Miracle-dz/UniFAR-FTUs huggingface dataset:
Degenerating Drosophila Larval Epidermal Cells Drive Thorax Closure

Adult thorax formation in Drosophila begins during pre-pupal development by fusion of its two contralateral progenitor halves, the heminotal epithelia (HE). HEs migrate and replace an underlying cell layer of thoracic larval epidermal cells (LECs) during a morphogenetic process called thorax closure. The LEC layer has so far been proposed to be a passive substrate over which HEs migrate before their zipping. By contrast, here we show that the pull forces generated within the LEC layer drive HE migration. During thorax closure, the LECs display actomyosin-mediated contraction, via enrichment of non-muscle myosin-II and actin, besides squamous-to-pseudostratified columnar epithelial transition and tissue shrinkage. This shrinkage of the LEC layer is further accompanied by cell extrusion and death, that prevent overcrowding of LECs, thereby promoting further shrinkage. The pull forces thus generated by the shrinking LEC layer are then relayed to the HEs by their mutual adhesions via βPS1 (Mys) and αPS3 (Scb) integrins. Suppression of cell death in the LEC layer by a gain of p35 leads to cell overcrowding, which impedes HE migration and zipping. Further, knockdown of sqh, the light chain of non-muscle myosin II, in LECs or integrins (mys or scb) in either the LEC layer or in the HEs, or both abrogate thorax closure. Mathematical modeling also reveals the biophysical underpinnings of the forces that drive this tissue closure process wherein a degenerating LEC layer mediates its succession by the future adult primodia. These essential principles of thorax closure appear ancient in origin and recur in multiple morphogenetic contexts and tissue repair.
Targeting Allostery in the Dynein Motor Domain with Small Molecule Inhibitors

Cytoplasmic dyneins are AAA (ATPase associated with diverse cellular activities) motor proteins responsible for microtubule minus-end-directed intracellular transport. Dynein’s unusually large size, four distinct nucleotide-binding sites, and the existence of closely-related isoforms with different functions, pose challenges for the design of potent and selective chemical inhibitors. Here we use structural approaches to develop a model for the inhibition of a well-characterized S. cerevisiae dynein construct by pyrazolo-pyrimidinone-based compounds. These data, along with single molecule experiments and mutagenesis studies, indicate that the compounds likely inhibit dynein by engaging the regulatory ATPase sites in the AAA3 and AAA4 domains, and not by interacting with dynein’s main catalytic site in the AAA1 domain. A double Walker B mutant in AAA3 and AAA4 is an inactive enzyme, suggesting that inhibiting these regulatory sites can have a similar effect to inhibiting AAA1. Our findings reveal how chemical inhibitors can be designed to disrupt allosteric communication across dynein’s AAA domains.
Chronic in vivo STED nanoscopy uncovers multiple drivers of shape volatility in stable cortical spines

Excitatory synapses on spines of pyramidal neurons are considered a central locus of cortical memory traces. Here we introduce chronic in vivo STED nanoscopy to superresolve dendritic spines in mouse neocortex for up to one month and assess on-going spine remodeling at nanoscale resolution. We find that distinct features of spine geometry, such as neck length and head size exhibit essentially uncorrelated dynamics, indicating multiple independent drivers of spine remodeling. For neck length, neck width and head size, the magnitude of this remodeling indicates substantial fluctuations in synaptic strength, which is exaggerated in a mouse model of neurodegeneration. Despite this high degree of volatility, all spine features influencing synaptic strength also exhibit persistent components that are maintained over long periods of time. Thus, at the nanoscale, stable dendritic spines exhibit a delicate balance of stability and volatility.
A Tale of Two Tails: Efficient Profiling of Protein Degraders by Specific Functional and Target Engagement Readouts

Targeted protein degradation represents an area of great interest, potentially offering improvements with respect to dosing, side effects, drug resistance, and reaching “undruggable” proteins compared with traditional small-molecule therapeutics. A major challenge in the design and characterization of degraders acting as molecular glues is that binding of the molecule to the protein of interest (PoI) is not needed for efficient and selective protein degradation; instead, one needs to understand the interaction with the responsible ligase. Similarly, for proteasome targeting chimeras (PROTACs), understanding the binding characteristics of the PoI alone is not sufficient. Therefore, simultaneously assessing the binding to both PoI and the E3 ligase as well as the resulting degradation profile is of great value. The cellular thermal shift assay (CETSA) is an unbiased cell-based method, designed to investigate the interaction of compounds with their cellular protein targets by measuring compound-induced changes in protein thermal stability. In combination with mass spectrometry (MS), CETSA can simultaneously evaluate compound-induced changes in the stability of thousands of proteins. We have used CETSA MS to profile a number of protein degraders, including molecular glues (e.g., immunomodulatory drugs) and PROTACs, to understand mode of action and to deconvolute off-target effects in intact cells. Within the same experiment, we were able to monitor both target engagement by observing changes in protein thermal stability as well as efficacy by simultaneous assessment of protein abundances. This allowed us to correlate target engagement (i.e., binding to the PoI and ligases) and functional readout (i.e., degrader induced protein degradation).
SingleChannelNet: A model for Automatic Sleep Stage Classification with Raw Single-Channel EEG

In diagnosing sleep disorders, sleep stage classification is a very essential yet time-consuming process. Most of the existing state-of-the-art approaches rely on hand-crafted features and multi-modality polysomnography (PSG) data, where prior knowledge is compulsory and high computation cost can be expected. Besides, few studies are able to obtain high accuracy sleep staging using raw single-channel electroencephalogram (EEG). To overcome these shortcomings, this paper proposes an end-to-end framework with a deep neural network, namely SingleChannelNet, for automatic sleep stage classification based on raw single-channel EEG. The proposed model utilizes a 90s epoch as the textual input and employs two multi-convolution blocks and several max-average pooling layers to learn different scales of feature representations. To demonstrate the efficiency of the proposed model, we evaluate our model using different raw single-channel EEGs (C4/A1 and Fpz-Cz) on two different datasets (SHHS and Sleep-EDF datasets). Experimental results show that the proposed architecture can achieve better overall accuracy and Cohen’s kappa (SHHS: 89.2%-84.8%, Sleep-EDF: 89.4%-85%) compared with state-of-the-art approaches. Additionally, the proposed model can learn features automatically for sleep stage classification using different single-channel EEGs with distinct sampling rates from different datasets without using any hand-engineered features.
Automated cell tracking using StarDist and TrackMate

The ability of cells to migrate is a fundamental physiological process involved in embryonic development, tissue homeostasis, immune surveillance, and wound healing. Therefore, the mechanisms governing cellular locomotion have been under intense scrutiny over the last 50 years. One of the main tools of this scrutiny is live-cell quantitative imaging, where researchers image cells over time to study their migration and quantitatively analyze their dynamics by tracking them using the recorded images. Despite the availability of computational tools, manual tracking remains widely used among researchers due to the difficulty setting up robust automated cell tracking and large-scale analysis. Here we provide a detailed analysis pipeline illustrating how the deep learning network StarDist can be combined with the popular tracking software TrackMate to perform 2D automated cell tracking and provide fully quantitative readouts. Our proposed protocol is compatible with both fluorescent and widefield images. It only requires freely available and open-source software (ZeroCostDL4Mic and Fiji), and does not require any coding knowledge from the users, making it a versatile and powerful tool for the field. We demonstrate this pipeline’s usability by automatically tracking cancer cells and T cells using fluorescent and brightfield images. Importantly, we provide, as supplementary information, a detailed step-by-step protocol to allow researchers to implement it with their images.
Higher-order sensorimotor circuit of the brain’s global network supports human consciousness

The neural correlates of consciousness, defined as the minimum neuronal mechanisms sufficient for any conscious percept, are usually subject to different interpretations depending on whether one uses measures of local or global brain activities. We argue that the local regions may support consciousness by serving as hubs within the brain’s global network. We adopt a unique functional magnetic resonance imaging resting state dataset that encompasses various conscious states, including non-rapid eye movement (NREM)-sleep, rapid eye movement (REM)-sleep, anesthesia, and brain injury patients. Using a graph-theoretical measure for detecting local hubs within the brain’s global network, we identify various higher-order sensory and motor regions as hubs with significantly reduced degree centrality during unconsciousness. Additionally, these regions form a sensorimotor circuit which correlates with levels of consciousness. Our findings suggest that integration of higher-order sensorimotor function may be a key mechanism of consciousness. This opens novel perspectives for therapeutic modulation of unconsciousness.
Attention affects overall gain but not selective contrast at meter frequencies in the neural processing of rhythm

When listening to music, humans spontaneously perceive and synchronize movement to periodic pulses of meter. A growing body of evidence suggests that this widespread ability is related to neural processes that selectively enhance meter periodicities. However, to what extent these neural processes are affected by the attentional state of the listener remains largely unknown. Here, we recorded EEG while participants listened to auditory rhythms and detected small changes in tempo or pitch of the stimulus, or performed a visual task. The overall neural response to the auditory input decreased when participants attended the visual modality, indicating generally lower sensitivity to acoustic information. However, the selective contrast at meter periodicities did not differ across the three tasks. Moreover, this selective contrast could be trivially accounted for by biologically-plausible models of subcortical auditory processing, but only when meter periodicities were already prominent in the acoustic input. However, when meter periodicities were not prominent in the auditory input, the EEG responses could not be explained by low-level processing. This was also confirmed by early auditory responses that originate predominantly in early auditory areas and were recorded in the same EEG. The contrast at meter periodicities in these early responses was consistently smaller than in the EEG responses originating mainly from higher-level processing stages. Together, these results demonstrate that selective contrast at meter periodicities involves higher-level neural processes that may be engaged automatically, irrespective of behavioral context. This robust shaping of the neural representation of rhythm might thus contribute to spontaneous and effortless synchronization to musical meter in humans across cultures.
Crystal structure of GH43 exo-β-1,3-galactanase from the basidiomycete Phanerochaete chrysosporium provides insights into the mechanism of bypassing side chains

Arabinogalactan proteins (AGPs) are functional plant proteoglycans, but their functions are largely unexplored, mainly because of the complexity of the sugar moieties, which are generally analyzed with the aid of glycoside hydrolases. In this study, we solved the apo and liganded structures of exo-β-1,3-galactanase from the basidiomycete Phanerochaete chrysosporium (Pc1,3Gal43A), which specifically cleaves AGPs. It is composed of a glycoside hydrolase family 43 subfamily 24 (GH43_sub24) catalytic domain together with a carbohydrate-binding module family (CBM) 35 binding domain. GH43_sub24 lacks the catalytic base Asp that is conserved among other GH43 subfamilies. Crystal structure and kinetic analyses indicated that the tautomerized imidic acid function of Gln263 serves instead as the catalytic base residue. Pc1,3Gal43A has three subsites that continue from the bottom of the catalytic pocket to the solvent. Subsite -1 contains a space that can accommodate the C-6 methylol of Gal, enabling the enzyme to bypass the β-1,6-linked galactan side chains of AGPs. Furthermore, the galactan-binding domain in CBM35 has a different ligand interaction mechanism from other sugar-binding CBM35s. Some of the residues involved in ligand recognition differ from those of galactomannan-binding CBM35, including substitution of Trp for Gly, which affects pyranose stacking, and substitution of Asn for Asp in the lower part of the binding pocket. Pc1,3Gal43A WT and its mutants at residues involved in substrate recognition are expected to be useful tools for structural analysis of AGPs. Our findings should also be helpful in engineering designer enzymes for efficient utilization of various types of biomass.
A highly conserved 310 helix within the kinesin motor domain is critical for kinesin function and human health

An uncharacterized structural element in the kinesin motor domain is shown to be critical for motor function and human health. KIF1A is a critical cargo transport motor within neurons. More than 100 known mutations result in KIF1A-associated neurological disorder (KAND), a degenerative condition for which there is no cure. A missense mutation, P305L, was identified in children diagnosed with KAND, but the molecular basis for the disease is unknown. We find that this conserved residue is part of an unusual 310 helix immediately adjacent to the family-specific K-loop, which facilitates a high microtubule-association rate. We find that the mutation negatively affects several biophysical parameters of the motor. However, the microtubule-association rate of the motor is most markedly affected, revealing that the presence of an intact K-loop is not sufficient for its function. We hypothesize that the 310 helix facilitates a specific K-loop conformation that is critical for its function. We find that the function of this proline is conserved in kinesin-1, revealing a fundamental principle of the kinesin motor mechanism.
Common Mental Disorders and Disability Across Cultures: Results From the WHO Collaborative Study on Psychological Problems in General Health Care

Objective. —To examine the impact of common mental illness on functional disability and the cross-cultural consistency of this relationship while controlling for physical illness. A secondary objective was to determine the level of disability associated with specific psychiatric disorders. Design. —A cross-sectional sample selected by two-stage sampling. Setting. —Primary health care facilities in 14 countries covering most major cultures and languages. Patients. —A total of 25916 consecutive attenders of these facilities were screened for psychopathology using the General Health Questionnaire (96% response). Screened patients were sampled from the General Health Questionnaire score strata for the second-stage Composite International Diagnostic Interview administered to 5447 patients (62% response). Main Outcome Measures. —Patient-reported physical disability, number of disability days, and interviewer-rated occupational role functioning. Results. —After controlling for physical disease severity, psychopathology was consistently associated with increased disability. Physical disease severity was an independent, although weaker, contributor to disability. A dose-response relationship was found between severity of mental illness and disability. Disability was most prominent among patients with major depression, panic disorder, generalized anxiety, and neurasthenia; disorder-specific differences were modest after controlling for psychiatric comorbidity. Results were consistent across disability measures and across centers. Conclusions. —The consistent relationship of psychopathology and disability indicates the compelling personal and socioeconomic impact of common mental illnesses across cultures. This suggests the importance of impairments of higher-order human capacities (eg, emotion, motivation, and cognition) as determinants of functional disability. (JAMA. 1994;272:1741-1748)
Working nights and lower leisure-time physical activity associate with chronic pain in Southern African truck drivers: a cross-sectional study

Background Globally the trucking industry employs millions of people. Recently the prevalence of chronic pain in Southern African truck drivers was reported at 10%. We investigated factors associated with chronic pain in truck drivers including mental health, physical activity, and sleep, as no study has done so. Methods Southern African truck drivers were recruited at truck stops in Gauteng and Free State Provinces, South Africa (n=614). Chronic pain was defined as pain present for at least the last three months. Depressive symptoms were assessed with the Patient Health Questionnaire-9, post-traumatic stress disorder with the Posttraumatic Stress Disorder Checklist for DSM-5 (PCL-5), exposure to traumatic events with the Life Events Checklist-5 (LEC-5) and daytime sleepiness with the Epworth Sleepiness Scale. Sleep quality was measured on a four-point Likert scale. Leisure-time physical activity was measured using the Godin-Shephard leisure-time physical activity questionnaire. Associations between these factors, demographic factors and chronic pain were investigated. Results Six hundred and fourteen male truck drivers were recruited. Multivariate analysis showed that working ≥ 2 nights/week (OR=2.68, 95% CI=1.55-4.68) was associated with chronic pain and physical activity was protective (OR=0.97, 95% CI 0.95-0.98). In an exploratory analysis, greater depressive symptoms (p=0.004), daytime sleepiness (p=0.01) and worse sleep quality (p=0.001) was associated with working ≥ 2 nights/week. Lower leisure-time physical activity was associated with worse sleep quality (p=0.006), but not daytime sleepiness or depressive symptoms (p>0.05). Conclusions There is a clear relationship between working nights and activity levels, and chronic pain, sleep quality, and depression in truck drivers.
Membrane binding controls ordered self-assembly of animal septins

Septins are conserved cytoskeletal proteins that regulate cell cortex mechanics. The mechanisms of their interactions with the plasma membrane remain poorly understood. Here we show by cell-free reconstitution that membrane binding requires electrostatic interactions of septins with anionic lipids and promotes the ordered self-assembly of fly septins into filamentous meshworks. Transmission electron microscopy reveals that both fly and mammalian septins form arrays of single and paired filaments. Atomic force microscopy and quartz crystal microbalance demonstrate that the fly filaments form mechanically rigid, 12 to 18 nm thick, double layers of septins. By contrast, C-terminally truncated septin mutants form 4 nm thin monolayers, indicating that stacking requires the C-terminal coiled coils on DSep2 and Pnut subunits. Our work shows that membrane binding is required for fly septins to form ordered arrays of single and paired filaments and provides new insights into the mechanisms by which septins may regulate cell surface mechanics.
Mycobacterium smegmatis expands across surfaces using hydraulic sliding

Mycobacterium smegmatis spreads over soft agar surfaces by sliding motility, a form of passive motility in which growth and reduction of surface adhesion enable the bacteria to push each other outwards. Hence, sliding motility is mostly associated with round colonies. However, M. smegmatis sliding colonies can also produce long, pointed dendrites. Round sliding colonies were readily reproduced, but our non-round colonies were different from those seen previously. The latter (named digitate colonies) had centimetre-long linear protrusions, containing a central channel filled with a free-flowing suspension of M. smegmatis and solid aggregates. Digitate colonies had both a surface pellicle and an inner biofilm component surrounding a central channel, which sat in a cleft in the agar. Time-lapse microscopy showed that the expansion of the fluid-filled channel enabled the lengthwise extension of the protrusions without any perceptible growth of the bacteria taking place. These observations represent a novel type of sliding motility, named hydraulic sliding, associated with a specialised colony structure and the apparent generation of force by expansion of a liquid core. As this structure requires pellicle formation without an initial liquid culture it implies the presence of an unstudied mycobacterial behaviour that may be important for colonisation and virulence. Originality-Significance Statement This study is the first to identify a new form of passive motility in the mycobacteria; hydraulic sliding, in which liquid expansion is the cause of motility. This form of motility has so far never been described in bacteria. The study also reveals new ways mycobacteria can form biofilms and colonize complex three-dimensional substrates, aspects of mycobacterial biology that are important for infection, pathogenesis and vaccine development. Author Summary Mycobacterium smegmatis is used as a non-pathogenic model organism for pathogenic mycobacteria. During growth, M. smegmatis can move passively over soft agar surfaces by a process called sliding motility, whereby colony growth directly pushes cells outwards. Although passive, sliding motility is believed to be important in allowing bacteria to colonise surfaces. Sliding motility however does not fully account for how M. smegmatis produces dendritic colonies. We attempted to generate dendritic colonies but found instead that the cells produced colonies that had larger protrusions radiating from them (digitate colonies). Digitate colonies are a previously unobserved phenomenon, in that the bacteria create a biofilm-lined, fluid-filled, pellicle-covered, deep cleft in the agar and move across the surface by the expansion of the contained liquid core of the protrusions. Given the new structure and the new mechanism of expansion we have termed this set of behaviours hydraulic sliding. These observations are important as it is a new variation in the way bacteria can move, generate biofilms (notably mycobacterial pellicle) and colonize complex three-dimensional substrates.
Structural biologists, let’s mind our colors

In structural biology, most figures of macromolecules are aimed at those well-versed in structure, requiring prior familiarity with scales and commonly used color schemes. Yet, as structural biology becomes democratized with the increasing pace of structure determination, the accessibility of structural data is paramount. Here, we identify three keys, and have written accompanying software plugins, for structural biologists to create figures truer to the hard-won data and clearer across different modes of color vision and to non-expert readers. Use perceptually uniform colormaps Consider readers with different modes of color vision Be explicit about scales and color usage