Dopamine regulates decision thresholds in human reinforcement learning in males.

Dopamine fundamentally contributes to reinforcement learning, but recent accounts also suggest a contribution to specific action selection mechanisms and the regulation of response vigour. Here, we examine dopaminergic mechanisms underlying human reinforcement learning and action selection via a combined pharmacological neuroimaging approach in male human volunteers (n = 31, within-subjects; Placebo, 150 mg of the dopamine precursor L-dopa, 2 mg of the D2 receptor antagonist Haloperidol). We found little credible evidence for previously reported beneficial effects of L-dopa vs. Haloperidol on learning from gains and altered neural prediction error signals, which may be partly due to differences experimental design and/or drug dosages. Reinforcement learning drift diffusion models account for learning-related changes in accuracy and response times, and reveal consistent decision threshold reductions under both drugs, in line with the idea that lower dosages of D2 receptor antagonists increase striatal DA release via an autoreceptor-mediated feedback mechanism. These results are in line with the idea that dopamine regulates decision thresholds during reinforcement learning, and may help to bridge action selection and response vigor accounts of dopamine.

Single cohesin molecules generate force by two distinct mechanisms.

Spatial organization of DNA is facilitated by cohesin protein complexes that move on DNA and extrude DNA loops. How cohesin works mechanistically as a molecular machine is poorly understood. Here, we measure mechanical forces generated by conformational changes in single cohesin molecules. We show that bending of SMC coiled coils is driven by random thermal fluctuations leading to a ~32 nm head-hinge displacement that resists forces up to 1 pN; ATPase head engagement occurs in a single step of ~10 nm and is driven by an ATP dependent head-head movement, resisting forces up to 15 pN. Our molecular dynamic simulations show that the energy of head engagement can be stored in a mechanically strained conformation of NIPBL and released during disengagement. These findings reveal how single cohesin molecules generate force by two distinct mechanisms. We present a model, which proposes how this ability may power different aspects of cohesin-DNA interaction.

Whether cycling around the city is in fact healthy in the light of air quality - Results of black carbon.

Studying the air quality and exposure of the inhabitants of urban agglomerations to pollution is the basis for the creation and development of more sustainable cities. Although research on black carbon (BC) has not yet reached the official acceptable levels and guidelines, the World Health Organization clearly indicates the need to measure and control the level of this pollutant. In Poland, monitoring of the level of BC concentration is not included in the air quality monitoring network. To estimate the extent of this pollutant to which pedestrians and cyclists are exposed, mobile measurements were carried out on over 26 km of bicycle paths in Wroclaw. The obtained results indicate the influence of urban greenery next to the bicycle path (especially if the cyclist is separated from the street lane by hedges or other tall plants) and the 'breathability' (i.e., associated with surrounding infrastructure) of the area on the obtained concentrations; the average concentration of BC in such places ranged from 1.3 to 2.2 µg/m3, whereas a cyclist riding directly on bike paths adjacent to the main roads in the city center is exposed to concentrations in the range of 2.3-14 µg/m3. The results of the measurements, also related to stationary measurements made at a selected point of one of the routes, clearly indicate the importance of the infrastructure surrounding the bicycle paths, their location, and the impact of urban traffic on the obtained BC concentrations. The results presented in our study are based only on short-term-field campaigns preliminary studies. To determine the quantitative impact of the characteristics of the bicycle route on the concentration of pollutants, and thus the exposure of users, the systematized research should cover a greater part of the city and be representative in terms of various hours of the day.

[Position of trauma surgery in emergency medicine]. / Stellung der Unfallchirurgie in der Notfallmedizin.

BACKGROUND: The decision of the Federal Joint Committee has resulted in the further development of in-hospital emergency medicine through the establishment of central emergency departments and staged emergency care. In addition, the additional training in clinical acute and emergency medicine was established. AIM: The aim of this work is on the one hand to make trauma surgeons aware of these structural changes. On the other hand, we would like to evaluate an opinion and discuss the position of trauma surgery in emergency medicine. METHODS: A web-based online survey was conducted at the Trauma Network and Trauma Registry Congress to collect participants' opinions on the position of trauma surgery in emergency medicine. RESULTS: Of 143 congress participants, 98 (67%) responded to the survey. The majority of participants were male (n = 78, 80%), over 40 years of age (n = 62, 63%), and in a professional position with staff responsibility (n = 73, 75%). Emergency medicine (mean: 84.8; SD: 18.7) and intensive care medicine (mean: 78.3; SD: 20.4) training appears important. On the other hand, subsequent work in these areas appears less important (prehospital emergency medicine: mean: 65.1; SD: 28.0; ICU: mean: 53.7, SD: 30.3); however, activity in an emergency department is rated higher (MW: 87.0; SD: 18.7). There is high agreement that the trauma leader should be a trauma surgeon (mean 87.9; SD: 19.7). DISCUSSION: A high volume of emergency trauma surgery patients and the care of severely injured patients in designated trauma centers show that trauma surgery expertise is mandatory in a central emergency department. Senior positions should also be sought to ensure high quality standards.

Cohesin mediates DNA loop extrusion by a "swing and clamp" mechanism.

Structural maintenance of chromosomes (SMC) complexes organize genome topology in all kingdoms of life and have been proposed to perform this function by DNA loop extrusion. How this process works is unknown. Here, we have analyzed how loop extrusion is mediated by human cohesin-NIPBL complexes, which enable chromatin folding in interphase cells. We have identified DNA binding sites and large-scale conformational changes that are required for loop extrusion and have determined how these are coordinated. Our results suggest that DNA is translocated by a spontaneous 50 nm-swing of cohesin's hinge, which hands DNA over to the ATPase head of SMC3, where upon binding of ATP, DNA is clamped by NIPBL. During this process, NIPBL "jumps ship" from the hinge toward the SMC3 head and might thereby couple the spontaneous hinge swing to ATP-dependent DNA clamping. These results reveal mechanistic principles of how cohesin-NIPBL and possibly other SMC complexes mediate loop extrusion.

X-ray Fluorescence Uptake Measurement of Functionalized Gold Nanoparticles in Tumor Cell Microsamples.

Quantitative cellular in vitro nanoparticle uptake measurements are possible with a large number of different techniques, however, all have their respective restrictions. Here, we demonstrate the application of synchrotron-based X-ray fluorescence imaging (XFI) on prostate tumor cells, which have internalized differently functionalized gold nanoparticles. Total nanoparticle uptake on the order of a few hundred picograms could be conveniently observed with microsamples consisting of only a few hundreds of cells. A comparison with mass spectroscopy quantification is provided, experimental results are both supported and sensitivity limits of this XFI approach extrapolated by Monte-Carlo simulations, yielding a minimum detectable nanoparticle mass of just 5 pg. This study demonstrates the high sensitivity level of XFI, allowing non-destructive uptake measurements with very small microsamples within just seconds of irradiation time.

Genome folding through loop extrusion by SMC complexes.

Genomic DNA is folded into loops and topologically associating domains (TADs), which serve important structural and regulatory roles. It has been proposed that these genomic structures are formed by a loop extrusion process, which is mediated by structural maintenance of chromosomes (SMC) protein complexes. Recent single-molecule studies have shown that the SMC complexes condensin and cohesin are indeed able to extrude DNA into loops. In this Review, we discuss how the loop extrusion hypothesis can explain key features of genome architecture; cellular functions of loop extrusion, such as separation of replicated DNA molecules, facilitation of enhancer-promoter interactions and immunoglobulin gene recombination; and what is known about the mechanism of loop extrusion and its regulation, for example, by chromatin boundaries that depend on the DNA binding protein CTCF. We also discuss how the loop extrusion hypothesis has led to a paradigm shift in our understanding of both genome architecture and the functions of SMC complexes.

STAG1 vulnerabilities for exploiting cohesin synthetic lethality in STAG2-deficient cancers.

The cohesin subunit STAG2 has emerged as a recurrently inactivated tumor suppressor in human cancers. Using candidate approaches, recent studies have revealed a synthetic lethal interaction between STAG2 and its paralog STAG1 To systematically probe genetic vulnerabilities in the absence of STAG2, we have performed genome-wide CRISPR screens in isogenic cell lines and identified STAG1 as the most prominent and selective dependency of STAG2-deficient cells. Using an inducible degron system, we show that chemical genetic degradation of STAG1 protein results in the loss of sister chromatid cohesion and rapid cell death in STAG2-deficient cells, while sparing STAG2-wild-type cells. Biochemical assays and X-ray crystallography identify STAG1 regions that interact with the RAD21 subunit of the cohesin complex. STAG1 mutations that abrogate this interaction selectively compromise the viability of STAG2-deficient cells. Our work highlights the degradation of STAG1 and inhibition of its interaction with RAD21 as promising therapeutic strategies. These findings lay the groundwork for the development of STAG1-directed small molecules to exploit synthetic lethality in STAG2-mutated tumors.

ESCO1 and CTCF enable formation of long chromatin loops by protecting cohesinSTAG1 from WAPL.

Eukaryotic genomes are folded into loops. It is thought that these are formed by cohesin complexes via extrusion, either until loop expansion is arrested by CTCF or until cohesin is removed from DNA by WAPL. Although WAPL limits cohesin's chromatin residence time to minutes, it has been reported that some loops exist for hours. How these loops can persist is unknown. We show that during G1-phase, mammalian cells contain acetylated cohesinSTAG1 which binds chromatin for hours, whereas cohesinSTAG2 binds chromatin for minutes. Our results indicate that CTCF and the acetyltransferase ESCO1 protect a subset of cohesinSTAG1 complexes from WAPL, thereby enable formation of long and presumably long-lived loops, and that ESCO1, like CTCF, contributes to boundary formation in chromatin looping. Our data are consistent with a model of nested loop extrusion, in which acetylated cohesinSTAG1 forms stable loops between CTCF sites, demarcating the boundaries of more transient cohesinSTAG2 extrusion activity.

PDS5 proteins are required for proper cohesin dynamics and participate in replication fork protection.

Cohesin is a chromatin-bound complex that mediates sister chromatid cohesion and facilitates long-range interactions through DNA looping. How the transcription and replication machineries deal with the presence of cohesin on chromatin remains unclear. The dynamic association of cohesin with chromatin depends on WAPL cohesin release factor (WAPL) and on PDS5 cohesin-associated factor (PDS5), which exists in two versions in vertebrate cells, PDS5A and PDS5B. Using genetic deletion in mouse embryo fibroblasts and a combination of CRISPR-mediated gene editing and RNAi-mediated gene silencing in human cells, here we analyzed the consequences of PDS5 depletion for DNA replication. We found that either PDS5A or PDS5B is sufficient for proper cohesin dynamics and that their simultaneous removal increases cohesin's residence time on chromatin and slows down DNA replication. A similar phenotype was observed in WAPL-depleted cells. Cohesin down-regulation restored normal replication fork rates in PDS5-deficient cells, suggesting that chromatin-bound cohesin hinders the advance of the replisome. We further show that PDS5 proteins are required to recruit WRN helicase-interacting protein 1 (WRNIP1), RAD51 recombinase (RAD51), and BRCA2 DNA repair associated (BRCA2) to stalled forks and that in their absence, nascent DNA strands at unprotected forks are degraded by MRE11 homolog double-strand break repair nuclease (MRE11). These findings indicate that PDS5 proteins participate in replication fork protection and also provide insights into how cohesin and its regulators contribute to the response to replication stress, a common feature of cancer cells.

DNA loop extrusion by human cohesin.

Eukaryotic genomes are folded into loops and topologically associating domains, which contribute to chromatin structure, gene regulation, and gene recombination. These structures depend on cohesin, a ring-shaped DNA-entrapping adenosine triphosphatase (ATPase) complex that has been proposed to form loops by extrusion. Such an activity has been observed for condensin, which forms loops in mitosis, but not for cohesin. Using biochemical reconstitution, we found that single human cohesin complexes form DNA loops symmetrically at rates up to 2.1 kilo-base pairs per second. Loop formation and maintenance depend on cohesin's ATPase activity and on NIPBL-MAU2, but not on topological entrapment of DNA by cohesin. During loop formation, cohesin and NIPBL-MAU2 reside at the base of loops, which indicates that they generate loops by extrusion. Our results show that cohesin and NIPBL-MAU2 form an active holoenzyme that interacts with DNA either pseudo-topologically or non-topologically to extrude genomic interphase DNA into loops.

Absolute quantification of cohesin, CTCF and their regulators in human cells.

The organisation of mammalian genomes into loops and topologically associating domains (TADs) contributes to chromatin structure, gene expression and recombination. TADs and many loops are formed by cohesin and positioned by CTCF. In proliferating cells, cohesin also mediates sister chromatid cohesion, which is essential for chromosome segregation. Current models of chromatin folding and cohesion are based on assumptions of how many cohesin and CTCF molecules organise the genome. Here we have measured absolute copy numbers and dynamics of cohesin, CTCF, NIPBL, WAPL and sororin by mass spectrometry, fluorescence-correlation spectroscopy and fluorescence recovery after photobleaching in HeLa cells. In G1-phase, there are ~250,000 nuclear cohesin complexes, of which ~ 160,000 are chromatin-bound. Comparison with chromatin immunoprecipitation-sequencing data implies that some genomic cohesin and CTCF enrichment sites are unoccupied in single cells at any one time. We discuss the implications of these findings for how cohesin can contribute to genome organisation and cohesion.

Werner syndrome helicase is a selective vulnerability of microsatellite instability-high tumor cells.

Targeted cancer therapy is based on exploiting selective dependencies of tumor cells. By leveraging recent functional screening data of cancer cell lines we identify Werner syndrome helicase (WRN) as a novel specific vulnerability of microsatellite instability-high (MSI-H) cancer cells. MSI, caused by defective mismatch repair (MMR), occurs frequently in colorectal, endometrial and gastric cancers. We demonstrate that WRN inactivation selectively impairs the viability of MSI-H but not microsatellite stable (MSS) colorectal and endometrial cancer cell lines. In MSI-H cells, WRN loss results in severe genome integrity defects. ATP-binding deficient variants of WRN fail to rescue the viability phenotype of WRN-depleted MSI-H cancer cells. Reconstitution and depletion studies indicate that WRN dependence is not attributable to acute loss of MMR gene function but might arise during sustained MMR-deficiency. Our study suggests that pharmacological inhibition of WRN helicase function represents an opportunity to develop a novel targeted therapy for MSI-H cancers.

A neurobiological pathway to smoking in adolescence: TTC12-ANKK1-DRD2 variants and reward response.

The TTC12-ANKK1-DRD2 gene-cluster has been implicated in adult smoking. Here, we investigated the contribution of individual genes in the TTC12-ANKK1-DRD2 cluster in smoking and their association with smoking-associated reward processing in adolescence. A meta-analysis of TTC12-ANKK1-DRD2 variants and self-reported smoking behaviours was performed in four European adolescent cohorts (N = 14,084). The minor G-allele of rs2236709, mapping TTC12, was associated with self-reported smoking (p = 5.0 × 10-4) and higher plasma cotinine levels (p = 7.0 × 10-5). This risk allele was linked to an increased ventral-striatal blood-oxygen level-dependent (BOLD) response during reward anticipation (n = 1,263) and with higher DRD2 gene expression in the striatum (p = 0.013), but not with TTC12 or ANKK gene expression. These data suggest a role for the TTC12-ANKK1-DRD2 gene-cluster in adolescent smoking behaviours, provide evidence for the involvement of DRD2 in the early stages of addiction and support the notion that genetically-driven inter-individual differences in dopaminergic transmission mediate reward sensitivity and risk to smoking.

A mechanism of cohesin-dependent loop extrusion organizes zygotic genome architecture.

Fertilization triggers assembly of higher-order chromatin structure from a condensed maternal and a naïve paternal genome to generate a totipotent embryo. Chromatin loops and domains have been detected in mouse zygotes by single-nucleus Hi-C (snHi-C), but not bulk Hi-C. It is therefore unclear when and how embryonic chromatin conformations are assembled. Here, we investigated whether a mechanism of cohesin-dependent loop extrusion generates higher-order chromatin structures within the one-cell embryo. Using snHi-C of mouse knockout embryos, we demonstrate that the zygotic genome folds into loops and domains that critically depend on Scc1-cohesin and that are regulated in size and linear density by Wapl. Remarkably, we discovered distinct effects on maternal and paternal chromatin loop sizes, likely reflecting differences in loop extrusion dynamics and epigenetic reprogramming. Dynamic polymer models of chromosomes reproduce changes in snHi-C, suggesting a mechanism where cohesin locally compacts chromatin by active loop extrusion, whose processivity is controlled by Wapl. Our simulations and experimental data provide evidence that cohesin-dependent loop extrusion organizes mammalian genomes over multiple scales from the one-cell embryo onward.

Topologically associating domains and chromatin loops depend on cohesin and are regulated by CTCF, WAPL, and PDS5 proteins.

Mammalian genomes are spatially organized into compartments, topologically associating domains (TADs), and loops to facilitate gene regulation and other chromosomal functions. How compartments, TADs, and loops are generated is unknown. It has been proposed that cohesin forms TADs and loops by extruding chromatin loops until it encounters CTCF, but direct evidence for this hypothesis is missing. Here, we show that cohesin suppresses compartments but is required for TADs and loops, that CTCF defines their boundaries, and that the cohesin unloading factor WAPL and its PDS5 binding partners control the length of loops. In the absence of WAPL and PDS5 proteins, cohesin forms extended loops, presumably by passing CTCF sites, accumulates in axial chromosomal positions (vermicelli), and condenses chromosomes. Unexpectedly, PDS5 proteins are also required for boundary function. These results show that cohesin has an essential genome-wide function in mediating long-range chromatin interactions and support the hypothesis that cohesin creates these by loop extrusion, until it is delayed by CTCF in a manner dependent on PDS5 proteins, or until it is released from DNA by WAPL.

Synthetic lethality between the cohesin subunits STAG1 and STAG2 in diverse cancer contexts.

Recent genome analyses have identified recurrent mutations in the cohesin complex in a wide range of human cancers. Here we demonstrate that the most frequently mutated subunit of the cohesin complex, STAG2, displays a strong synthetic lethal interaction with its paralog STAG1. Mechanistically, STAG1 loss abrogates sister chromatid cohesion in STAG2 mutated but not in wild-type cells leading to mitotic catastrophe, defective cell division and apoptosis. STAG1 inactivation inhibits the proliferation of STAG2 mutated but not wild-type bladder cancer and Ewing sarcoma cell lines. Restoration of STAG2 expression in a mutated bladder cancer model alleviates the dependency on STAG1. Thus, STAG1 and STAG2 support sister chromatid cohesion to redundantly ensure cell survival. STAG1 represents a vulnerability of cancer cells carrying mutations in the major emerging tumor suppressor STAG2 across different cancer contexts. Exploiting synthetic lethal interactions to target recurrent cohesin mutations in cancer, e.g. by inhibiting STAG1, holds the promise for the development of selective therapeutics.

Where There is Smoke There is Fear-Impaired Contextual Inhibition of Conditioned Fear in Smokers.

The odds-ratio of smoking is elevated in populations with neuropsychiatric diseases, in particular in the highly prevalent diagnoses of post-traumatic stress and anxiety disorders. Yet, the association between smoking and a key dimensional phenotype of these disorders-maladaptive deficits in fear learning and fear inhibition-is unclear. We therefore investigated acquisition and memory of fear and fear inhibition in healthy smoking and non-smoking participants (N=349, 22% smokers). We employed a well validated paradigm of context-dependent fear and safety learning (day 1) including a memory retrieval on day 2. During fear learning, a geometrical shape was associated with an aversive electrical stimulation (classical fear conditioning, in danger context) and fear responses were extinguished within another context (extinction learning, in safe context). On day 2, the conditioned stimuli were presented again in both contexts, without any aversive stimulation. Autonomic physiological measurements of skin conductance responses as well as subjective evaluations of fear and expectancy of the aversive stimulation were acquired. We found that impairment of fear inhibition (extinction) in the safe context during learning (day 1) was associated with the amount of pack-years in smokers. During retrieval of fear memories (day 2), smokers showed an impairment of contextual (safety context-related) fear inhibition as compared with non-smokers. These effects were found in physiological as well as subjective measures of fear. We provide initial evidence that smokers as compared with non-smokers show an impairment of fear inhibition. We propose that smokers have a deficit in integrating contextual signs of safety, which is a hallmark of post-traumatic stress and anxiety disorders.

Effects of Medial Orbitofrontal Cortex Lesions on Self-Control in Intertemporal Choice.

Many decisions involve a trade-off between the temporal proximity of a reward and its magnitude. A range of clinical conditions are associated with poor self-control during such intertemporal choices, such that smaller rewards that are received sooner are preferred over larger rewards that are received later to a greater extent [1, 2]. According to a prominent neural model of self-control [3-6], subjective reward values are represented in the medial orbitofrontal cortex (mOFC) at the time of choice [7-9]. Successful self-control in this model is then thought to depend on a modulation of these mOFC value representations via the lateral prefrontal cortex (lPFC) [3, 6]. Here we directly tested three key predictions of this model in patients with lesions to the mOFC (n = 9) and matched controls (n = 19). First, we show that mOFC lesions disrupt choice-free valuation ratings. This finding provides causal evidence for a role of the mOFC in reward valuation and contrasts with the effects of lPFC disruption [6]. Second, we show that mOFC damage indeed decreases self-control during intertemporal choice, replicating previous findings [10]. Third, extending these previous observations, we show that the effect of mOFC damage on intertemporal choice depends on the actual self-control demands of the task. Our findings thus provide causal evidence for a role of mOFC in reward valuation and are compatible with the idea that mOFC damage affects self-control specifically under conditions that might normally require a modulation of mOFC value representations, e.g., by the lPFC.

ARHGEF17 is an essential spindle assembly checkpoint factor that targets Mps1 to kinetochores.

To prevent genome instability, mitotic exit is delayed until all chromosomes are properly attached to the mitotic spindle by the spindle assembly checkpoint (SAC). In this study, we characterized the function of ARHGEF17, identified in a genome-wide RNA interference screen for human mitosis genes. Through a series of quantitative imaging, biochemical, and biophysical experiments, we showed that ARHGEF17 is essential for SAC activity, because it is the major targeting factor that controls localization of the checkpoint kinase Mps1 to the kinetochore. This mitotic function is mediated by direct interaction of the central domain of ARHGEF17 with Mps1, which is autoregulated by the activity of Mps1 kinase, for which ARHGEF17 is a substrate. This mitosis-specific role is independent of ARHGEF17's RhoGEF activity in interphase. Our study thus assigns a new mitotic function to ARHGEF17 and reveals the molecular mechanism for a key step in SAC establishment.