Miracle-dz/UniFAR-FTUs huggingface dataset:
Membrane receptor MerTK is a newly identified transcriptional regulator that associates to chromatin as nanoclusters during human DC differentiation

MerTK is a transmembrane receptor tyrosine kinase (RTK) mainly expressed in dendritic cells (DCs) and macrophages where it plays an important role in immunotolerance, but also in activating oncogenic signalling pathways. Albeit MerTK is exploited as clinical target in cancer and auto-immune disorders, the mechanisms that regulate its diverse functions are poorly understood. Here, we identified a remarkably high pool of the full receptor in the nucleus of human DCs. Nuclear translocation was ligand-dependent. Importantly, MerTK nuclear levels correlated to DC differentiation and were spatiotemporally regulated by the transmembrane receptor LRP-1. Using dual-colour super-resolution nanoscopy we discovered that nuclear MerTK forms nanoclusters, whose strength strongly depends on chromatin accessibility during DC differentiation. We finally revealed high transcription capacity of MerTK. Overall, our work indicates that nuclear MerTK acts as a transcription factor regulating DC differentiation, thus implicating for the first time a physiological function for RTK nuclear translocation in immunity.
Rhodobacterales and Rhizobiales Are Associated With Stony Coral Tissue Loss Disease and Its Suspected Sources of Transmission

In 2014, Stony Coral Tissue Loss Disease (SCTLD) was first detected off the coast of Miami, FL, United States, and continues to persist and spread along the Florida Reef Tractr (FRT) and into the Caribbean. SCTLD can have up to a 61% prevalence in reefs and has affected at least 23 species of scleractinian corals. This has contributed to the regional near-extinction of at least one coral species, Dendrogyra cylindrus. Initial studies of SCTLD indicate microbial community shifts and cessation of lesion progression in response to antibiotics on some colonies. However, the etiology and abiotic sources of SCTLD transmission are unknown. To characterize SCTLD microbial signatures, we collected tissue samples from four affected coral species: Stephanocoenia intersepta, Diploria labyrinthiformis, Dichocoenia stokesii, and Meandrina meandrites. Tissue samples were from apparently healthy (AH) corals, and unaffected tissue (DU) and lesion tissue (DL) on diseased corals. Samples were collected in June 2018 from three zones: (1) vulnerable (ahead of the SCTLD disease boundary in the Lower Florida Keys), (2) endemic (post-outbreak in the Upper Florida Keys), and (3) epidemic (SCTLD was active and prevalent in the Middle Florida Keys). From each zone, sediment and water samples were also collected to identify whether they may serve as potential sources of transmission for SCTLD-associated microbes. We used 16S rRNA gene amplicon high-throughput sequencing methods to characterize the microbiomes of the coral, water, and sediment samples. We identified a relatively higher abundance of the bacteria orders Rhodobacterales and Rhizobiales in DL tissue compared to AH and DU tissue. Also, our results showed relatively higher abundances of Rhodobacterales in water from the endemic and epidemic zones compared to the vulnerable zone. Rhodobacterales and Rhizobiales identified at higher relative abundances in DL samples were also detected in sediment samples, but not in water samples. Our data indicate that Rhodobacterales and Rhizobiales may play a role in SCTLD and that sediment may be a source of transmission for Rhodobacterales and Rhizobiales associated with SCTLD lesions.
Human organs-on-chips as tools for repurposing approved drugs as potential influenza and COVID19 therapeutics in viral pandemics

Rapidly spreading viral pandemics, such as those caused by influenza and SAR-CoV-2 (COVID19), require rapid action and the fastest way to combat this challenge is by repurposing existing drugs as anti-viral therapeutics. Here we first show that human organ-on-a-chip (Organ Chip) microfluidic culture devices lined by a highly differentiated, primary, human lung airway epithelium cultured under an air-liquid interface and fed by continuous medium flow can be used to model virus entry, replication, strain-dependent virulence, host cytokine production, and recruitment of circulating immune cells in response to infection by influenza, as well as effects of existing and novel therapeutics. These Airway Chips, which contain human lung epithelial cells that express high levels of ACE2 and TMPRSS2, were then used to assess the inhibitory activities of 7 clinically approved drugs (chloroquine, arbidol, toremifene, clomiphene, amodiaquine, verapamil, and amiodarone) that we found inhibit infection by viral pseudoparticles expressing SARS-CoV-2 spike protein in human Huh-7 cells, and others recently showed suppress infection by native SARS-CoV-2 in Vero cells. However, when these drugs were administered under flow at the maximal concentration in blood reported in clinical studies in human Airway Chips, only two of these drugs - amodiaquine and toremifene - significantly inhibited entry of the pseudotyped SARS-CoV-2 virus. This work suggests that human Organ Chip technology may be used in conjunction with existing rapid cell-based screening assays to study human disease pathogenesis and expedite drug repurposing in biothreat crises caused by pandemic viruses.
Disordered Proteins Enable Histone Chaperoning on the Nucleosome

Proteins with highly charged disordered regions are abundant in the nucleus, where many of them interact with nucleic acids and control key processes such as transcription. The functional advantages conferred by protein disorder, however, have largely remained unclear. Here we show that disorder can facilitate a remarkable regulatory mechanism involving molecular competition. Single-molecule experiments demonstrate that the human linker histone H1 binds to the nucleosome with ultra-high affinity. However, the large-amplitude dynamics of the positively charged disordered regions of H1 persist on the nucleosome and facilitate the interaction with the highly negatively charged and disordered histone chaperone prothymosin α. Consequently, prothymosin α can efficiently invade the H1-nucleosome complex and displace H1 via competitive substitution. By integrating experiments and simulations, we establish a molecular model that rationalizes this process structurally and kinetically. Given the abundance of charged disordered regions in the nuclear proteome, this mechanism may be widespread in cellular regulation.
Loss of Caspase-8 function in combination with SMAC mimetic treatment sensitizes Head and Neck Squamous Carcinoma to radiation through induction of necroptosis

Caspase-8 (CASP8) is one of the most frequently mutated genes in head and neck squamous carcinomas (HNSCC), and mutations of CASP8 are associated with poor overall survival. The distribution of these mutations in HNSCC suggests that they are likely to be inactivating. Inhibition of CASP8 has been reported to sensitize cancer cells to necroptosis, a unique cell death mechanism. Here, we evaluated how CASP8 regulates necroptosis in HSNCC using cell line models and syngeneic mouse xenografts. In vitro, knockdown of CASP8 rendered HNSCCs susceptible to necroptosis induced by a second mitochondria-derived activator of caspase (SMAC) mimetic, Birinapant, when combined with pan-caspase inhibitors Z-VAD-FMK or Emricasan. Strikingly, inhibition of CASP8 function via knockdown or Emricasan treatment was associated with enhanced radiation killing by Birinapant through induction of necroptosis. In a syngeneic mouse model of oral cancer, Birinapant, particularly when combined with radiation delayed tumor growth and enhanced survival under CASP8 loss. Exploration of molecular underpinnings of necroptosis sensitivity confirmed that the level of functional receptor-interacting serine/threonine-protein kinase-3 (RIP3), a key enzyme in the necroptosis pathway was crucial in determining susceptibility to this mode of death. Although an in vitro screen revealed that many HNSCC cell lines were resistant to necroptosis due to low levels of RIP3, patient tumors maintain RIP3 expression and should therefore remain sensitive. Collectively, these results suggest that targeting the necroptosis pathway with SMAC mimetics, especially in combination with radiation, may be a relevant therapeutic approach in HNSCC with compromised CASP8 status, provided that RIP3 function is maintained. Significance CASP8 status regulates necroptotic death in HNSCC and this pathway can be exploited therapeutically.
capCLIP: a new tool to probe protein synthesis in human cells through capture and identification of the eIF4E-mRNA interactome

Translation of eukaryotic mRNAs starts with binding of the m7G cap to the protein eIF4E followed by recruitment of other translation initiation factors. eIF4E’s essential role in translation suggests the cellular eIF4E-mRNA interactome (or ‘eIF4E cap-ome’) may serve as a faithful proxy of cellular translational activity. Here we describe capCLIP, a novel method to systematically capture and quantify the eIF4E cap-ome. To validate capCLIP, we identified the cap-omes in human cells ± the partial mTORC1 inhibitor rapamycin. As expected, TOP (terminal oligopyrimidine) mRNA representation is systematically reduced in rapamycin-treated cells. capCLIP tag data permits refinement of a 7-nucleotide TOP motif (5′-CUYUYYC-3′). We also apply capCLIP to probe the consequences of phosphorylation of eIF4E, whose function had remained unclear. eIF4E phosphorylation drives an overall reduction in eIF4E-mRNA association; strikingly, mRNAs most sensitive to phosphorylation possess short 5′-UTRs. capCLIP provides a sensitive and comprehensive measure of cellular translational activity. We foresee its application as a high-throughput way to assess translation in contexts not amenable to existing methodologies.
Locus coeruleus patterns differentially modulate learning and valence in rat via the ventral tegmental area and basolateral amygdala respectively

The locus coeruleus (LC), the main source of forebrain norepinephrine, produces phasic and tonic firing patterns that are theorized to have distinct functional consequences. However, how different firing modes affect learning and valence coding of sensory information are unknown. Here bilateral optogenetic activation of rat LC neurons using 10-Hz phasic trains of either 300 msec or 10 sec accelerates acquisition of a food-rewarded similar odor discrimination, but not a dissimilar odor discrimination, consistent with LC-supported enhanced pattern separation and plasticity. Similar odor discrimination learning is impaired by noradrenergic blockade in the piriform cortex (PC). However, here 10-Hz LC phasic light-mediated learning facilitation is prevented by a dopaminergic antagonist in the PC, or by ventral tegmental area (VTA) silencing with lidocaine, suggesting an LC-VTA-PC dopamine circuitry mediates 10-Hz phasic learning facilitation. Tonic stimulation at 10 Hz did not alter odor discrimination acquisition, and was less effective in activating VTA DA neurons. For valence encoding, tonic stimulation at 25 Hz induced freezing, anxiety and conditioned odor aversion, while 10-Hz phasic stimulation produced an odor preference consistent with positive valence. Noradrenergic blockade in the basolateral amygdala (BLA) prevented conditioned odor preference and aversion induced by 10-Hz phasic and 25-Hz tonic light respectively. CTB retro-labeling showed relatively larger engagement of nucleus accumbens projecting neurons over central amygdala projecting neurons in the BLA with 10-Hz LC phasic activation, compared to 25-Hz tonic. These outcomes argue that LC pauses, as well as LC firing frequencies, differentially influence both target networks and behaviour.
ISG15 drives immune pathology and respiratory failure during viral infection

Cytokine storm during respiratory viral infection is an indicator of disease severity and poor prognosis. Type 1 interferon (IFN-I) production and signaling has been reported to be causal in cytokine storm-associated pathology in several respiratory viral infections, however, the mechanisms by which IFN-I promotes disease pathogenesis remain poorly understood. Here, using Usp18-deficient, USP18 enzymatic-inactive and Isg15-deficient mouse models, we report that lack of deISGylation during persistent viral infection leads to severe immune pathology characterized by hematological disruptions, cytokine amplification, lung vascular leakage and death. This pathology requires T cells but not T cell-intrinsic deletion of Usp18. However, lack of Usp18 in myeloid cells mimicked the pathological manifestations observed in Usp18-/- or Usp18C61A mice which were dependent on Isg15. We further mechanistically demonstrate that interrupting the ISGylation/deISGylation circuit increases extracellular levels of ISG15 which is accompanied by inflammatory neutrophil accumulation to the lung. Importantly, neutrophil depletion reversed morbidity and mortality in Usp18C61A mice. In summary, we reveal that the enzymatic function of Usp18 is crucial for regulating extracellular release of ISG15. This is accompanied by altered neutrophil differentiation, cytokine amplification and mortality following persistent viral infection. Moreover, our results suggest that extracellular ISG15 may drive the inflammatory pathology observed and could be both a prospective predictor of disease outcome and a therapeutic target during severe respiratory viral infections.
Acetylcholine is released in the basolateral amygdala in response to predictors of reward and enhances learning of cue-reward contingency

The basolateral amygdala (BLA) is critical for associating initially neutral cues with appetitive and aversive stimuli and receives dense neuromodulatory acetylcholine (ACh) projections. We measured BLA ACh signaling and principal neuron activity in mice during cue-reward learning using a fluorescent ACh sensor and calcium indicators. We found that ACh levels and activity of nucleus basalis of Meynert (NBM) cholinergic terminals in the BLA (NBM-BLA) increased sharply in response to reward-related events and shifted as mice learned the tone-reward contingency. BLA principal neuron activity followed reward retrieval and moved to the reward-predictive tone after task acquisition. Optical stimulation of cholinergic NBM-BLA terminal fibers during cue-reward learning led to more rapid learning of the cue-reward contingency. These results indicate that BLA ACh signaling carries important information about salient events in cue-reward learning and provides a framework for understanding how ACh signaling contributes to shaping BLA responses to emotional stimuli.
Electrical Automaticity and Intercellular Synchronization via Shared Extracellular Compartments

Electrically excitable cells often spontaneously and synchronously depolarize in vitro and in vivo. It remains unknown how cells synchronize and autorhythmically activate above the intrinsic mean activation frequency of isolated cells without pacemaking mechanisms. Recent insights into ephaptic coupling (non-gap junction or synaptic coupling) suggest that cyclic ion accumulation and depletion in diffusion limited extracellular volumes densely expressing ion channels modifies action potentials. This report explores how potassium accumulation and depletion in a restricted extracellular domain promotes spontaneous oscillations in the Hodgkin Huxley action potential model, which does not spontaneously activate on its own without external stimulus. Simulations demonstrate cells sharing a diffusion limited extracellular compartment can become autorhythmic and synchronous despite intercellular electrical heterogeneity. Autorhythmic frequency can be determined by net potassium flux into the cleft and the cleft volume. Additionally, inexcitable cells can induce autorhythmic activity in an excitable cell via a shared cleft and sufficient potassium fluxes contributed by each cell. Importantly, the synchronization and autorhythmic activity conferred by shared cleft with reduced potassium efflux can occur in the absence of gap junctions. Lastly, not only can potassium oscillations in shared restricted clefts initiate, support, and suppress autorhythmic depolarizations, the same mechanism can homogenize repolarization. The work has implications for understanding how automaticity is coordinated among excitable cells and suggests a new role for non-excitable cells such as fibroblasts, macrophages, or astrocytes with sarcolemmal potassium handling proteins facing shared and restricted intercellular clefts. SIGNIFICANCE A mechanism of cyclic ion accumulation and depletion in diffusion limited extracellular compartments can suppress, initiate, and support autorhythmic activity. Additionally, autorhythmicity can emerge from electrophysiologically heterogeneous cell pairs sharing a diffusion limited extracellular compartment, even if the individual cells will not spontaneously depolarize on their own. Sustained and synchronous autorhythmic activity can occur in the absence of gap junction coupling. Lastly, the shared diffusion limited extracellular compartment can also reduce action potential duration gradients by synchronizing repolarization.
Immune-challenged vampire bats produce fewer contact calls

Infections can affect how animals vocalize and, hence, interact with conspecifics. While this effect has been shown for mate-attraction calls, other vocalizations that facilitate social contact have received less attention. When isolated, vampire bats produce contact calls which attract highly associated groupmates. Here, we test the effect of an immune challenge on contact calling rates of individually isolated vampire bats. Sickness behavior did not appear to change call structure, but it decreased the number of contact calls produced. This effect could decrease contact with groupmates and augment other established mechanisms by which sickness reduces social encounters (e.g. mortality, lethargy, and social withdrawal or disinterest).
Single-nucleus RNA-seq and FISH reveal coordinated transcriptional activity in mammalian myofibers

Skeletal muscle fibers are large syncytia but it is currently unknown whether gene expression is coordinately regulated in their numerous nuclei. By snRNA-seq and snATAC-seq, we showed that slow, fast, myotendinous and neuromuscular junction myonuclei each have different transcriptional programs, associated with distinct chromatin states and combinations of transcription factors. In adult mice, identified myofiber types predominantly express either a slow or one of the three fast isoforms of Myosin heavy chain (MYH) proteins, while a small number of hybrid fibers can express more than one MYH. By snRNA-seq and FISH, we showed that the majority of myonuclei within a myofiber are synchronized, coordinately expressing only one fast Myh isoform with a preferential panel of muscle-specific genes. Importantly, this coordination of expression occurs early during post-natal development and depends on innervation. These findings highlight a unique mechanism of coordination of gene expression in a syncytium.
The influence of data type and functional traits on native bee phenology metrics: Opportunistic versus inventory records

Efforts to understand activity patterns of bees, our most important pollinators, often rely on opportunistically collected museum records to model temporal shifts or declines. This type of data, however, may not be suitable for this purpose given high spatiotemporal variability of native bee activity. By comparing phenological metrics calculated from intensive systematic inventory data with those from opportunistic museum records for bee species spanning a range of functional traits, we explored biases and limitations of data types to determine best practices for bee monitoring and assessment. We compiled half a million records of wild bee occurrence from opportunistic museum collections and six systematic inventory efforts, focusing analyses on 45 well-represented species that spanned five functional traits: sociality, nesting habits, floral specialization, voltinism, and body size. We then used permutation tests to evaluate differences between data types in estimating three phenology metrics: flight duration, number of annual abundance peaks, and date of the highest peak. We used GLMs to test for patterns of data type significance across traits. All 45 species differed significantly in the value of at least one phenology metric depending on the data type used. The date of the highest abundance peak differed for 40 species, flight duration for 34 species, and the number of peaks for 15 species. The number of peaks was more likely to differ between data types for larger bees, and flight duration was more likely to differ for larger bees and specialist bees. Our results reveal a strong influence of data type on phenology metrics that necessitates consideration of data source when evaluating changes in phenological activity, possibly applicable to many taxa. Accurately assessing phenological change may require expanding wild bee monitoring and data sharing.
Generalized gametic relationships for flexible analyses of parent-of-origin effects

Genomic imprinting causes alleles to influence the phenotype in a parent-of-origin-specific manner. In attempts to determine the effects of imprinted loci, gametic relationship matrices have widely been used in pedigree-based parent-of-origin analyses of population data. One drawback of this is the size of these matrices because they represent each individual by two gametic effects. Significantly fewer equations are needed if a previously published reduced imprinting model is used that relates observations from progeny without its own offspring to the transmitting abilities of their parents. This can be accomplished using a numerator relationship matrix, with only a single row and column per parent and ancestors. However, the reduced model is not applicable when the parents have records. To better handle the curse of dimensionality, we propose a combination of average gametic effects (transmitting abilities) for individuals without their own records and single gametic effects for others. The generalized gametic relationship matrix is the covariance of this mixture of genetic effects that allows for a significant reduction in the number of equations in gametic models depending on the trait, depth of pedigree, and population structure. It can also render the reduced model much more flexible by including observations from parents. Rules for setting-up its inverse from a pedigree are derived and implemented on an open-source program. The application of the same principles to phased marker data leads to a genomic version of the generalized gametic relationships. The implementation of generalized gametic models to the ASReml package is illustrated through worked examples.