The Mitochondrial RNA Granule Modulates Manganese-Dependent Cell Toxicity Prolonged manganese exposure causes manganism, a neurodegenerative movement disorder. The identity of adaptive and non-adaptive cellular processes targeted by manganese remains mostly unexplored. Here we study mechanisms engaged by manganese in genetic cellular models known to increase susceptibility to manganese exposure, the plasma membrane manganese efflux transporter SLC30A10 and the mitochondrial Parkinsons gene PARK2. We found that SLC30A10 and PARK2 mutations as well as manganese exposure compromised the mitochondrial RNA granule as well as mitochondrial transcript processing. These RNA granule defects led to impaired assembly and function of the mitochondrial respiratory chain. Notably, cells that survived a cytotoxic manganese challenge had impaired RNA granule function, thus suggesting that this granule phenotype was adaptive. CRISPR gene editing of subunits of the mitochondrial RNA granule, FASTKD2 or DHX30, as well as pharmacological inhibition of mitochondrial transcription-translation, were protective rather than deleterious for survival of cells acutely exposed to manganese. Similarly, adult Drosophila mutants with defects in the mitochondrial RNA granule component scully were safeguarded from manganese-induced mortality. We conclude that the downregulation of the mitochondrial RNA granule function is a protective mechanism for acute metal toxicity.

Significance StatementMutations in the manganese efflux transporter SLC30A10 and the mitochondrial Parkinsons gene PARK2, cause neurodegeneration and increased susceptibility to toxic manganese exposure. Thus, molecular processes affected in both mutants could offer insight into fundamental mechanisms conferring susceptibility or resilience to environmental and genetic factors associated with neurodegeneration. Here we report that SLC30A10 and PARK2 mutations compromise an understudied structure, the mitochondrial RNA granule, which is required for processing polycistronic mitochondrial RNAs. Cells and Drosophila lacking mitochondrial RNA granule components were resistant to manganese exposure. We conclude that the downregulation of the mitochondrial RNA granule function is an adaptive mechanism for cells exposed to manganese.
The cell growth cycle in budding yeast: testing hypotheses. Metabolic oscillation percolate throughout cellular physiology. The transcriptome oscillates between gene cohorts encoding for biosynthesis and growth, and for catabolism and stress response. However, long protein half-lives dampen effects on protein abundance, and the function of periodic transcription remains elusive. To test three prevailing hypotheses, we performed RNA-seq analysis during a dynamic state of the system, just preceding a bifurcation of system dynamics to a longer period. A few enzymes with short protein half-live and high transcript abundance amplitude could play a direct role in metabolic dynamics (hypothesis H1), e.g., carbonic anhydrase, sulfate uptake and the glyoxylate cycle. Substantial changes in the relative duration of expression of the antagonistic cohorts precede the bifurcation, supporting the idea of a function in cellular resource allocation (H2). The pulse-width modulation model, a mathematical formulation of this idea, explains the dependence of the cycle period on the culture growth rate. This pulse-like model of cell growth provides a first theoretical framework, where the phenomenon is understood as a mechanism of cellular resource allocation and protein homeostasis. The universal temporal program of transcription encodes the spatial structure of the cell, and we further suggest a function in pattern formation of the growing cell (H3).
Protein N-glycosylation is essential for SARS-CoV-2 infection SARS-CoV-2 extensively N-glycosylates its spike proteins, which are necessary for host cell invasion and the target of both vaccines and immunotherapies. These sugars are predicted to help mediate spike binding to the host receptor by stabilizing its  open conformation and evading host immunity. Here, we investigated both the essentiality of the host N-glycosylation pathway and SARS-CoV-2 N-glycans for infection. Inhibition of host N-glycosylation using RNAi or FDA-approved drugs reduced virus infectivity, including that of several variants. Under these conditions, cells produced less virions and some completely lost their infectivity. Furthermore, partial deglycosylation of intact virions showed that surface-exposed N-glycans are critical for cell invasion. Altogether, spike N-glycosylation is a targetable pathway with clinical potential for treatment or prevention of COVID-19. An epigenomic shift in amygdala marks the transition to maternal behaviors in alloparenting virgin female mice In many species, adults care for young offspring that are not their own, a phenomenon called alloparenting. However, most nonparental adults must be sensitized by repeated or extended exposures to newborns before they will robustly display parental-like behaviors. To capture neurogenomic events underlying the transition to active parental caring behaviors, we analyzed brain gene expression and chromatin profiles of virgin female mice co-housed with mothers during pregnancy and after birth. After an initial display of antagonistic behaviors and a surge of defense-related gene expression, we observed a dramatic shift in the chromatin landscape specifically in amygdala of the pup-exposed virgin females, accompanied by a dampening of anxiety-related gene expression. This epigenetic shift coincided with hypothalamic expression of the oxytocin gene and the emergence of behaviors and gene expression patterns classically associated with maternal care. The results outline a neurogenomic program associated with dramatic behavioral changes and suggest molecular networks relevant to human postpartum mental health. Predictive coding during action observation - a depth-resolved intersubject functional correlation study at 7T While the brain regions involved in action observation are relatively well documented in humans and primates, how these regions communicate to help understand and predict actions remains poorly understood. Traditional views emphasized a feed-forward architecture in which visual features are organized into increasingly complex representations that feed onto motor programs in parietal and then premotor cortices where the matching of observed actions upon the observers own motor programs contributes to action understanding. Predictive coding models place less emphasis on feed-forward connections and propose that feed-back connections from premotor regions back to parietal and visual neurons represent predictions about upcoming actions that can supersede visual inputs when actions become predictable, with visual input then merely representing prediction errors. Here we leverage the notion that feed-back connections target specific cortical layers to help adjudicate across these views. Specifically, we test whether observing sequences of hand actions in their natural order, which permits participants to predict upcoming actions, triggers more feed-back input to parietal regions than seeing the same actions in a scrambled sequence that hinders making predictions. Using submillimeter fMRI acquisition at 7T, we find that watching predictable sequences triggers more action-related activity (as measured using intersubject functional correlation) in the parietal cortical area PFt at depths receiving feed-back connections (layers III and V/VI) than watching the exact same actions in scrambled and hence unpredictable sequence. In addition, functional connectivity analysis performed using intersubject functional connectivity confirms that these increased action-related signals in PFt could originate from ventral premotor region BA44. This data showcases the utility of intersubject functional correlation in combination with 7T MRI to explore the architecture of social cognition under more naturalistic conditions, and provides evidence for models that emphasize the importance of feed-back connections in action prediction. High-throughput analysis of single human cells reveals the complex nature of DNA replication timing control DNA replication initiates from replication origins firing throughout S phase. Debate remains about whether origins are a fixed set of loci, or a loose agglomeration of potential sites used stochastically in individual cells, and about how consistent their firing time is. We developed an approach to profile DNA replication from whole-genome sequencing of thousands of single cells, which includes "in silico flow cytometry", a method for discriminating replicating and non-replicating cells. Using two microfluidic platforms, we analyzed up to 2,437 replicating cells from a single sample. The resolution and scale of the data allow focused analysis of replication initiation sites, demonstrating that most occur in confined genomic regions. While initiation order is remarkably similar across cells, we unexpectedly identify several subtypes of initiation regions in late-replicating regions. Taken together, high throughput, high resolution sequencing of individual cells reveals previously underappreciated variability in replication initiation and progression. Notch signaling via Hey1 and Id2b regulates Muller glia's regenerative response to retinal injury Zebrafish Muller glia (MG) respond to retinal injury by suppressing Notch signaling and producing progenitors for retinal repair. A certain threshold of injury-derived signal must be exceeded in order to engage MG in a regenerative response (MGs injury-response threshold). Pan-retinal Notch inhibition expands the zone of injury-responsive MG at the site of focal injury, suggesting that Notch signaling regulates MGs injury-response threshold. We found that Notch signaling enhanced chromatin accessibility and gene expression at a subset of regeneration-associated genes in the uninjured retina. Two Notch effector genes, hey1 and id2b, were identified that reflect bifurcation of the Notch signaling pathway, and differentially regulate MGs injury-response threshold and proliferation of MG-derived progenitors. Furthermore, Notch signaling component gene repression in the injured retina suggests a role for Dll4, Dlb, and Notch3 in regulating Notch signaling in MG and epistasis experiments confirm that the Dll4/Dlb-Notch3-Hey1/Id2b signaling pathway regulates MGs injury-response threshold and proliferation. A bispecific monomeric nanobody induces spike trimer dimers and neutralizes SARS-CoV-2 in vivo Antibodies binding to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike have therapeutic promise, but emerging variants show the potential for virus escape. This emphasizes the need for therapeutic molecules with distinct and novel neutralization mechanisms. Here we isolated a nanobody that interacts simultaneously with two RBDs from different spike trimers of SARS-CoV-2, rapidly inducing the formation of spike trimer-dimers leading to the loss of their ability to attach to the host cell receptor, ACE2. We show that this nanobody potently neutralizes SARS-CoV-2, including the B.1.351 variant, and cross-neutralizes SARS-CoV. Furthermore, we demonstrate the therapeutic potential of the nanobody against SARS-CoV-2 and the B.1.351 variant in a human ACE2 transgenic mouse model. This naturally elicited bispecific monomeric nanobody establishes a novel strategy for potent inactivation of viral antigens and represents a promising antiviral against emerging SARS-CoV-2 variants. The transmission of African Animal Trypanosomiasis (Nagana) in two districts in the forest zone of Ghana The African Animal Trypanosomiasis (nagana) is caused by several species of trypanosome species which results in significant clinical diseases. This consequently leads to huge economic losses. We carried out a cross-sectional survey to investigate the composition of vectors, parasite diversity and intensities in two districts in the Eastern region of Ghana. Cytochrome c oxidase subunit 1 (COI) and internal transcribed spacer (ITS1) dependent PCR assays identified tsetse fly species and trypanosome parasites circulating in the area. A total of 229 tsetse flies, 65 pigs and 20 were sampled. Female and male flies 155 (51.8%), 74 (32.3%) respectively were collected with biconical traps. The overall vector density of 4.3 flies/trap/day was observed. Typanosome prevalence of 58.9% (95% CI: 52.5-65.1), 46.2% (95% CI: 34.6-58.1) and 0.0% (95% CI: 0.0-16.1) in tsetse flies, pigs and cattle respectively were detected. Trypanosoma congolense was the predominant species with a prevalence of 80.7% (95% CI: 73.3-86.5) in flies and 60.0% (95% CI: 42.3-75.4). There was evidence of multiple trypanosome infection with T. congolense/T. simiae occurring highest with prevalence of 38.0% (95% CI: 30.7-46.9). The parasite prevalence in pigs across the communities was high with significant differences associated between locations (Chi2 = 28.06, 95% CI: 0.05-0.81, P=0.0009). Blood meal with flies revealed feeding on both domestic Sus scrofa domesticus (pig) and Phacochoerus africanus (warthog). Tsetse flies in this area remain highly infected with trypanosomes and continue to transmit the parasites to livestock and human populations in the communities. Accommodating site variation in neuroimaging data using normative and hierarchical Bayesian models AO_SCPLOWBSTRACTC_SCPLOWThe potential of normative modeling to make individualized predictions from neuroimaging data has enabled inferences that go beyond the case-control approach. However, site effects are often confounded with variables of interest in a complex manner and can bias estimates of normative models, which has impeded the application of normative models to large multi-site neuroimaging data sets. In this study, we suggest accommodating for these site effects by including them as random effects in a hierarchical Bayesian model. We compared the performance of a linear and a non-linear hierarchical Bayesian model in modeling the effect of age on cortical thickness. We used data of 570 healthy individuals from the ABIDE (autism brain imaging data exchange) data set in our experiments. In addition, we used data from individuals with autism to test whether our models are able to retain clinically useful information while removing site effects. We compared the proposed single stage hierarchical Bayesian method to several harmonization techniques commonly used to deal with additive and multiplicative site effects using a two stage regression, including regressing out site and harmonizing for site with ComBat, both with and without explicitly preserving variance related to age and sex as biological variation of interest. In addition, we made predictions from raw data, in which site has not been accommodated for. The proposed hierarchical Bayesian method showed the best predictive performance according to multiple metrics. Beyond that, the resulting z-scores showed little to no residual site effects, yet still retained clinically useful information. In contrast, performance was particularly poor for the regression model and the ComBat model in which age and sex were not explicitly modeled. In all two stage harmonization models, predictions were poorly scaled, suffering from a loss of more than 90 % of the original variance. Our results show the value of hierarchical Bayesian regression methods for accommodating site variation in neuroimaging data, which provides an alternative to harmonization techniques. While the approach we propose may have broad utility, our approach is particularly well suited to normative modelling where the primary interest is in accurate modelling of inter-subject variation and statistical quantification of deviations from a reference model.

1 HighlightsO_LIDevelopment and presentation of normative modeling approach based on hierarchical Bayesian modeling that can be applied to large multi-site neuroimaging data sets.
C_LIO_LIComparison of performance of Hierarchical Bayesian model including site as predictor to several common ways to harmonize for multi-site effects.
C_LIO_LIPresentation of normative modeling as site correction tool.
C_LI Second Heart Field-derived Cells Contribute to Angiotensin II-mediated Ascending Aortopathies BackgroundThe ascending aorta is a common location for aneurysm and dissection. This aortic region is populated by a mosaic of medial and adventitial cells that are embryonically derived from either the second heart field (SHF) or the cardiac neural crest. SHF-derived cells populate areas that coincide with the spatial specificity of thoracic aortopathies. The purpose of this study was to determine whether and how SHF-derived cells contribute to ascending aortopathies.

MethodsAscending aortic pathologies were examined in patients with sporadic thoracic aortopathies and angiotensin II (AngII)-infused mice. Ascending aortas without overt pathology from AngII-infused mice were subjected to mass spectrometry assisted proteomics, and molecular features of SHF-derived cells were determined by single cell transcriptomic analyses. Genetic deletion of either low-density lipoprotein receptor-related protein 1 (Lrp1) or transforming growth factor-{beta} receptor 2 (Tgfbr2) in SHF- derived cells was conducted to examine the impact of SHF-derived cells on vascular integrity.

ResultsPathologies in human ascending aortic aneurysmal tissues were predominant in outer medial layers and adventitia. This gradient was mimicked in mouse aortas following AngII infusion that was coincident with the distribution of SHF-derived cells. Proteomics indicated that brief AngII infusion, prior to overt pathology, evoked downregulation of SMC proteins and differential expression of extracellular matrix proteins, including several LRP1 ligands. LRP1 deletion in SHF-derived cells augmented AngII-induced ascending aortic aneurysm and rupture. Single cell transcriptomic analysis revealed that brief AngII infusion decreased Lrp1 and Tgfbr2 mRNA abundance in SHF-derived cells and induced a unique fibroblast population with low abundance of Tgfbr2 mRNA. SHF-specific Tgfbr2 deletion led to embryonic lethality at E12.5 with dilatation of the outflow tract and retroperitoneal hemorrhage. Integration of proteomic and single cell transcriptomics results identified plasminogen activator inhibitor 1 (PAI1) as the most increased protein in SHF-derived SMCs and fibroblasts during AngII infusion. Immunostaining revealed a transmural gradient of PAI1 in both ascending aortas of AngII-infused mice and human ascending aneurysmal aortas that mimicked the gradient of medial and adventitial pathologies.

ConclusionSHF-derived cells exert a critical role in maintaining vascular integrity through LRP1 and TGF-{beta} signaling associated with increases of aortic PAI1.



O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/930917v6_ufig1.gif" ALT="Figure 1">
View larger version (34K):
org.highwire.dtl.DTLVardef@185e3d9org.highwire.dtl.DTLVardef@131d702org.highwire.dtl.DTLVardef@11afa42org.highwire.dtl.DTLVardef@242786_HPS_FORMAT_FIGEXP  M_FIG C_FIG Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABS- SHF-derived SMCs and fibroblasts associate with AngII-induced aortic pathologies.
- AngII induces a distinct fibroblast sub-cluster that is less abundant for mRNAs related to major extracellular components and TGF{beta} ligands and receptors, but more abundant for proliferative genes.
- TGFBR2 deletion in SHF-derived cells are embryonic lethal with significant dilatation of the outflow tract in mice.
- SHF-specific deletion of LRP1 leads to aortic pathologies in mice, supporting the importance of SHF-derived cells in maintaining ascending aortic wall integrity.


What are the clinical implications?- Heterogeneity of the embryonic origins of SMCs and fibroblasts contributes to complex mechanisms of vasculopathy formation, which should be considered when investigating the pathogenesis of thoracic aortopathies. A network approach to genetic circuit designs As genetic circuits become more sophisticated, the size and complexity of data about their designs increases. This data captured goes beyond monolithic genetic sequences and towards circuit modularity and functional details, which are beneficial for analyzing circuit performance and establishing design automation techniques. However, the accessibility, visualisation and usability of design data (and metadata) have received relatively little attention to date. Here, we present a method to turn circuit designs into networks and showcase its potential to enhance the utility of design data. Since networks are dynamic structures, initial graphs can be interactively shaped into sub-networks of relevant information based on requirements such as abstraction, hierarchy and protein interactions. Additionally, several visual changes can be applied, such as colouring or clustering nodes based on types (e.g., genes or promoters), resulting in easier comprehension from a user perspective. This approach allows circuit designs to be coupled to other networks, such as metabolic pathways or implementation protocols captured in graph-like formats. Therefore, we advocate using networks to structure, access and improve synthetic biology information. Development of a novel PROTAC using the nucleic acid aptamer as a targeting ligand for tumor-selective degradation of nucleolin PROteolysis TArgeting Chimeras (PROTACs) induce targeted protein degradation by hijacking the intracellular ubiquitin proteasome system, thus emerging as a new strategy for drug development. However, most PROTACs generated lack cell-type selectivity and have poorly soluble in water. To address this drawback, we developed a novel PROTAC ZL216 using aptamer AS1411 as a targeting ligand of nucleolin to conjugate with a small molecule ligand of E3 ligase VHL, which shows high aqueous solubility and serum stability. Based on the differential expression of nucleolin on the cell surface, ZL216 could bind to and internalize into breast cancer cells, but not normal breast cells. Furthermore, we revealed that ZL216 promoted the formation of a nucleolin-ZL216-VHL ternary complex in breast cancer cells and potently induced nucleolin degradation in vitro and in vivo. As a result, ZL216 inhibited the proliferation and migration of breast cancer cells. These studies demonstrate that in addition to peptides and small molecule compounds, nuclei acid aptamers can also be used to generate PROTACs, which broadens the toolbox constructing PROTACs and provides a promising strategy for development of tumor-selective PROTACs. Topology and adenocarcinoma cell localization dataset on the labyrinthin biomarker. The discovery and characterization of tumor associated antigens is increasingly important to advance the field of immuno-oncology. Data on the, 1) topology, 2) amino acid (a.a.) homology analyses and 3) cell surface localization are presented as evidence in support of labyrinthin as a pan-adenocarcinoma marker. Multiple bioinformatics tools were used to collectively predict calcium binding domain(s), N-myristoylation sites, kinase II phosphorylation sites, and transmembrane domain(s) with predicted N- and C-termini orientations. Sequence homology comparisons were found and alignments made for labyrinthin (255 a.a.) vs. the intracellular aspartyl/asparaginyl beta-hydroxylase (ASPH; 758 a.a.) and the ASPH-gene related protein junctate (299 a.a.), which are both type II proteins. The epitopes for commonly used MCA 44-3A6 anti-labyrinthin and FB-50 anti-ASPH/junctate antibodies were present in all three proteins. The selective expression of labyrinthin was investigated by third party fluorescent activated cell sorter (FACS) analyses of MCA 44-3A6 binding to non-permeabilized adenocarcinoma and normal cell line models. Labyrinthin was detected on non-permeabilized A549 human lung adenocarcinoma cells, but not on normal WI-38 human lung fibroblasts nor primary cultures of normal human glandular-related cells. Microscopic images of immunofluorescent labelled MCA 44-3A6 binding to A549 cells at random cell cycle stages complement the FACS results by further showing that labyrinthin persisted on the cell surfaces along with some cell internalization within 20 minutes. The dataset presented herein is related to research article: "Labyrinthin: A distinct pan-adenocarcinoma diagnostic and immunotherapeutic tumor specific antigen" by Babich et al., Heliyon, 2022 [1].

Specifications Table

O_TBL View this table:
org.highwire.dtl.DTLVardef@19222bborg.highwire.dtl.DTLVardef@1264bcdorg.highwire.dtl.DTLVardef@13a8158org.highwire.dtl.DTLVardef@1dcf0org.highwire.dtl.DTLVardef@503a9f_HPS_FORMAT_FIGEXP  M_TBL C_TBL Value of the data O_LIBecause identification and characterization of neoantigens [3, 4, 5] are rare yet increasingly important for diagnostics and therapeutics, the identification of labyrinthin as a pan-adenocarcinoma target is significant for further study by clinicians and cancer researchers.
C_LIO_LIThe data provide a basis to determine to what extent labyrinthin is a distinguishing marker for adenocarcinomas vs. conventional methods.
C_LIO_LIThe results indicate that therapeutic strategies can be employed that take advantage of labyrinthins convenient presence on adenocarcinoma cell surfaces.
C_LIO_LIThe discovery of a calcium binding sequence in labyrinthin provides a basis to research if it is key to explaining the long-known phenomena of elevated calcium and irregular signaling in cancer cells [8, 9].
C_LIO_LIThe data reveal a need to explore distinctions between ASPH and labyrinthin, in particular because ASPH [10, 11] and labyrinthin-based treatments have already been FDA approved for clinical trials.
C_LI The pH Dependence of Niclosamide Solubility, Dissolution, and Morphology Motivates Potentially Universal Mucin-Penetrating Nasal and Throat Sprays for COVID19, its Contagious Variants, and Other Respiratory Viral Infections MotivationWith the coronavirus pandemic still raging, prophylactic nasal and early treatment throat sprays could help prevent infection and reduce viral load. Niclosamide has the potential to treat a broad range of viral infections if local bioavailability is optimized as mucin-penetrating solutions as opposed to more traditional microparticle-based sprays that cannot penetrate the mucin.

ExperimentalpH-dependence of supernatant concentrations and dissolution rates of niclosamide were measured in buffered solutions by Nanodrop-UV/Vis-spectroscopy for niclosamide from different suppliers, as precipitated material, and as cosolvates. Data was compared to predictions from Henderson Hasselbalch and precipitation pH models. Optimal microscopy was used to observe the morphologies of precipitated and converted niclosamide.

ResultsSupernatant-concentrations of niclosamide increased with increasing pH, from 1.77uM at pH 3.66 to 30uM at pH 8, and more rapidly from 90uM at pH8.5 to 300uM at pH 9.1, reaching 641uM at pH 9.5. Logarithmic rates for dissolution increased by [~]3x for pHs 8.62 to 9.44. However, when precipitated from supersaturated solution, niclosamide equilibrated to much lower final supernatant concentrations, reflective of more stable polymorphs at each pH that were also apparent for niclosamide from other suppliers and cosolvates.

ConclusionsGiven niclosamides activity against COVID19, its more contagious variants, and other respiratory viral infections, these niclosamide solutions, that put the virus in lockdown, could represent universal prophylactic nasal and early treatment throat sprays. As solutions they would be the simplest and potentially most effective formulations from both an efficacy standpoint as well as manufacturing and distribution, with no cold chain. They now just need testing. Microbial colonization of tannin-rich tropical plants: interplay between degradability, methane production and tannin disappearance in the rumen AO_SCPLOWBSTRACTC_SCPLOWCondensed tannins in plants are found free and attached to protein and fibre but it is not known whether these fractions influence rumen degradation and microbial colonization. This study explored the rumen degradation of tropical tannins-rich plants and the relationship between condensed tannins fractions disappearance and microbial communities colonising plant particles. Leaves from Calliandra calothyrsus, Gliricidia sepium, and Leucaena leucocephala, pods from Acacia nilotica and the leaves of two agricultural by-products: Manihot esculenta and Musa spp. were incubated in situ in the rumen of dairy cows. An in vitro approach was also used to assess the effects of these plants on rumen fermentation parameters. All plants contained more than 100 g/kg of condensed tannins with a large proportion (32 to 61%) bound to proteins. Calliandra calothyrsus had the highest concentration of condensed tannins at 361 g/kg, whereas Acacia nilotica was particularly rich in hydrolysable tannins (350 g/kg). Hydrolysable and free condensed tannins from all plants completely disappeared after 24 h incubation in the rumen. Disappearance of protein-bound condensed tannins was variable with values ranging from 93% for Gliricidia sepium to 21% for Acacia nilotica. In contrast, fibre-bound condensed tannins disappearance averaged ~82% and did not vary between plants. Disappearance of bound fractions of condensed tannins was not associated with degradability of plant fractions. The presence of tannins interfered with the microbial colonisation of plants. Each plant had distinct bacterial and archaeal communities after 3 and 12 h of incubation in the rumen and distinct protozoal communities at 3 h. Adherent communities in tannin-rich plants had a lower relative abundance of fibrolytic microbes, notably Fibrobacter spp. Whereas, archaea diversity was reduced in high tannin-containing Calliandra calothyrsus and Acacia nilotica at 12 h of incubation. Concurrently, in vitro methane production was lower for Calliandra calothyrsus, Acacia nilotica and Leucaena leucocephala although for the latter total volatile fatty acids production was not affected and was similar to control. Here we show that the total amount of hydrolysable and condensed tannins contained in a plant govern the interaction with rumen microbes affecting degradability and fermentation. The effect of protein- and fibre-bound condensed tannins on degradability is less important. Generation of nonlinear and spatially-organized 3D cultures on a microfluidic chip using photoreactive thiol-ene and methacryloyl hydrogels Micropatterning techniques for 3D cell cultures enable the recreation of tissue-level structures, but their combination with well-defined, microscale fluidic systems for perfusion remains challenging. To address this technological gap, we developed a user-friendly in-situ micropatterning protocol that integrates photolithography of crosslinkable, cell-laden hydrogels with a simple microfluidic housing, and tested the impact of crosslinking chemistry on stability and spatial resolution. Working with gelatin functionalized with photo-crosslinkable moieties, we found that inclusion of cells at high densities ([&ge;] 107/mL) during crosslinking did not impede thiol-norbornene gelation, but decreased the storage moduli of methacryloyl hydrogels. Hydrogel composition and light dose were selected to match the storage moduli of soft tissues. The cell-laden precursor solution was flowed into a microfluidic chamber and exposed to 405 nm light through a photomask to generate the desired pattern. The on-chip 3D cultures were self-standing, and the designs were interchangeable by simply swapping out the photomask. Thiol-ene hydrogels yielded highly accurate feature sizes from 100 - 900 m in diameter, whereas methacryloyl hydrogels yielded slightly enlarged features. Furthermore, only thiol-ene hydrogels were mechanically stable under perfusion overnight. Repeated patterning readily generated multi-region cultures, either separately or adjacent, including non-linear boundaries that are challenging to obtain on-chip. As a proof-of-principle, primary human T cells, were patterned on-chip with high regional specificity. Viability remained high (> 85%) after overnight culture with constant perfusion. We envision that this technology will enable researchers to pattern 3D cultures under fluidic control in biomimetic geometries that were previously difficult to obtain. Mouse lemur, a new animal model to investigate cardiac pacemaker activity in primates. BackgroundGray mouse lemurs (Microcebus murinus) are one of the smallest and most ancestral primates known. Their size falls in between that of mice and rats, and their genetic proximity to humans makes them an emerging model to study age-related neurodegeneration. Since Microcebus murinus replicates similar senescence processes of humans, it constitutes a useful model also for studying cardiovascular dysfunctions. However, its cardiac physiology is unknown. Thus, we investigated sinoatrial node (SAN) pacemaker activity of mouse lemurs, presenting the first characterization of heart automaticity in nonhuman primates.

Methods and ResultsWe recorded the heart automaticity in freely moving mouse lemurs and in their SAN tissues and pacemaker myocytes. Mouse lemurs have a heart rate (HR) in between those of mice and rats and a similar generation of electrical impulses in the SAN tissue. Conversely, their ventricular depolarization is more similar to large mammals. Mouse lemur SAN myocytes express the main pacemaker currents at densities similar to mice: the hyperpolarization-activated current (If) and the L-type (ICa,L) and T-type (ICa,T) calcium currents. Moreover, mouse lemur SAN myocytes display spontaneous ryanodine receptor (RyR)-dependent Ca2+ transients. Using muscle-derived stem cells (MDSCs) from the Microcebus murinus skeletal muscle, we also differentiated pacemaker-like (PML) cells showing spontaneous automaticity and expressing markers of native SAN myocytes (HCN4 and connexin-45).

ConclusionsIn conclusion, our characterization of heart automaticity in Microcebus murinus provides new opportunities for comparative cardio-physiology studies in primates and with humans, and for pharmacologic testing of molecules that could assist in modulating age-related dysfunctions of automaticity. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. MODEL PREDICTIVE CONTROL OF CANCER CELLULAR DYNAMICS: A NEW STRATEGY FOR THERAPY DESIGN Recent advancements in Cybergenetics have led to the development of new computational and experimental platforms that enable to robustly steer cellular dynamics by applying external feedback control. Such technologies have never been applied to regulate intracellular dynamics of cancer cells. Here, we show in silico that adaptive model predictive control (MPC) can effectively be used to steer signalling dynamics in Non-Small Cell Lung Cancer (NSCLC) cells to resemble those of wild-type cells, and to support the design of combination therapies. Our optimisation-based control algorithm enables tailoring the cost function to force the controller to alternate different drugs and/or reduce drug exposure, minimising both drug-induced toxicity and resistance to treatment. Our results pave the way for new cybergenetics experiments in cancer cells for drug combination therapy design. Citation needed? Wikipedia and the COVID-19 pandemic BackgroundWith the COVID-19 pandemics outbreak, millions flocked to Wikipedia for updated information. Amid growing concerns regarding an "infodemic", ensuring the quality of information is a crucial vector of public health. Investigating if and how Wikipedia remained up to date and in line with science is key to formulating strategies to counter misinformation. Using citation analyses, we asked: which sources informed Wikipedias COVID-19-related articles before and during the pandemics first wave (January-May 2020).

ResultsWe found that coronavirus-related articles referenced trusted media sources and high-quality academic research. Moreover, despite a surge in COVID-19 preprints, Wikipedia had a clear preference for open-access studies published in respected journals and made little use of preprints. Building a timeline of English COVID-19 articles from 2001-2020 revealed a nuanced trade-off between quality and timeliness. It further showed how preexisting articles on key topics related to the virus created a framework for integrating new knowledge. Supported by a rigid sourcing policy, this "scientific infrastructure" facilitated contextualization and regulated the influx of new information. Lastly, we constructed a network of DOI-Wikipedia articles, which showed the shifting landscape of pandemic-related knowledge on Wikipedia and how academic citations create a web of shared knowledge supporting topics like COVID-19 vaccine development.

ConclusionsUnderstanding how scientific research interacts with the digital knowledge-sphere during the pandemic provides insight into how Wikipedia can facilitate access to science. It also reveals how, aided by what we term its "citizen encyclopedists", it successfully fended off COVID-19 disinformation and how this unique model may be deployed in other contexts. ConVarT: a search engine for orthologous variants and functional inference of human genetic variants The availability of genetic variants, together with phenotypic annotations from model organisms, facilitates comparing these variants with equivalent variants in humans. However, existing databases and search tools do not make it easy to scan for equivalent variants, namely "matching variants" (MatchVars) between humans and other organisms. Therefore, we developed an integrated search engine called ConVarT (http://www.convart.org/) for matching variants between humans, mice, and C. elegans. ConVarT incorporates annotations (including phenotypic and pathogenic) into variants, and these previously unexploited phenotypic MatchVars from mice and C. elegans can give clues about the functional consequence of human genetic variants. Our analysis shows that many phenotypic variants in different genes from mice and C. elegans, so far, have no counterparts in humans, and thus, can be useful resources when evaluating a relationship between a new human mutation and a disease. Targeted biallelic integration of an inducible Caspase 9 suicide gene for safer cellular therapies prevents development of drug-resistant escapees in human iPSCs Drug-inducible suicide systems may help to minimize risks of cellular therapies due to the tumor forming potential of human induced pluripotent stem cells (hiPSCs). Recent research challenged the usefulness of such systems since rare drug-resistant subclones were observed that showed elimination or silencing of the transgene.

We have introduced a drug-inducible Caspase9 suicide system (iCASP9) into the AAVS1 safe harbor locus of hiPSCs. In these cells, apoptosis could be efficiently induced in vitro. In mice, drug treatment generally led to rapid elimination of teratomas, but individual animals subsequently formed tumor tissue from monoallelic iCASP9 hiPSCs. Very rare drug-resistant subclones of monoallelic iCASP9 hiPSCs appeared in vitro with frequencies of ~ 3x10-8. Transgene elimination, presumably via Loss of Heterozygosity (LoH), or methylation of the CAG promoter but not methylation of the O_SCPLOWPPPC_SCPLOWO_SCPCAP1C_SCPCAPO_SCPLOWRC_SCPLOWO_SCPCAP12C_SCPCAPO_SCPLOWCC_SCPLOW locus were identified as underlying mechanisms. In contrast, we never observed any escapees from biallelic iCASP9 hiPSCs, even after treatment of up to 0.8 billion hiPSCs.

In conclusion, biallelic integration of an iCASP9 system in the AAVS1 locus may substantially contribute to the safety level of iPSC-based therapies, which should be calculated by relating clonal escapee frequencies to the cell number in tumors of a size that is readily detectable during routine screening procedures. Modeling Adsorption, Conformation, and Orientation of the Fis1 Tail Anchor at the Mitochondrial Outer Membrane Proteins can be targeted to organellar membranes using a tail anchor (TA), a stretch of hydrophobic amino acids found at the polypeptide carboxyl-terminus. The Fis1 protein (Fis1p), which promotes mitochondrial and peroxisomal division in the yeast Saccharomyces cerevisiae, is targeted to those organelles by its TA. Substantial evidence suggests that Fis1p insertion into the mitochondrial outer membrane can occur without the need for a translocation machinery. However, recent findings raise the possibility that Fis1p insertion into mitochondria might be promoted by a proteinaceous complex. Here, we have performed atomistic and coarse-grained simulations to analyze the adsorption, conformation and orientation of the Fis1(TA). Our results support stable insertion at the mitochondrial outer membrane in a monotopic, rather than a bitopic (transmembrane), configuration. Once inserted in the monotopic orientation, unassisted transition to the bitopic orientation is expected to be blocked by the highly charged nature of the TA carboxyl-terminus and by the Fis1p cytosolic domain. Our results are consistent with a model in which Fis1p does not require a translocation machinery for insertion at mitochondria. Two Types of Cinnamoyl-CoA Reductase Function Divergently in Tissue Lignification, Phenylpropanoids Flux Control, and Inter-pathway Cross-talk with Glucosinolates as Revealed in Brassica napus Cinnamoyl-CoA reductase (CCR) is the entry point of lignin pathway and a crucial locus in dissection and manipulation of associated traits, but its functional dissection in Brassicaceae plants is largely lagged behind though Arabidopsis thaliana CCR1 has been characterized to certain extent. Here, 16 CCR genes are identified from Brassica napus and its parental species B. rapa and B. oleracea. Brassicaceae CCR genes are divided into CCR1 subfamily and CCR2 subfamily with divergent organ-specificity, yellow-seed trait participation and stresses responsiveness. CCR1 is preferential in G- and H-lignins biosynthesis and vascular development, while CCR2 has a deviation to S-lignin biosynthesis and interfascicular fiber development. CCR1 has stronger effects on lignification-related development, lodging resistance, phenylpropanoid flux control and seed coat pigmentation, whereas CCR2 controls sinapates levels. CCR1 upregulation could delay bolting and flowering time, while CCR2 upregulation weakens vascular system in leaf due to suppressed G lignin accumulation. Besides, CCR1 and CCR2 are deeply but almost oppositely linked with glucosinolates metabolism through inter-pathway crosstalk. Strangely, upregulation of both CCR1 and CCR2 could not enhance resistance to UV-B and S. sclerotiorum though CCR2 is sharply induced by them. These results provide systemic dissection on Brassica CCRs and CCR1-CCR2 divergence in Brassicaceae.

HighlightBrassicaceae contains two types of Cinnamoyl-CoA reductase. As revealed in Brassica napus, they are divergently involved in lignin monomer biosynthesis, tissue lignification, phenylpropanoid flux control, and inter-pathway crosstalk with glucosinolates. Evolution of thermal physiology alters the projected range of threespine stickleback under climate change Species distribution models (SDMs) are widely used to predict range shifts but could be unreliable under climate change scenarios because they do not account for evolution. The thermal physiology of a species is a key determinant of range and thus incorporating thermal trait evolution into SDMs might be expected to alter projected ranges. We identified a genetic basis for physiological and behavioural traits that evolve in response to temperature change in natural populations of threespine stickleback (Gasterosteus aculeatus). Using these data, we created geographic range projections using a mechanistic niche area approach under two climate change scenarios. Under both scenarios, trait data was either static ( no evolution models), allowed to evolve at observed evolutionary rates ( evolution models), or allowed to evolve at a rate of evolution scaled by the trait variance that is explained by quantitative trait loci (QTL;  scaled evolution models). We show that incorporating these traits and their evolution substantially altered the projected ranges for a widespread panmictic marine population, with over 7-fold increases in area under climate change projections when traits are allowed to evolve. Evolution-informed SDMs should improve the precision of forecasting range dynamics under climate change, and aid in their application to management and the protection of biodiversity.