about how the Spc24‐25 heterodimer connects to centromeric chromatin. Here, we present molecular details of Spc24‐25 in complex with the histone‐fold protein Cnn1/CENP‐T illustrating how this connection ultimately links microtubules to chromosomes. The conserved Ndc80 receptor motif of Cnn1 is bound as an α helix in a hydrophobic cleft at the interface between Spc24 and Spc25. Point mutations that disrupt the Ndc80–Cnn1 interaction also abrogate binding to the Mtw1 complex and are lethal in yeast. We identify a Cnn1‐related motif in the Dsn1 subunit of the Mtw1 complex, necessary for Ndc80 binding and essential for yeast growth. Replacing this region with the Cnn1 peptide restores viability demonstrating functionality of the Ndc80‐binding module in different molecular contexts. Finally, phosphorylation of the Cnn1 N‐terminus coordinates the binding of the two competing Ndc80 interaction partners. Together, our data provide structural insights into the modular binding mechanism of the Ndc80 complex to its centromere recruiters. The yeast kinetochore receptors Cnn1 and Dsn1 employ a related peptide motif to recruit the microtubule‐binding Ndc80 complex in a competitive, phosphorylation‐regulated manner.

The majority of individuals with problematic and pathological gambling remain untreated, and treatment barriers are high. Internet-based interventions can help to address existing barriers, and first studies suggest their potential for this target group. Within a randomized controlled trial ( N  = 150) with two assessment times (baseline and post-intervention), we aimed to investigate the feasibility, acceptance, and effectiveness of a self-guided Internet-based intervention targeted at gambling problems. We expected a significant reduction in gambling symptoms (primary outcome) and depressive symptoms as well gambling-specific dysfunctional thoughts (secondary outcomes) in the intervention group (IG) compared to a wait-list control group with access to treatment-as-usual (control group, CG) after the intervention period of 8 weeks. Results of the complete cases, per protocol, intention-to-treat (ITT), and frequent user analyses showed significant improvements in both groups for primary and secondary outcomes but no significant between-group differences (ITT primary outcome, F (1,147) = .11, p  = .739, ηp2 < .001). Moderation analyses indicated that individuals in the IG with higher gambling and depressive symptoms, older age, and comorbid anxiety symptoms showed significant improvement relative to the CG. The intervention was positively evaluated (e.g., 96.5% rated the program as useful). Possible reasons for the nonsignificant between-group differences are discussed. Future studies should include follow-up assessments and larger samples to address limitations of the present study. Trial Registration ClinicalTrials.gov (NCT03372226), http://clinicaltrials.gov/ct2/show/NCT03372226 , date of registration (13/12/2017).

Schizophrenia is a complex psychiatric disorder with unknown and presumably heterogeneous etiology. While the disorder can have various outcomes, research is predominantly “deficit-oriented” emphasizing the hardship that the disorder inflicts on sufferers as well as their families and society. Beyond symptom reduction, imparting patients with hope and meaning in life is increasingly considered an important treatment target, which may raise self-esteem, and reduce self-stigma and suicidal ideation. The present study compared a psychotherapeutic treatment aimed at improving cognitive insight, individualized metacognitive intervention (MCT+), with an active control in order to elucidate if personal meaning-making and hope can be improved in patients with psychosis across time. A total of 92 patients were randomized to either individualized metacognitive therapy (MCT+) or CogPack (neuropsychological training) and followed up for up to 6 months. The “Subjective Sense in Psychosis Questionnaire” (SUSE) was administered which covers different salutogenetic vs pathogenetic views of the disorder, valence of symptom experiences and the consequences of psychosis. Patients in the MCT+ group showed a significant positive shift in attitudes towards the consequences of their illness over time relative to patients in the active control condition. There was some evidence that MCT+ also enhanced meaning-making. The perceived negative consequences of psychosis were highly correlated with depression and low self-esteem, as well as suicidality. The study shows that a cognitive insight training can improve meaning-making in patients and help them come to terms with their diagnosis.

Healthy sleep, positive general affect, and the ability to regulate emotional experiences are fundamental for well-being. In contrast, various mental disorders are associated with altered rapid eye movement (REM) sleep, negative affect, and diminished emotion regulation abilities. However, the neural processes mediating the relationship between these different phenomena are still not fully understood. In the present study of 42 healthy volunteers, we investigated the effects of selective REM sleep suppression (REMS) on general affect, as well as on feelings of social exclusion, cognitive reappraisal (CRA) of emotions, and their neural underpinnings. Using functional magnetic resonance imaging we show that, on the morning following sleep suppression, REMS increases general negative affect, enhances amygdala responses and alters its functional connectivity with anterior cingulate cortex during passively experienced experimental social exclusion. However, we did not find effects of REMS on subjective emotional ratings in response to social exclusion, their regulation using CRA, nor on functional amygdala connectivity while participants employed CRA. Our study supports the notion that REM sleep is important for affective processes, but emphasizes the need for future research to systematically investigate how REMS impacts different domains of affective experience and their neural correlates, in both healthy and (sub-)clinical populations.

Background Evidence shows that internet-based self-help interventions are effective in reducing symptoms for a wide range of mental disorders. To date, online interventions treating psychotic disorders have been scarce, even though psychosis is among the most burdensome disorders worldwide. Furthermore, the implementation of cognitive-behavioral therapy (CBT) for psychosis in routine health care is challenging. Internet-based interventions could narrow this treatment gap. Thus, a comprehensive CBT-based online self-help intervention for people with psychosis has been developed. The aim of this study is the evaluation of the feasibility and efficacy of the intervention compared with a waiting list control group. Methods The intervention includes modules on delusion, voice hearing, social competence, mindfulness, and seven other domains. Participants are guided through the program by a personal moderator. Usage can be amended by an optional smartphone app. In this randomized controlled trial, participants are allocated to a waiting list or an intervention of eight weeks. Change in positive psychotic symptoms of both groups will be compared (primary outcome) and predictors of treatment effects will be assessed. Discussion To our knowledge, this project is one of the first large-scale investigations of an internet-based intervention for people with psychosis. It may thus be a further step to broaden treatment options for people suffering from this disorder. Trial registration NCT02974400 (clinicaltrials.gov), date of registration: November 28th 2016.

The development of brain imaging techniques, in particular functional magnetic resonance imaging (fMRI), made it possible to non-invasively study the hemispheric lateralization of cognitive brain functions in large cohorts. Comprehensive models of hemispheric lateralization are, however, still missing and should not only account for the hemispheric specialization of individual brain functions, but also for the interactions among different lateralized cognitive processes (e.g., language and visuospatial processing). This calls for robust and reliable paradigms to study hemispheric lateralization for various cognitive functions. While numerous reliable imaging paradigms have been developed for language, which represents the most prominent left-lateralized brain function, the reliability of imaging paradigms investigating typically right-lateralized brain functions, such as visuospatial processing, has received comparatively less attention. In the present study, we aimed to establish an fMRI paradigm that robustly and reliably identifies right-hemispheric activation evoked by visuospatial processing in individual subjects. In a first study, we therefore compared three frequently used paradigms for assessing visuospatial processing and evaluated their utility to robustly detect right-lateralized brain activity on a single-subject level. In a second study, we then assessed the test-retest reliability of the so-called Landmark task-the paradigm that yielded the most robust results in study 1. At the single-voxel level, we found poor reliability of the brain activation underlying visuospatial attention. This suggests that poor signal-to-noise ratios can become a limiting factor for test-retest reliability. This represents a common detriment of fMRI paradigms investigating visuospatial attention in general and therefore highlights the need for careful considerations of both the possibilities and limitations of the respective fMRI paradigm-in particular, when being interested in effects at the single-voxel level. Notably, however, when focusing on the reliability of measures of hemispheric lateralization (which was the main goal of study 2), we show that hemispheric dominance (quantified by the lateralization index, LI, with |LI| >0.4) of the evoked activation could be robustly determined in more than 62% and, if considering only two categories (i.e., left, right), in more than 93% of our subjects. Furthermore, the reliability of the lateralization strength (LI) was \"fair\" to \"good\". In conclusion, our results suggest that the degree of right-hemispheric dominance during visuospatial processing can be reliably determined using the Landmark task, both at the group and single-subject level, while at the same time stressing the need for future refinements of experimental paradigms and more sophisticated fMRI data acquisition techniques.

Hang on in there A long-standing mystery of mitosis research is the mechanism that transports chromosomes to the nuclear-spindle poles during anaphase. The chromosomes seem to gain their objective by clinging on to the kinetochore microtubule polymers despite the fact that they are disassembling at the time. Fluorescence microscopy has now been used to produce movies and electron micrographs that show how it's done. The microtubule polymer disassembles via a conformational change that pushes the Dam1 ring complex, an important microtubule binding element in the budding yeast kinetochore, along its lattice. This elegant mechanism may be the key to the conversion of force generated by microtubule depolymerization into the chromosome movements seen in mitosis. The Dam1 ring complex is a molecular device that can translate the force generated by microtubule depolymerization into movement along the lattice to facilitate chromosome segregation. Chromosomes interact through their kinetochores with microtubule plus ends and they are segregated to the spindle poles as the kinetochore microtubules shorten during anaphase A of mitosis. The molecular natures and identities of coupling proteins that allow microtubule depolymerization to pull chromosomes to poles during anaphase have long remained elusive 1 . In budding yeast, the ten-protein Dam1 complex is a critical microtubule-binding component of the kinetochore 2 that oligomerizes into a 50-nm ring around a microtubule in vitro 3 , 4 . Here we show, with the use of a real-time, two-colour fluorescence microscopy assay, that the ring complex moves processively for several micrometres at the ends of depolymerizing microtubules without detaching from the lattice. Electron microscopic analysis of ‘end-on views’ revealed a 16-fold symmetry of the kinetochore rings. This out-of-register arrangement with respect to the 13-fold microtubule symmetry is consistent with a sliding mechanism based on an electrostatically coupled ring–microtubule interface. The Dam1 ring complex is a molecular device that can translate the force generated by microtubule depolymerization into movement along the lattice to facilitate chromosome segregation.

Background Only a small fraction of individuals with pathological or problematic gambling seek professional help despite available evidence-based treatments such as cognitive behavioral therapy (CBT). Anonymous Internet-based interventions may help to overcome treatment barriers. Results of a pilot study using an Internet-based intervention for depression in a sample of individuals with problematic or pathological gambling behavior show that both depressive and gambling-related symptomatology can be reduced with a generic depression program compared with a wait-list control group. Based on encouraging results of the pilot study, we developed a low-threshold, anonymous and cost-free online self-help program (“Restart”) to test whether a program tailored to the needs of gamblers yields better results compared to the effects of the intervention evaluated in the pilot study. The online self-help program is based on CBT, targeting emotional problems and gambling-related symptoms and is accompanied by a smartphone application to sustain treatment benefits. Method A randomized controlled trial with two conditions (intervention group and wait-list control group), two assessment times (reassessment after 8 weeks) and a total of 136 participants is planned. The primary outcome will be change in pathological gambling measured with the Pathological Gambling Adaptation of Yale-Brown Obsessive-Compulsive Scale from pre to post intervention. The change in depressive symptoms (assessed with the Patient Health Questionnaire - 9 depression module) and gambling-related dysfunctional thoughts (assessed with the Gambling Attitudes and Beliefs Survey) will represent secondary outcomes. The intervention includes modules on debt management, impulse control, gambling-specific cognitive biases, self-esteem, social competence, sleep hygiene, mindfulness and positive activities. Discussion This study is one of the first investigations of Internet-based self-help programs in a sample of problematic gamblers. Self-guided Internet-based interventions represent a promising possibility to narrow the existing treatment gap while saving expensive and scarce resources (e.g., psychotherapists). The expected findings will add substantial knowledge in the development of effective Internet-based treatments for individuals with gambling problems. The empirical and clinical implications (e.g., broader use and promotion of such interventions in the future) and the limitations of the study will be discussed. Trial registration ClinicalTrials.gov, NCT03372226 . Registered on 13 December 2017.