title: Increased excursions to functional networks in schizophrenia in the absence of task

abstract: Brain activity during rest has been demonstrated to evolve through a repertoire of functional connectivity (FC) patterns, whose alterations may provide biomarkers of schizophrenia - a psychotic disorder characterized by dysfunctional brain connectivity. In this study, differences between the dynamic exploration of resting-state networks using functional magnetic resonance imaging (fMRI) data from 71 schizophrenia patients and 74 healthy controls were investigated using a method focusing on the dominant fMRI signal phase coherence pattern at each time point. Through the lens of dynamical systems theory, brain activity in the form of temporal FC state trajectories was examined for intergroup differences by calculating the fractional occupancy, dwell time, limiting probability of each state and the transition probabilities between states. Results showed reduced fractional occupancy of a globally synchronized state in schizophrenia. Conversely, FC states overlapping with canonical functional subsystems exhibited increased fractional occupancy and limiting probability in schizophrenia. Furthermore, state-to-state transition probabilities were altered in schizophrenia. This revealed a reduced probability of remaining in a global integrative state, increased probability of switching from this state to functionally meaningful networks and reduced probability of remaining in a state related to the Default Mode network. These results revealed medium to large effect sizes. Finally, this study showed that using K-medoids clustering did not influence the observed intergroup differences - highlighting the utility of dynamical systems theory to better understand brain activity. Combined, these findings expose pronounced differences between schizophrenia patients and healthy controls - supporting and extending current knowledge regarding disrupted brain dynamics in schizophrenia.

category: neuroscience


title: Reactivation of hedonic but not sensory representations in human emotional learning

abstract: Learning which stimuli in our environment co-occur with painful or pleasurable events is critical for survival. Previous research has established the basic neural and behavioural mechanisms of aversive and appetitive conditioning; however, it is unclear what precisely is learned. Here we examined what aspects of the unconditioned stimulus (US) - sensory and hedonic - are transferred to the conditioned stimulus (CS). To decode the content of brain activation patterns elicited during appetitive (soft touch) and aversive (painful touch) conditioning of faces, a novel variation of representational similarity analysis (RSA) based on theoretically driven representational patterns of interest (POIs) was applied to fMRI data. Once face associations were learned through conditioning, globally the CS reactivated US representational patterns showing conditioning-dependent reactivation. More specifically, in higher order brain regions, the CS only reactivated hedonic but not sensory aspects of the US - suggesting that affective conditioning primarily carries forward the valence of the experience rather than its sensory origins.


title: Bitter taste perception in BaYaka hunter-gatherers

abstract: Aversion towards bitter tastes evolved across vertebrate species to enable the recognition of harmful plant toxins. Genetic background, mode of subsistence, and dietary factors may explain variation in bitter taste sensitivity between human ecologies. We are the first to examine bitter taste perception within a population at different levels of market-integration, conducting an experiment using phenylthiocarbamide (PTC) and thiourea (Thiourea) infused paper strips amongst the Mbendjele BaYaka hunter-gatherers from Congo. We investigated (i.) prevalence of bitter tasting phenotypes amongst the BaYaka; (ii.) differences between BaYaka who grew up in town and forest camps. We found that 45.1% of BaYaka experience PTC as bitter, and 42.5% experience Thiourea as bitter. There were no sex differences, however bitter tasting phenotypes were significantly more prevalent in town than in forest camps (PTC:64.1%vs35.1%; Thiourea:64.1%vs31.1%). Despite a shared genetic background, we found that BaYaka who grew up in town were more sensitive to bitter taste than those living in the forest, suggesting a developmental component in taste perception. We suggest that the dietary transition from wild to cultivated foods in BaYaka living in town and a decreased use of traditional plant medicine may underlie this variation in bitter taste perception.

category: ecology


title: A microbial mutualist within host individuals increases parasite transmission between host individuals: Evidence from a field mesocosm experiment

abstract: The interactions among host-associated microbes and parasites can have clear consequences for disease susceptibility and progression within host individuals. Yet, empirical evidence for how these interactions impact parasite transmission between host individuals remains scarce. We address this scarcity by using a field mesocosm experiment to investigate the interaction between a systemic fungal endophyte, Epichloe coenophiala, and a fungal parasite, Rhizoctonia solani, in leaves of a grass host, tall fescue. Specifically, we investigated how this interaction impacted parasite transmission under field conditions in replicated experimental host populations. Epichloe-inoculated populations tended to have greater disease prevalence over time, though this difference had weak statistical support. More clearly, Epichloe-inoculated populations experienced higher peak parasite prevalences than Epichloe-free populations. Epichloe conferred a benefit in growth; Epichloe-inoculated populations had greater aboveground biomass than Epichloe-free populations. Using biomass as a proxy, host density was correlated with peak parasite prevalence, but Epichloe still increased peak parasite prevalence after controlling for the effect of biomass. Together, these results suggest that within-host microbial interactions can impact disease at the population level. Further, while Epichloe is clearly a mutualist of tall fescue, it may not be a defensive mutualist in relation to R. solani.


title: Genomic insights into the recent population history of Mapuche Native Americans

abstract: The last few years have witnessed an explosive generation of genomic data from ancient and modern Native American populations. These data shed light on key demographic shifts that occurred in geographically diverse territories of South America, such as the Andean highlands, Southern Patagonia and the Amazon basin. We used genomic data to study the recent population history of the Mapuche, who are the major Native population from the Southern Cone (Chile and Argentina). We found evidence of specific shared genetic ancestry between the Mapuche and ancient populations from Southern Patagonia, Central Chile and the Argentine Pampas. Despite previous evidence of cultural influence of Inca and Tiwanaku polities over the Mapuche, we did not find evidence of specific shared ancestry between them, nor with Amazonian groups. We estimated the effective population size dynamics of the Mapuche ancestral population during the last millennia, identifying a population bottle-neck around 1650 AD, coinciding with a period of Spaniards invasions into the territory inhabited by the Mapuche. Finally, we show that admixed Chileans underwent post-admixture adaptation in their Mapuche subancestry component in genes related with lipid metabolism, suggesting adaptation to scarce food availability.

category: genetics


title: Potent neutralizing anti-SARS-CoV-2 human antibodies cure infection with SARS-CoV-2 variants in hamster model

abstract: Treatment with neutralizing monoclonal antibodies (mAbs) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to COVID-19 management. Unfortunately, SARS-CoV-2 variants can escape several of these recently approved mAbs, highlighting the need for additional discovery and development. In a convalescent COVID-19 patient, we identified six mAbs, classified in four epitope groups, that potently neutralized SARS-CoV-2 Wuhan, alpha, beta, gamma and delta infection in vitro. In hamsters, mAbs 3E6 and 3B8 potently cured infection with SARS-CoV-2 Wuhan, beta and delta when administered post-viral infection at 5 mg/kg. Even at 0.2 mg/kg, 3B8 still reduced viral titers. Intramuscular delivery of DNA-encoded 3B8 resulted in in vivo mAb production of median serum levels up to 90 g/ml, and protected hamsters against delta infection. Overall, our data mark 3B8 as a promising candidate against COVID-19, and highlight advances in both the identification and gene-based delivery of potent human mAbs.

category: immunology


title: Metabolic strategies of sharing pioneer bacteria mediating fresh macroalgae breakdown

abstract: Macroalgae represent huge amounts of biomass worldwide, largely recycled by marine heterotrophic bacteria. We investigated the strategies of "pioneer" bacteria within the flavobacterial genus Zobellia to initiate the degradation of fresh brown macroalgae, which has received little attention compared to the degradation of isolated polysaccharides. Zobellia galactanivorans DsijT could use macroalgae as a sole carbon source and extensively degrade algal tissues without requiring physical contact, via the secretion of extracellular enzymes. This indicated a sharing behaviour, whereby pioneers release public goods that can fuel other bacteria. Comparisons of eight Zobellia strains, and strong transcriptomic shifts in Z. galactanivorans cells using fresh macroalgae vs. isolated polysaccharides, revealed potential overlooked traits of pioneer bacteria. Besides brown algal polysaccharide degradation, they notably include stress resistance proteins, type IX secretion system proteins and novel uncharacterized Polysaccharide Utilization Loci. Overall, this work highlights the relevance of studying fresh macroalga degradation to fully understand the niche, metabolism and evolution of pioneer degraders, as well as their cooperative interactions within microbial communities, as key players in macroalgal biomass turnover.

category: microbiology


title: Negative Binomial factor regression with application to microbiome data analysis

abstract: The human microbiome provides essential physiological functions and helps maintain host homeostasis via the formation of intricate ecological host-microbiome relationships. While it is well established that the lifestyle of the host, dietary preferences, demographic background, and health status can influence microbial community composition and dynamics, robust generalizable associations between specific host-associated factors and specific microbial taxa have remained largely elusive. Here, we propose factor regression models that allow the estimation of structured parsimonious associations between host-related features and amplicon-derived microbial taxa. To account for the overdispersed nature of the amplicon sequencing count data, we propose Negative Binomial reduced rank regression (NB-RRR) and Negative Binomial co-sparse factor regression (NB-FAR). While NB-RRR encodes the underlying dependency among the microbial abundances as outcomes and the host-associated features as predictors through a rank-constrained coefficient matrix, NB-FAR uses a sparse singular value decomposition of the coefficient matrix. The latter approach avoids the notoriously difficult joint parameter estimation by extracting sparse unit-rank components of the coefficient matrix sequentially. To solve the non-convex optimization problems associated with these factor regression models, we present a novel iterative block-wise majorization procedure. Extensive simulation studies and an application to the microbial abundance data from the American Gut Project demonstrate the efficacy of the proposed procedure. In the American Gut Project data, we identify key factors that strongly link dietary habits and host life style to specific microbial families.


title: Nrf1 is not a direct target gene of SREBP1, albeit both are integrated into the rapamycin-responsive regulatory network in human hepatoma cells

abstract: It is worth interrogating why no more experimental evidence confirming those findings, since being reported by Mannings group in 2014s Nature (doi: 10.1038/nature13492), has been provided in the hitherto known literature. A key issue arising from their work is of particular concern about whether the mTORC1 signaling to upregulation of Nrf1-targeted proteasomal expression profiles occurs directly by SREBP1. In this study, our experiment evidence revealed that Nrf1 is not a direct target of SREBP1, although both are involved in the rapamycin-responsive regulatory networks. Closely scrutinizing two distinct transcriptomic datasets unraveled no significant changes in transcriptional expression of Nrf1 and almost all proteasomal subunits in siSREBP1 or SREBP1-/- cells, when compared to equivalent controls. However, distinct upstream signaling to Nrf1 dislocation by p97 and its processing by DDI1/2, along with downstream proteasomal expression, may be monitored by mTOR signaling, to various certain extents, depending on distinct experimental settings in different types of cells. Our further evidence has been obtained from DDI1-/-(DDI2insC) cells, demonstrating that putative effects of mTOR on the rapamycin-responsive signaling to Nrf1 and proteasomes may also be executed partially through a DDI1/2-independent mechanism, albeit the detailed regulatory events remain to be determined.

category: molecular biology


title: The VINE complex is a VPS9-domain GEF-containing SNX-BAR coat involved in endosomal sorting

abstract: Membrane trafficking pathways perform important roles in establishing and maintaining the endolysosomal network. Retrograde protein sorting from the endosome is promoted by conserved SNX-BAR-containing coat complexes including retromer which enrich cargo at tubular microdomains and generate transport carriers. In metazoans, retromer cooperates with VARP, a conserved VPS9-domain GEF, to direct an endosomal recycling pathway. The function of the yeast VARP homolog Vrl1 has been overlooked due an inactivating mutation in commonly studied strains. Here, we demonstrate that Vrl1 has features of a SNX-BAR coat protein and forms an obligate complex with Vin1, the paralog of the retromer SNX-BAR protein Vps5. Unique features in the Vin1 N-terminus allow Vrl1 to distinguish it from Vps5, thereby forming what we have named the VINE complex. VINE occupies endosomal tubules and promotes the delivery of a conserved mannose 6-phosphate receptor-like protein to the vacuolar membrane. In addition to sorting functions, membrane recruitment by Vin1 is essential for Vrl1 GEF activity, suggesting that VINE is a multifunctional coat complex that regulates trafficking and signaling events at the endosome.

category: cell biology


title: Exploring the relationships between autozygosity, educational attainment, and cognitive ability in a contemporary, trans-ancestral American sample

abstract: Previous studies have found significant associations between estimated autozygosity - the proportion of an individuals genome contained in homozygous segments due to distant inbreeding - and multiple traits, including educational attainment (EA) and cognitive ability. In one study, estimated autozygosity showed a stronger association with parental EA than the subjects own EA. This was likely driven by parental EAs association with mobility: more educated parents tended to migrate further from their hometown, therefore choosing more genetically diverse partners. We examined the associations between estimated autozygosity, cognitive ability, and parental EA in a contemporary sub-sample of adolescents from the Adolescent Brain and Cognitive Development StudySM (ABCD Study(R)) (analytic N=6,504). We found a negative association between autozygosity and child cognitive ability consistent with previous studies, while the associations between autozygosity and parental EA were in the expected direction of effect (with greater levels of autozygosity being associated with lower EA) but the effect sizes were significantly weaker than those estimated in previous work. We also found a lower mean level of autozygosity in the ABCD sample compared to previous autozygosity studies, which may reflect overall decreasing levels of autozygosity over generations. Variation in migration and mobility patterns in the ABCD study compared to other studies may explain the pattern of associations between estimated autozygosity, EA, and cognitive ability in the current study.


title: Anticipating future SARS-CoV-2 variants of concern through ab initio quantum mechanical modeling

abstract: Evolved SARS-CoV-2 variants are currently challenging the efficacy of first-generation vaccines, largely through the emergence of spike protein mutants. Among these variants, Delta is presently the most concerning. We employ an ab initio quantum mechanical model based on Density Functional Theory to characterize the spike protein Receptor Binding Domain (RBD) interaction with host cells and gain mechanistic insight into SARS-CoV-2 evolution. The approach is illustrated via a detailed investigation of the role of the E484K RBD mutation, a signature mutation of the Beta and Gamma variants. The simulation is employed to: predict the depleting effect of the E484K mutation on binding the RBD with select antibodies; identify residue E484 as a weak link in the original interaction with the human receptor hACE2; and describe SARS-CoV-2 Wuhan strand binding to the bat Rhinolophus macrotis ACE2 as more optimized than the human counterpart. Finally, we predict the hACE2 binding efficacy of a hypothetical E484K mutation added to the Delta variant RBD, identifying a potential future variant of concern. Results can be generalized to other mutations, and provide useful information to complement existing experimental datasets of the interaction between randomly generated libraries of hACE2 and viral spike mutants. We argue that ab initio modeling is at the point of being aptly employed to inform and predict events pertinent to viral and general evolution.

category: biophysics


title: Salmonella Typhimurium infection drives NK cell loss and conversion to ILC1-like cells, and CIS inhibition enhances antibacterial immunity

abstract: Immunotherapy has revolutionized cancer therapy by reactivating tumor-resident cytotoxic lymphocytes. More recently, immunotherapy has emerged to restore immunity against infectious agents, including bacterial infections. Immunotherapy primarily targets inhibitory pathways in tumor-resident T cells, however interest in other effector populations, such as natural killer (NK) cells, is growing. We have previously discovered that NK cell metabolism, proliferation, and activation can be neutralized through the TGF-{beta} immunosuppressive pathway by inducing plasticity of NK cells and differentiation into ILC1-like subsets. NK cells are also regulated through cytokine-inducible SH2-containing protein (CIS), which is induced by IL-15 and is a potent intracellular checkpoint suppressing NK cell survival and function. Targeting these two distinct pathways to restore NK cell function has shown promise is cancer models, but their application in bacterial infection remains unknown. Here, we investigate whether enhancement of NK cell function can improve anti-bacterial immunity, using Salmonella Typhimurium as a model. We identified conversion of NK cells to ILC1-like for the first time in the context of bacterial infection, however TGF-{beta} signaling was curiously redundant in this plasticity. Future work should focus on identifying drivers of ILC1 plasticity and its functional implication in bacterial infection models. We further describe that CIS-deficient mice displayed enhanced pro-inflammatory function and dramatically enhanced anti-infection immunity. Inhibition of CIS may present as a viable therapeutic option to enhance immunity towards bacterial infection.


title: Fracture healing is delayed in the absence of gasdermin signaling

abstract: Amino-terminal fragments from proteolytically cleaved gasdermins (GSDMs) form plasma membrane pores that enable the secretion of interleukin-1{beta} (IL-1{beta}) and IL-18. Excessive GSDM- mediated pore formation can compromise the integrity of the plasma membrane thereby causing the lytic inflammatory cell death, pyroptosis. We found that GSDMD and GSDME were the only GSDMs that were readily expressed in bone microenvironment. Therefore, we tested the hypothesis that GSDMD and GSDME are implicated in fracture healing owing to their role in the obligatory inflammatory response following injury. We found that bone callus volume and biomechanical properties of injured bones were significantly reduced in mice lacking either GSDM compared with wild-type (WT) mice, indicating that fracture healing was compromised in mutant mice. However, compound loss of GSDMD and GSDME did not exacerbate the outcomes, suggesting shared actions of both GSDMs in fracture healing. Mechanistically, bone injury induced IL-1{beta} and IL-18 secretion in vivo, a response that was mimicked in vitro by bone debris and ATP, which function as inflammatory danger signals. Importantly, the secretion of these cytokines was attenuated in conditions of GSDMD deficiency. Finally, deletion of IL-1 receptor reproduced the phenotype of Gsdmd or Gsdme deficient mice, implying that inflammatory responses induced by the GSDM-IL-1 axis promote bone healing after fracture.


title: The exploration of Thermococcus barophilus lipidome reveals the widest variety of phosphoglycolipids in Thermococcales.

abstract: One of the most distinctive characteristics of Archaea is their unique lipids. While the general nature of archaeal lipids has been linked to their tolerance to extreme conditions, little is known about the diversity of lipidic structures Archaea are able to synthesize, which hinders the elucidation of the physicochemical properties of their cell membrane. In an effort to widen the known lipid repertoire of the piezophilic and hyperthermophilic model archaeon Thermococcus barophilus, we comprehensively characterized its intact polar lipid (IPL), core lipid (CL), and polar head group compositions using a combination of cutting-edge liquid chromatography and mass spectrometric ionization systems. We tentatively identified 82 different IPLs based on five distinct CLs and 10 polar head group derivatives of phosphatidylhexoses, including compounds reported here for the first time, e.g., di-N-acetylhexosamine phosphatidylhexose-bearing lipids. Despite having extended the knowledge on the lipidome, our results also indicate that the majority of T. barophilus lipids remain inaccessible to current analytical procedures and that improvements in lipid extraction and analysis are still required. This expanded yet incomplete lipidome nonetheless opens new avenues for understanding the physiology, physicochemical properties, and organization of the membrane in this archaeon as well as other Archaea.


title: Regulation and mechanistic basis of macrolide resistance by the ABC-F ATPase MsrD

abstract: Antibiotic resistance ABC-Fs (ARE ABC-Fs) are translation factors currently proliferating among human pathogens that provide resistance against clinically important ribosome-targeting antibiotics. Here, we combine genetic and structural approaches to determine the activity of the streptococcal ARE ABC-F protein MsrD on the ribosome and its regulation in response to macrolide exposure. We show that cladinose-containing macrolides lead to insertion of MsrDL leader peptide into a conserved crevice of the ribosomal exit tunnel, which remained thus far undocumented, concomitantly with 23S rRNA rearrangements that preclude proper accommodation of release factors and inhibits termination. The stalled ribosome obstructs formation of a Rho-independent terminator which prevents msrD transcriptional attenuation. This stalled ribosome is rescued by MsrD powered by its two functionally asymmetric ATPase sites, but not by MsrD mutants which do not provide antibiotic resistance, showing evidence of equivalence between MsrD function in antibiotic resistance and its action on this complex.


title: PTEN differentially regulates endocytosis, migration, and proliferation in the enteric protozoan parasite Entamoeba histolytica

abstract:
PTEN is a lipid phosphatase that is highly conserved and involved in a broad range of biological processes including cytoskeletal reorganization, endocytosis, signal transduction, and cell migration in all eukaryotes. Although regulation of phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P3] signaling via PTEN has been well established in model organisms and mammals, it remains elusive in the parasitic protist E. histolytica, which heavily relies on PtdIns phosphate(s)-dependent membrane traffic, migration, and phago- and trogocytosis for its pathogenesis. In this study, we characterized the major PTEN from E. histolytica, EhPTEN1, which shows the highest expression at the transcript level in the trophozoite stage among 6 possible PTENs, to understand the significance of PtdIns(3,4,5)P3 signaling in this parasite. Live imaging of GFP-EhPTEN1 expressing amebic trophozoites showed localization mainly in the cytosol with a higher concentration at pseudopods and the extending edge of the phago- and trogocytic cups. Furthermore, quantitative analysis of phago- and trogocytosis using a confocal image cytometer showed that overexpression of EhPTEN1 caused reduction in trogo- and phagocytosis while transcriptional gene silencing of EhPTEN1 gene caused opposite phenotypes. These data suggest that EhPTEN1 has an inhibitory role in these biological processes. Conversely, EhPTEN1 acts as a positive regulator for fluid-phase and receptor-mediated endocytosis in E. histolytica trophozoites. Moreover, we showed that EhPTEN1 was required for optimal growth and migration of this parasite. Finally, the phosphatase activity of EhPTEN1 towards PtdIns(3,4,5)P3 was demonstrated, suggesting that the biological roles of EhPTEN1 are likely linked to its catalytic function. Taken together, these results indicate that EhPTEN1 differentially regulates multiple cellular activities essential for proliferation and pathogenesis of the organism, via PtdIns(3,4,5)P3 signaling. Elucidation of biological roles of PTEN and PtdIns(3,4,5)P3 signaling at the molecular levels promotes our understanding of the pathogenesis of this parasite and potentially leads to the design of novel therapeutics against amebiasis.

Author summaryEntamoeba histolytica is an intestinal protozoan parasite that causes amoebic dysentery and liver abscesses in humans. It has been well understood how the amoebas ability to ingests and destroy human cells and invade tissues contributes to disease symptoms such as bloody diarrhea. The underlying mechanisms for such activities, called pathogenicity, include trafficking (transport) and secretion of cytolytic proteins, migration (ameboid movement), and ingestion and destruction of human cells, heavily rely on the signal transduction system via metabolism (synthesis and decomposition) of phosphoinositides (phosphatidylinositols containing 0-5 phosphates), and downstream regulation of cytoskeleton (dynamic network of interlinking protein filaments, such as actin, in the cytoplasm). In this study, we characterized one enzyme called EhPTEN1, which degrades and inactivate PtdIns(3,4,5)P3. We have shown that EhPTEN1 is involved in migration, internalization of soluble and solid materials (endocytosis, trogo-, and phagocytosis). EhPTEN1 apparently regulates cell migration, endocytosis, trogo-, phagocytosis, and proliferation in a complex fashion. Our findings help in the elucidation of the physiological significance of PTEN and cellular events regulated via phosphoinositides in this enteric parasite and other pathogenic parasites, and potentially lead to the development of new control measures against parasitic diseases.


title: Bet-hedging antimicrobial strategies in macrophage phagosome acidification drive the dynamics of Cryptococcus neoformans intracellular escape mechanisms

abstract: The fungus Cryptococcus neoformans is a major human pathogen with a remarkable intracellular survival strategy that includes exiting macrophages through non-lytic exocytosis (Vomocytosis) and transferring between macrophages (Dragotcytosis) by a mechanism that involves sequential events of non-lytic exocytosis and phagocytosis. Vomocytosis and Dragotcytosis are fungal driven processes, but their triggers are not understood. We hypothesized that the dynamics of Dragotcytosis could inherit the stochasticity of phagolysosome acidification and that Dragotcytosis was triggered by fungal cell stress. Consistent with this view, fungal cells involved in Dragotcytosis reside in phagolysosomes characterized by low pH and/or high oxidative stress. Using fluorescent microscopy, qPCR, live cell video microscopy, and fungal growth assays we found that the that mitigating pH or oxidative stress abrogated Dragotcytosis frequency, that ROS susceptible mutants of C. neoformans underwent Dragotcytosis more frequently. Dragotcytosis initiation was linked to phagolysosomal pH and oxidative stresses and correlated with the macrophage polarization state. Dragotcytosis manifested stochastic dynamics thus paralleling the dynamics of phagosomal acidification, which correlated with the inhospitality of phagolysosomes in differently polarized macrophages. Hence, randomness in phagosomal acidification randomly created a population of inhospitable phagosomes where fungal cell stress triggered stochastic C. neoformans non-lytic exocytosis dynamics to escape a non-permissive intracellular macrophage environment.


title: Ectosome effect on endothelial monolayers in hyperglycemic and normoglycemic conditions

abstract: Extracellular vesicles, namely those larger ones - Ectosomes (Ect), are thought to be important cell-to-cell communication medium. Ect are considered as a potential therapeutic for type-1 and type-2 diabetes mellitus. Ect can be internalized by endothelial cells and, owing to their cargo, they modulate targeted cell behavior. Under hyperglycemic conditions (HGC), endothelial cells changed their properties and became stiffer and less mobile which causes endothelial dysfunction and abnormalities in micro- and macrovascular systems. The aim of this study was to find whether Ect restore mobility and motility of macrovascular endothelial cells under HGC. Uptake of Ect, cell morphology, cytoskeleton organization and membrane stiffness (by atomic force microscopy) were analyzed after the exposure to isolated Ect. To find which cellular pathways were deregulated by HGC and whether Ect could potentially restore gene expression profile, transcriptome analysis was done. We observed that endothelial cells internalized more Ect under normoglycemic conditions (NGC) then HGC. Hyperglycemic cells (HG) were bigger and showed the stiffer surface with denser actin cytoskeleton in comparison to normoglycemic cells. Number of metabolic pathways was influenced under HGC, especially those related to intracellular transport, metabolism and cellular component organization and Ect did not restore HGC impaired cell signaling. Ectosomes cannot reverse this harmful effect of hyperglycemia in endothelial cells, which can have clinical implication in use Ect as therapeutic target in diabetes treatment.


title: A mechanical G2 checkpoint controls epithelial cell division through E-cadherin-mediated regulation of Wee1-Cdk1

abstract: Epithelial cell divisions must be tightly coordinated with cell loss to preserve epithelial integrity. However, it is not well understood how the rate of epithelial cell division adapts to changes in cell number, for instance during homeostatic turnover or upon wounding of epithelia. Here, we show epithelial cells sense local cell density through mechanosensitive E-cadherin adhesions to control G2/M cell cycle progression. We demonstrate that tensile forces on E-cadherin adhesions are reduced as local cell density increases, which prompts the accumulation of the G2 checkpoint kinase Wee1. This elevated abundance of Wee1 results in inhibitory phoshorylation of Cdk1, and thereby establishes a pool of cells that is temporarily halted in G2-phase. Importantly, these cells are readily triggered to divide upon epithelial wounding, due to the consequent increase in intercellular forces and resulting degradation of Wee1. Our data thus demonstrate that epithelial cell division is controlled by a mechanical G2 checkpoint, which is regulated by cell density-dependent intercellular forces sensed and transduced by E-cadherin adhesions.


title: Charting ESCRT function reveals distinct and non-compensatory roles in blood progenitor maintenance and lineage choice in Drosophila

abstract:
Tissue heterogeneity permits diverse biological outputs in response to systemic signals but requires context-dependent spatiotemporal regulation of a limited number of signaling circuits. In addition to their stereotypical roles of transport and cargo sorting, endocytic networks provide rapid, adaptable, and often reversible means of signaling. Aberrant function of the Endosomal Sorting Complex Required for Transport (ESCRT) components results in ubiquitinated cargo accumulation, uncontrolled signaling and neoplastic transformation. However, context-specific effects of ESCRT on developmental decisions are not resolved. By a comprehensive spatiotemporal profiling of ESCRT in Drosophila hematopoiesis in vivo, here we show that pleiotropic ESCRT components have distinct effects on blood progenitor maintenance, lineage choice and response to immune challenge. Of all 13 core ESCRT components tested, only Vps28 and Vp36 were required in all progenitors, whereas others maintained spatiotemporally defined progenitor subsets. ESCRT depletion also sensitized posterior progenitors that normally resist differentiation, to respond to immunogenic cues. Depletion of the critical Notch signaling regulator Vps25 did not promote progenitor differentiation at steady state but made younger progenitors highly sensitive to wasp infestation, resulting in robust lamellocyte differentiation. We identify key heterotypic roles for ESCRT in controlling Notch activation and thereby progenitor proliferation and differentiation. Further, we show that ESCRT ability to regulate Notch activation depends on progenitor age and position along the anterior-posterior axis. The phenotypic range and disparity in signaling upon depletion of components provides insight into how ESCRT may tailor developmental diversity. These mechanisms for subtle control of cell phenotype may be applicable in multiple contexts.

SignificanceThe Endosomal Sorting Complex Required for Transport (ESCRT) machinery sorts ubiquitinated cargo for degradation or recycling. Aberrant ESCRT function is associated with many blood disorders. We did a comprehensive functional analysis of all 13 core ESCRT components in maintenance and differentiation of Drosophila larval blood progenitors. We show that ESCRT have diverse and non-compensatory functions in blood progenitors. ESCRT depletion from progenitors affects ubiquitination status cell autonomously and independent of progenitor maintenance. ESCRT function is more critical to maintain older progenitors and to prevent Notch-dependent crystal cell differentiation. Further, ESCRT depletion sensitizes refractile younger progenitors for lamellocyte differentiation. Our in situ developmental map of ESCRT function reveals critical checkpoints for cell fate choice and new paradigms for generating progenitor heterogeneity.

category: developmental biology


title: Epistasis is not a strong constraint on the recurrent evolution of toxin-resistant Na+,K+-ATPases among tetrapods

abstract:
A growing body of theoretical and experimental evidence suggests that intramolecular epistasis is a major determinant of rates and patterns of protein evolution and imposes a substantial constraint on the evolution of novel protein functions. Here, we examine the role of intramolecular epistasis in the recurrent evolution of resistance to cardiotonic steroids (CTS) across tetrapods, which occurs via specific amino acid substitutions to the -subunit family of Na,K-ATPases (ATP1A). After identifying a series of recurrent substitutions at two key sites of ATP1A that are predicted to confer CTS resistance in diverse tetrapods, we then performed protein engineering experiments to test the functional consequences of introducing these substitutions onto divergent species backgrounds. In line with previous results, we find that substitutions at these sites can have substantial background-dependent effects on CTS resistance. Globally, however, these substitutions also have pleiotropic effects that are consistent with additive rather than background-dependent effects. Moreover, the magnitude of a substitutions effect on activity does not depend on the overall extent of ATP1A sequence divergence between species. Our results suggest that epistatic constraints on the evolution of CTS-resistant forms of Na,K-ATPase likely depend on a small number of sites, with little dependence on overall levels of protein divergence. We propose that dependence on a limited number sites may account for the observation of convergent CTS resistance substitutions observed among taxa with highly divergent Na,K-ATPases.

Significance StatementIndividual amino acid residues within a protein work in concert to produce a functionally coherent structure that must be maintained even as orthologous proteins in different species diverge over time. Given this dependence, we expect identical mutations to have more similar effects on protein function in more closely related species. We tested this hypothesis by performing protein-engineering experiments on ATP1A, an enzyme mediating target-site insensitivity to cardiotonic steroids (CTS) in diverse animals. These experiments reveal that the phenotypic effects of substitutions can sometimes be background-dependent, but also that the magnitude of these phenotypic effects does not correlate with overall levels of ATP1A sequence divergence. Our results suggest that epistatic constraints are determined by states at a small number of sites, potentially explaining the frequent convergent CTS resistance substitutions among Na,K-ATPases of highly divergent taxa.

category: evolutionary biology


title: Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence


title: Origins and breadth of pairwise epistasis in an α-helix of β-lactamase TEM-1

abstract: Epistasis affects genome evolution together with our ability to predict individual mutation effects. The mechanistic basis of epistasis remains, however, largely unknown. To quantify and better understand interactions between fitness-affecting mutations, we focus on a 11 amino-acid -helix of the protein {beta}-lactamase TEM-1, and build a comprehensive library of more than 15,000 double mutants. Analysis of the growth rates of these mutants shows pervasive epistasis, which can be largely explained by a non-linear two-state model, where inactivating, destabilizing, neutral, or stabilizing mutations additively contribute to the phenotype. Hence, most epistatic interactions can be predicted by a non-linear model informed by single-point mutational measurements only. Deviations from the two-state model are consistently found for few pairs of residues, in particular when they are in contact. This result, as well as single-point mutation parameters, can be quantitatively found back through direct-coupling-analysis-based statistical models inferred from homologous sequence data. Our results thus shed light on the existence and the origins of the multiple determinants of the epistatic landscape, even at the level of small structural components of a protein, and suggest that the corresponding constraints shape the entire {beta}-lactamase family.


title: Systems genetics uncovers microbe-lipid-host connections in the murine gut

abstract: The molecular bases of how host genetic variation impact gut microbiome remain largely unknown. Here, we used a genetically diverse mouse population and systems genetics strategies to identify interactions between molecular phenotypes, including microbial functions, intestinal transcripts and cecal lipids that influence microbe-host dynamics. Quantitative trait loci (QTL) analysis identified genomic regions associated with variations in bacterial taxa, bacterial functions, including motility, sporulation and lipopolysaccharide production, and levels of bacterial- and host-derived lipids. We found overlapping QTL for the abundance of Akkermansia muciniphila and cecal levels of ornithine lipids (OL). Follow-up studies revealed that A. muciniphila is a major source of these lipids in the gut, provided evidence that OL have immunomodulatory effects and identified intestinal transcripts co-regulated with these traits. Collectively, these results suggest that OL are key players in A. muciniphila-host interactions and support the role of host genetics as a determinant of responses to gut microbes.


title: Translational suppression via IFG-1/eIF4G confers resistance to stress-induced RNA alternative splicing in Caenorhabditis elegans

abstract: Splicing of pre-mRNA is an essential process for dividing cells and splicing defects have been linked to aging and various chronic diseases. Environmental stress has recently been shown to alter splicing fidelity and molecular mechanisms that protect against splicing disruption remains unclear. Using an in vivo RNA splicing reporter, we performed a genome-wide RNAi screen in Caenorhabditis elegans and found that protein translation suppression via silencing of the conserved initiation factor 4G (IFG-1/eIF4G) protects against cadmium-induced splicing disruption. Transcriptome analysis of an ifg-1 deficient mutant revealed an overall increase in splicing fidelity and resistance towards cadmium-induced alternative splicing compared to the wild-type. We found that the ifg-1 mutant up-regulates >80 RNA splicing regulatory genes that are controlled by the TGF-{beta} transcription factor SMA-2. The extended lifespan of the ifg-1 mutant is partially reduced upon sma-2 depletion and completely nullified when core spliceosome genes including snr-1, snr-2, and uaf-2 are knocked down. Together, these data describe a molecular mechanism that provides resistance towards stress-induced alternative splicing and demonstrate an essential role for RNA homeostasis in promoting longevity in a translation-compromised mutant.


title: Effect of gastrointestinal alterations mimicking elderly conditions on in vitro digestion of meat and soy proteins

abstract: We evaluated the digestion of meat (chicken, beef, and pork) and soy proteins under in vitro conditions mimicking gastrointestinal (GI) conditions of adults (control, C) and elderly with achlorhydria (EA). The changes in degree of hydrolysis (DH), SDS-PAGE profiles, peptide concentration, and proteomics profiles during the digestion process were investigated. Digestion under the EA condition markedly decreased the DH of all protein sources, especially for soy protein. SDS-PAGE profiling and proteomics showed that myofibrillar/sarcoplasmic protein from meat and glycinin/beta-conglycinin from soy were the proteins most affected by the different digestive conditions. Our results indicated that the difference in the digestibility of meat protein between EA and control conditions gradually narrowed from the gastric to the intestinal phase, while a pronounced difference between control and EA conditions was maintained also in the intestinal phase. This work provides new insights of value for future dietary recommendations for elderly individuals.

category: biochemistry


title: Orthoformimycin inhibits translation elongation by displacing the A-site tRNA and preventing peptide bond formation

abstract: The ribosome is a major target for antibiotics owing to its essential cellular role in protein synthesis. Structural analysis of ribosome-antibiotic complexes provides insight into the molecular basis for their inhibitory action and highlights possible avenues to improve their potential or overcome existing resistance mechanisms. Here we use X-ray crystallography and pre-steady state kinetics to detail the inhibitory mechanism of the antimicrobial on the large ribosomal subunit.


title: Modeling Alternate Conformations with Alphafold2 via Modification of the Multiple Sequence Alignment

abstract: The unprecedented performance of Deepminds Alphafold2 in predicting protein structure in CASP XIV and the creation of a database of structures for multiple proteomes is reshaping structural biology. Moreover, the availability of Alphafold2s architecture and code has stimulated a number of questions on how to harness the capabilities of this remarkable tool. A question of central importance is whether Alphafold2s architecture is amenable to predict the intrinsic conformational heterogeneity of proteins. A general approach presented here builds on a simple manipulation of the multiple sequence alignment, via in silico mutagenesis, and subsequent modeling by Alphafold2. The approach is based in the concept that the multiple sequence alignment encodes for the structural heterogeneity, thus its rational manipulation will enable Alphafold2 to sample alternate conformations and potentially structural alterations due to point mutations. This modeling pipeline is benchmarked against canonical examples of protein conformational flexibility and applied to interrogate the conformational landscape of membrane proteins. This work broadens the applicability of Alphafold2 by generating multiple protein conformations to be tested biologically, biochemically, biophysically, and for use in structure-based drug design.

category: bioinformatics


title: Mechanism of FtsZ assembly dynamics revealed by filament structures in different nucleotide states

abstract: Treadmilling protein filaments perform essential cellular functions by growing from one end while shrinking from the other, driven by nucleotide hydrolysis. Bacterial cell division relies on the primitive tubulin homolog FtsZ, a target for antibiotic discovery that assembles into single treadmilling filaments that hydrolyse GTP at an active site formed upon subunit association. We determined high-resolution filament structures of FtsZ from the pathogen Staphylococcus aureus in complex with different nucleotide analogues and cations, including mimetics of the ground and transition states of catalysis. Together with mutational and biochemical analyses, our structures reveal interactions made by the GTP {gamma}-phosphate and Mg2+ at the subunit interface, a K+ ion stabilizing loop T7 for co-catalysis, new roles of key residues at the active site and a nearby crosstalk area, and rearrangements of a dynamic water shell bridging adjacent subunits upon GTP hydrolysis. We propose a mechanistic model that integrates nucleotide hydrolysis signalling with assembly-associated conformational changes and filament treadmilling. Equivalent assembly mechanisms may apply to more complex tubulin and actin cytomotive filaments that share analogous features with FtsZ.


title: Asking the Wrong Questions About American Science Education: Insights from a Longitudinal Study of High School Biotechnology Lab Instruction

abstract: The discussion of American science education is often framed by the questions: Why do American precollege students do poorly on international science assessments and what we are doing wrong? Rather we need to ask: Why do so many international students come to US universities for science, what are we doing right in science, and how do we stay ahead in science education? Poor scores on international assessments belie the fact that the U.S. has the best science education system in the world. Our study of 6,200 high school teachers in 1998 and 2018 documented striking success in retooling classrooms for labbased instruction in biotechnology and provided a pre-COVID-19 snapshot of what is right with American biology education. However, it also highlights the need revitalize our precollege teaching resource with a renewed National Science Foundation commitment to in-service training.

category: scientific communication and education


title: OPTN limits herpes stromal keratitis severity and demyelination through negative regulation of IL-17 and hyperinflammatory T-cell response

abstract: Herpes stromal keratitis (HSK) is a result of the inflammatory sequelae following primary and recurrent Herpes simplex virus type-1 (HSV-1) infections. This pathology is known to be mediated by immunopathogenic T cell responses against viral antigens, however most individuals infected with HSV-1 never exhibit signs of this immunopathology. Recent studies have identified the host restriction factor, optineurin (OPTN), as an inhibitor of viral spread in the central nervous system, protecting hosts from viral encephalopathy. In an HSV-1 corneal infection mouse model on OPTN knockout mice, we assess the contribution of OPTN to ameliorating the clinical manifestations of HSK. We identify that OPTN protects the host from loss of ocular and whisker sensitivity and opacification of the cornea. scRNA-seq of the trigeminal ganglion (TG) reveals that transcription changes to the peripheral neurons and immune cell populations drive the expression of Il-17A in CD4 and CD8 T cells, as well as increased infiltration of T cells into the TG. This leads to demyelination and the observed HSK pathology.


title: Fast maturation of splenic dendritic cells upon TBI is associated with FLT3/FLT3L signaling

abstract: Systemic inflammatory consequences remain a significant burden after traumatic brain injury (TBI), with almost all organs affected. The spleen is connected with the brain by autonomic innervation and by soluble mediators, and the cross-talk between brain and spleen may be important to establish the systemic inflammatory response to TBI. Ethanol intoxication, the most common comorbidity of TBI, is posited to influence the peripheral inflammatory response either directly or through the brain-spleen cross-talk. Here we show that TBI causes a substantial change in transcription of genes associated with dendritic cells activation in the spleen, in particular a FLT3/FLT3L induction 3h after TBI, which was enhanced by EI. The FLT3L induction was associated with the phosphorylation of FLT3 receptor in CD11c+ dendritic cells, which enhanced the protein synthesis of a subset of mRNAs, as shown by the increase in pS6, peIF2A levels in dendritic cells. This corresponded to the upregulation of proteins associated with maturation process and immunostimulatory properties such MHC-II, LAMP1 and CD68, and of pro-inflammatory cytokines such as TNF. Notably, EI enhanced the maturation of dendritic cells. However, whereas TBI decreases expression of the adrenergic 2b receptors on dendritic cells, EI increased it, thus augmenting the chances of cross-talk regulation of immune function by the autonomic system. In conclusion, this data indicates that TBI induces a fast maturation of the immunomodulatory functions of dendritic cells which is associated by FLT3/FLT3L signaling and which is enhanced by EI prior to TBI.


title: Synaptotagmin-1 C2B domains cooperatively stabilize the fusion stalk via a master-servant mechanism

abstract: Synaptotagmin-1 is a low-affinity Ca2+ sensor that triggers synchronous vesicle fusion. It contains two similar C2 domains (C2A and C2B) that cooperate in membrane binding, being the C2B domain the main responsible for the membrane fusion process due to its polybasic patch KRLKKKKTTIKK (321-332). In this work, a master-servant mechanism between two identical C2B domains is shown to control the formation of the fusion stalk. Two regions in C2B are essential for the process, the well-known polybasic patch and a recently described pair of arginines (398,399). The master domain shows strong PIP2 interactions with its polybasic patch and its pair of arginines. At the same time, the servant analogously cooperates with the master to reduce the total work to form the fusion stalk. The strategic mutation (T328E,T329E) in both master and servant domains disrupts the cooperative mechanism, drastically increasing the free energy needed to induce the fusion stalk, however with negligible effects on the master domain interactions with PIP2. These data point to a difference in the behavior of the servant domain, which is unable to sustain its PIP2 interactions neither through its polybasic patch nor through its pair of arginines, in the end losing its ability to assist the master in the formation of the fusion stalk.


title: Several different sequences are implicated in bloodstream-form-specific gene expression in Trypanosoma brucei

abstract: The parasite Trypanosoma brucei grows as bloodstream forms in mammalian hosts, and as procyclic forms in tsetse flies. In trypanosomes, gene expression regulation depends heavily on post-transcriptional mechanisms. Both the RNA-binding protein RBP10 and glycosomal phosphoglycerate kinase PGKC are expressed only in mammalian-infective forms. RBP10 targets procyclic-specific mRNAs for destruction, while PGKC is required for bloodstream-form glycolysis. Developmental regulation of both is essential: expression of either RBP10 or PGKC in procyclic forms inhibits their proliferation. We show that the 3'-untranslated region of the RBP10 mRNA is extraordinarily long - 7.3kb - and were able to identify six different sequences, scattered across the untranslated region, which can independently cause bloodstream-form-specific expression. The 3'-untranslated region of the PGKC mRNA, although much shorter, still contains two different regions, of 125 and 153nt, that independently gave developmental regulation. No short consensus sequences were identified that were enriched either within these regulatory regions, or when compared with other mRNAs with similar regulation, suggesting that more than one regulatory RNA-binding protein is important for repression of mRNAs in procyclic forms. We also identified regions, including an AT repeat, that increased expression in bloodstream forms, or suppressed it in both forms. Trypanosome mRNAs that encode RNA-binding proteins often have extremely extended 3'-untranslated regions. We suggest that one function of this might be to act as a fail-safe mechanism to ensure correct regulation even if mRNA processing or expression of trans regulators is defective.


title: Ubiquitination drives COPI priming and Golgi SNARE localization

abstract: Deciphering mechanisms controlling SNARE localization within the Golgi complex is crucial to understanding protein trafficking patterns within the secretory pathway. SNAREs are also thought to prime COPI assembly to ensure incorporation of these essential cargoes into vesicles but the regulation of these events is poorly understood. Here we report a roles for ubiquitin recognition by COPI in SNARE trafficking and in stabilizing interactions between Arf, COPI, and Golgi SNAREs. The ability of COPI to bind ubiquitin through its N-terminal WD repeat domain of {beta}COP or through an unrelated ubiquitin-binding domain (UBD) is essential for the proper localization of Golgi SNAREs Bet1 and Gos1. We find that COPI, the ArfGAP Glo3 and multiple Golgi SNAREs are ubiquitinated. Notably, the binding of Arf and COPI to Gos1 is markedly enhanced by ubiquitination of these components. Glo3 is thought to prime COPI-SNARE interactions; however, Glo3 is not enriched in the ubiquitin-stabilized SNARE-Arf-COPI complex but is instead enriched with COPI complexes that lack SNAREs. These results support a new model for how posttranslational modifications drive COPI priming events crucial for Golgi SNARE localization.


title: Asymmetrical dose-response shape the evolutionary trade-off between antifungal resistance and nutrient use

abstract: Antimicrobial resistance is an emerging threat for public health. The success of resistance mutations depends on the trade-off between the benefits and costs they incur. This trade-off is largely unknown and uncharacterized for antifungals. Here, we systematically measure the effect of all amino acid substitutions in the yeast cytosine deaminase Fcy1, the target of the antifungal 5-FC (flucytosine). We identify over 900 missense mutations granting resistance to 5-FC, a large fraction of which appear to act through destabilisation of the protein. The relationship between 5-FC resistance and growth sustained by cytosine deamination is characterized by a sharp trade-off, such that small gains in resistance universally lead to large losses in canonical enzyme function. We show that this steep relationship can be explained by differences in the dose-response functions of 5-FC and cytosine. Finally, we observe the same trade-off shape for the ortholog of FCY1 in Cryptoccocus neoformans, a human pathogen. Our results provide a powerful resource and platform for interpreting drug target variants in fungal pathogens as well as unprecedented insights into resistance-function trade-offs.


title: Asymmetrical dose-responses shape the evolutionary trade-off between antifungal resistance and nutrient use


title: An amino residue that guides the correct photoassembly the water-oxidation complex but not required for high affinity Mn2+ binding

abstract: The assembly of the Mn4O5Ca cluster of the photosystem II (PSII) starts from the initial binding and photooxidation of the first Mn2+ at a high affinity site (HAS). Recent cryo-EM apo-PSII structures reveal an altered geometry of amino ligands in this region and suggest the involvement of D1-Glu189 ligand in the formation of the HAS. We now find that Gln and Lys substitution mutants photoactivate with reduced quantum efficiency compared to the wild-type. However, the affinity of Mn2+ at the HAS in D1-E189K was very similar to the wild-type (~2.2 M). Thus, we conclude that D1-E189 does not form the HAS (~2.9 M) and that the reduced quantum efficiency of photoactivation in D1-E189K cannot be ascribed to the initial photooxidation of Mn2+ at the HAS. Besides reduced quantum efficiency, the D1-E189K mutant exhibits a large fraction of centers that fail to recover activity during photoactivation starting early in the assembly phase, becoming recalcitrant to further assembly. Fluorescence relaxation kinetics indicate on the presence of an alternative route for the charge recombination in Mn-depleted samples in all studied mutants and exclude damage to the photochemical reaction center as the cause for the recalcitrant centers failing to assemble and show that dark incubation of cells reverses some of the inactivation. This reversibility would explain the ability of these mutants to accumulate a significant fraction of active PSII during extended periods of cell growth. The failed recovery in the fraction of inactive centers appears to a reversible mis-assembly involving the accumulation of photooxidized, but non-catalytic high valence Mn at the donor side of photosystem II, and that a reductive mechanism exists for restoration of assembly capacity at sites incurring mis-assembly. Given the established role of Ca2+ in preventing misassembled Mn, we conclude that D1-E189K mutant impairs the ligation of Ca2+ at its effector site in all PSII centers that consequently leads to the mis-assembly resulting in accumulation of non-catalytic Mn at the donor side of PSII. Our data indicate that D1-E189 is not functionally involved in Mn2+ oxidation\binding at the HAS but rather involved in Ca2+ ligation and steps following the initial Mn2+ photooxidation.


title: An amino residue that guides the correct photoassembly the water-oxidation complex but is not required for high affinity Mn2+ binding


title: Habitat Suitability modeling of Urial (Ovis orientalis arkal) in the Samelghan plain by using Maximum Entropy Method

abstract: Habitat suitability models are useful tools for a variety of wildlife management objectives. Distributions of wildlife species can be predicted for geographical areas that have not been extensively surveyed. The basis of these models work is to minimize the relationship between species distribution and biotic and abiotic environments. For some species, there is information about presence and absence that allows the use of a variety of standard statistical methods, however, the absence data is not available for most species. Nowadays, the methods that need presence-only data are expanded. One of these methods is the Maximum Entropy (MaxEnt) modeling. The purpose of this study is to model the habitat of Urial (Ovis orientalis arkal) in the Samelghan plain in the North East of Iran with the MaxEnt method. This algorithm uses the Jackknife plot and percent contribution values to determine the significance of the variables. The results showed that variables such as southern aspects, Juniperus-Acer, Artemisia-Perennial plants, slope 0-5%, and asphalt road were the most important factors affecting the species habitat selection. The area under curve (AUC) Receiver Operating Characteristic (ROC) showed an excellent model performance. Suitable habitat was classified based on the threshold value (0.0513) and the ROC, which based on the results 28% of the area was a suitable habitat for Urial.


title: Similarities Between Bacterial GAD and Human GAD65:Implications in Gut Mediated Autoimmune Type 1 Diabetes

abstract: A variety of islet autoantibodies (AAbs) can predict and possibly dictate eventual type 1 diabetes (T1D) diagnosis. Upwards of 75% of those with T1D are positive for AAbs against glutamic acid decarboxylase (GAD65), a producer of gamma-aminobutyric acid (GABA) in human pancreatic beta cells. Interestingly, bacterial populations within the human gut also express GAD65 and produce GABA. Evidence suggests that dysbiosis of the microbiome may correlate with T1D pathogenesis and physiology. Therefore, autoimmune linkages between the gut microbiome and islets susceptible to autoimmune attack need to be further elucidated. Utilizing silico analyses, we show here that 25 GAD sequences from different human gut bacterial sources show sequence and motif similarities to human beta cell GAD65. Our motif analyses determined that a majority of gut GAD sequences contain the pyroxical dependent decarboxylase domain of human GAD65 which is important for its enzymatic activity. Additionally, we showed overlap with known human GAD65 T-cell receptor epitopes which may implicate the immune destruction of beta cells. Thus, we propose a physiological hypothesis in which changes in the gut microbiome in those with T1D result in a release of bacterial GAD, thus causing miseducation of the host immune system. Due to the notable similarities, we found between humans and bacterial GAD, these deputized immune cells may then go on to target human beta cells leading to the development of T1D.

category: genomics


title: Similarities Between Bacterial GAD and Human GAD65: Implications in Gut Mediated Autoimmune Type 1 Diabetes


title: Design of novel specific primers for helicobacter detection with diagnostic utility evaluation of different 16srRNA genus-specific primers: A comparative study

abstract: Molecular diagnosis of helicobacters by PCR is simpler, more accurate, and feasible compared to other diagnostic methods. Validity and accuracy are highly dependent on the PCR primer design, diffusion time, and mutation rate of helicobacters. This study aimed to design 16srRNA -specific primers for Helicobacter spp. and H. pylori. Application of comparative statistical analysis of the diagnostic utility of the most available 16srRNA genus-specific primers. The new primers were designed using bioinformatics tools (MAFFT MSA and Gblocks command line). A comparative study was applied on nine genus-specific 16srRNA primers in comparison to the ConsH using Insilco and laboratory evaluation. The results demonstrated that the best specificity and sensitivity of the primers designed for this study compared to other primers. The comparative study revealed that the heminested outer/inner primers were the worst. Although H276,16srRNA(a), HeliS/Heli-nest, and Hcom had acceptable diagnostic utility, false positive and false negative results were obtained. Specificity testing on clinical samples indicated a surprising result; that H. pylori was not the sole enemy that we were looking for, but the NPH should be considered as a real risk prognostic for gastric diseases, consequently, a specific diagnosis and treatment should be developed. This study concluded that our designed primers were the most specific and sensitive in comparison with other primers. in addition, Insilco evaluation is not accurate enough for primer assessment and that the laboratory evaluation is mandatory.


title: Dexamethasone induces senescence of lung epithelial cells and augments TGF-β1-mediated production of the fibrosis mediator serpin E1 (plasminogen activator inhibitor-1)

abstract:
BackgroundIdiopathic pulmonary fibrosis (IPF) is a progressive, incurable scarring disease of the lungs with a prognosis worse than most cancers. Pathologically, IPF is characterised by upregulation of the pro-fibrotic cytokine transforming growth factor-{beta}1 (TGF-{beta}1), activation of fibroblasts, and deposition of collagen in the alveolar interstitium. Recent evidence has highlighted the role of senescent type 2 alveolar epithelial cells in the pathogenesis of IPF. In a clinical trial, a treatment regimen containing a corticosteroid drug accelerated pulmonary fibrosis leading to more hospitalizations and deaths, particularly in patients with telomere shortening which drives cellular senescence.

AimTo investigate the potential pro-fibrotic actions of corticosteroids on lung epithelial cells in vitro, including effects on cellular senescence and interactions with TGF-{beta}1.

MethodsThe synthetic glucocorticoid dexamethasone (DEX) was incubated with A549 and BEAS-2B human lung epithelial cells in the presence or absence of TGF-{beta}1. Cellular senescence was assessed by morphology, senescence-associated beta-galactosidase (SA {beta}-Gal) expression, and qPCR for transcription of senescence-associated molecular markers. Conditioned media were screened for growth factors and cytokines and cultured with human lung fibroblasts. An IPF lung tissue RNA array dataset was re-analysed with a focus on senescence markers.

ResultsDEX induced senescence in lung epithelial cells associated with increased p21 (CDKN1A) expression independently of p16 (CDKN2A) or p53 (TP53). DEX amplified upregulation of the pro-fibrotic mediator serpin E1/plasminogen activator inhibitor-1 (PAI-1) in the presence of TGF-{beta}1. The senescence-associated secretory phenotype from lung epithelial cells treated with DEX plus TGF-{beta}1-treated contained increased concentrations of GM-CSF and IL-6 and when incubated with primary human lung fibroblasts there were trends to increased senescence and production of fibrosis markers. Upregulation of senescence markers was demonstrated by analysis of an IPF transcriptomic dataset.

DiscussionDEX induces senescence in lung epithelial cell lines in vitro and interacts with TGF-{beta}1 to amplify production of the pro-fibrotic mediator serpin E1 (PAI-1). This may be a mechanism by which corticosteroids promote pulmonary fibrosis in susceptible individuals. Serpin E1/PAI-1 is a potential druggable target in pulmonary fibrosis.


title: The non-random structure of multi-trophic ecological interactions maximizes species coexistence within ecologically realistic constraints

abstract: Theory posits that the persistence of species in ecological communities is shaped by their interactions within and across trophic levels. However, we lack empirical evaluations of how the structure, strength and sign of these interactions drive the potential to coexist in diverse multi-trophic communities. Here we model community feasibility domains, a theoretically-informed measure of coexistence probability, from empirical data on communities comprising more than 50 species for three trophic guilds (plants, pollinators, and herbivores). Although feasibility domains vary depending on the number of trophic guilds considered, we show that higher network connectance leads to lower coexistence opportunities. Moreover, empirical estimations of the feasibility domains were higher with respect to random network structures but lower than a mean-field approach, suggesting that observed interaction structures tend to maximize coexistence within its imposed limits. Our results stress the importance of incorporating empirically-informed interaction structures within and across guilds to better understand how species coexist in diverse multi-trophic communities.


title: The interaction between neural populations: Additive versus diffusive coupling

abstract:
Models of networks of populations of neurons commonly assume that the interactions between neural populations are via additive or diffusive coupling. When using the additive coupling, a populations activity is affected by the sum of the activities of neighbouring populations. In contrast, when using the diffusive coupling a neural population is affected by the sum of the differences between its activity and the activity of its neighbours. These two coupling functions have been used interchangeably for similar applications. Here, we show that the choice of coupling can lead to strikingly different brain network dynamics. We focus on a model of seizure transitions that has been used both with additive and diffusive coupling in the literature. We consider networks with two and three nodes, and large random and scale-free networks with 64 nodes. We further assess functional networks inferred from magnetoencephalography (MEG) from people with epilepsy and healthy controls. To characterize the seizure dynamics on these networks, we use the escape time, the brain network ictogenicity (BNI) and the node ictogenicity (NI), which are measures of the networks global and local ability to generate seizures. Our main result is that the level of ictogenicity of a network is strongly dependent on the coupling function. We find that people with epilepsy have higher additive BNI than controls, as hypothesized, while the diffusive BNI provides the opposite result. Moreover, individual nodes that are more likely to drive seizures with one type of coupling are more likely to prevent seizures with the other coupling function. Our results on the MEG networks and evidence from the literature suggest that the additive coupling may be a better modelling choice than the diffusive coupling, at least for BNI and NI studies. Thus, we highlight the need to motivate and validate the choice of coupling in future studies.

Author summaryMost models of brain dynamics assume that distinct brain regions interact in either an additive or a diffusive way. With additive coupling, each brain region sums incoming signals. In contrast, with diffusive coupling, each region sums the differences between its own signal and incoming signals. Although they are different, these two couplings have been used for very similar applications, particularly within models of epilepsy. Here we assessed the effect of this choice on seizure behaviour. Using a model of seizures and both artificial and real brain networks, we showed that the coupling choice can lead to very different seizure dynamics. We found that networks that are more prone to seizures using one coupling, are less prone to them using the other. Likewise, individual brain regions that are more likely to drive seizures when using additive coupling, are more likely to prevent them when using diffusive coupling. Using real brain networks, we found that the additive coupling predicted higher seizure propensity in people with epilepsy compared to healthy controls, whereas the diffusive coupling did not. Our results highlight the need to justify the choice of coupling used and show that the additive coupling may be a better option in some applications.


title: Highly accurate retinotopic maps of the physiological blind spot in human visual cortex

abstract: The physiological blind spot is a naturally occurring scotoma corresponding with the optic disc in the retina of each eye. Even during monocular viewing, observers are usually oblivious to the scotoma, in part because the visual system extrapolates information from the surrounding area. Unfortunately, studying this visual field region with neuroimaging has proven difficult, as it occupies only a small part of retinotopic cortex. Here we used functional magnetic resonance imaging and a novel data-driven method for mapping the retinotopic organization in and around the blind spot representation in V1. Our approach allowed for highly accurate reconstructions of the extent of an observers blind spot, and out-performed conventional model-based analyses. This method opens exciting opportunities to study the plasticity of receptive fields after visual field loss, and our data add to evidence suggesting that the neural circuitry responsible for impressions of perceptual completion across the physiological blind spot most likely involves regions of extrastriate cortex - beyond V1.


title: Structured connectivity in the cerebellum enables noise-resilient pattern separation

abstract: The cerebellum is thought to detect and correct errors between intended and executed commands1-3 and is critical for social behaviors, cognition and emotion4-6. Computations for motor control must be performed quickly to correct errors in real time and should be sensitive to small differences between patterns for fine error correction while being resilient to noise7. Influential theories of cerebellar information processing have largely assumed random network connectivity, which increases the encoding capacity of the networks first layer8-13. However, maximizing encoding capacity reduces resiliency to noise7. To understand how neuronal circuits address this fundamental tradeoff, we mapped the feedforward connectivity in the mouse cerebellar cortex using automated large-scale transmission electron microscopy (EM) and convolutional neural network-based image segmentation. We found that both the input and output layers of the circuit exhibit redundant and selective connectivity motifs, which contrast with prevailing models. Numerical simulations suggest these redundant, non-random connectivity motifs increase discriminability of similar input patterns at a minimal cost to the networks overall encoding capacity. This work reveals how neuronal network structure can balance encoding capacity and redundancy, unveiling principles of biological network architecture with implications for artificial neural network design.


title: Multi-task representations in human cortex transform along a sensory-to-motor hierarchy

abstract: Human cognition recruits diverse neural processes, yet the organizing computational and functional architectures remain unclear. Here, we characterized the geometry and topography of multi-task representations across human cortex using functional MRI during 26 cognitive tasks in the same subjects. We measured the representational similarity across tasks within a region, and the alignment of representations between regions. We found a cortical topography of representational alignment following a hierarchical sensory-association-motor gradient, revealing compression-then-expansion of multi-task dimensionality along this gradient. To investigate computational principles of multi-task representations, we trained multi-layer neural network models to transform empirical visual to motor representations. Compression-then-expansion organization in models emerged exclusively in a training regime where internal representations are highly optimized for sensory-to-motor transformation, and not under generic signal propagation. This regime produces hierarchically structured representations similar to empirical cortical patterns. Together, these results reveal computational principles that organize multi-task representations across human cortex to support flexible cognition.


title: Using child-friendly movie stimuli to study the development of face, place, and object regions from age 3 to 12 years

abstract: Scanning young children while watching short, engaging, commercially-produced movies has emerged as a promising approach for increasing data retention and quality. Movie stimuli also evoke a richer variety of cognitive processes than traditional experiments - allowing the study of multiple aspects of brain development simultaneously. However, because these stimuli are uncontrolled, it is unclear how effectively distinct profiles of brain activity can be distinguished from the resulting data. Here we develop an approach for identifying multiple distinct subject-specific Regions of Interest (ssROIs) using fMRI data collected during movie-viewing. We focused on the test case of higher-level visual regions selective for faces, scenes, and objects. Adults (N=13) were scanned while viewing a 5.5 minute child-friendly movie, as well as a traditional experiment with isolated faces, scenes, and objects. We found that just 2.7 minutes of movie data could identify subject-specific face, scene, and object regions. While successful, the movie approach was still less effective than a traditional localizer. Having validated our approach in adults, we then used the same methods on movie data collected from 3-12-year-old children (N=122). Movie response timecourses in 3-year-old childrens face, scene, and object regions were already significantly and specifically predicted by timecourses from the corresponding regions in adults. We also found evidence of continued developmental change, particularly in the face-selective posterior superior temporal sulcus. Taken together, our results reveal both early maturity and functional change in face, scene, and object regions, and more broadly highlight the promise of short, child-friendly movies for developmental cognitive neuroscience.


title: General factors of white matter microstructure from DTI and NODDI in the developing brain

abstract:
Preterm birth is closely associated with diffuse white matter dysmaturation inferred from diffusion MRI and neurocognitive impairment in childhood. Diffusion tensor imaging (DTI) and neurite orientation dispersion and density imaging (NODDI) are distinct dMRI modalities, yet metrics derived from these two methods share variance across tracts. This raises the hypothesis that dimensionality reduction approaches may provide efficient whole-brain estimates of white matter microstructure that capture (dys)maturational processes. To investigate the optimal model for accurate classification of generalised white matter dysmaturation in preterm infants we assessed variation in DTI and NODDI metrics across 16 major white matter tracts using principal component analysis and structural equation modelling, in 79 term and 141 preterm infants at term equivalent age. We used logistic regression models to evaluate performances of single-metric and multimodality general factor frameworks for efficient classification of preterm infants based on variation in white matter microstructure. Single-metric general factors from DTI and NODDI capture substantial shared variance (41.8-72.5%) across 16 white matter tracts, and two multimodality factors captured 93.9% of variance shared between DTI and NODDI metrics themselves. General factors associate with preterm birth and a single model that includes all seven DTI and NODDI metrics provides the most accurate prediction of microstructural variations associated with preterm birth. This suggests that despite global covariance of dMRI metrics in neonates, each metric represents information about specific (and additive) aspects of the underlying microstructure that differ in preterm compared to term subjects.

HighlightsO_LIWe measured variation of 7 DTI and NODDI metrics across 16 major tracts
C_LIO_LIGeneral factors for DTI and NODDI capture substantial shared variance across tracts
C_LIO_LIGeneral factors also capture substantial shared variance between DTI and NODDI
C_LIO_LISingle-metric and multimodality factors associate with gestational age at birth
C_LIO_LIThe best preterm prediction model contains all 7 single-metric g-factors
C_LI


title: Non-uniform distribution of dendritic nonlinearities differentially engages thalamostriatal and corticostriatal inputs onto cholinergic interneurons

abstract: The tonic activity of striatal cholinergic interneurons (CINs) is modified differentially by their afferent inputs. Although their unitary synaptic currents are identical, cortical inputs onto distal dendrites only weakly entrain CINs, whereas proximal thalamic inputs trigger abrupt pauses in discharge in response to salient external stimuli. To test whether the dendritic expression of the active conductances that drive autonomous discharge contribute to the CINs capacity to dissociate cortical from thalamic inputs, we used an optogenetics-based method to quantify dendritic excitability. We found that the persistent sodium (NaP) current gave rise to dendritic boosting and that the hyperpolarization-activated cyclic nucleotide-gated (HCN) current gave rise to a subhertz membrane resonance. This resonance may underlie our novel finding of an association between CIN pauses and internally-generated slow wave events in sleeping non-human primates. Moreover, our method indicated that dendritic NaP and HCN currents were preferentially expressed in proximal dendrites. We validated this non-uniform distribution with two-photon imaging of dendritic back-propagating action potentials, and by demonstrating boosting of thalamic, but not cortical, inputs by NaP currents. Thus, the localization of active dendritic conductances in CIN dendrites mirrors the spatial distribution of afferent terminals and may promote their differential responses to thalamic vs. cortical inputs.


title: A positive feedback loop mediates crosstalk between calcium, cyclic nucleotide and lipid signalling in Toxoplasma gondii

abstract: Fundamental processes of obligate intracellular parasites, such as Toxoplasma gondii and Plasmodium falciparum, are controlled by a set of plant-like calcium dependent kinases (CDPKs), the conserved cAMP- and cGMP-dependent protein kinases (PKA and PKG), secondary messengers and lipid signalling. While some major components of the signalling networks have been identified, how these are connected remains largely unknown. Here, we compare the phospho-signalling networks during Toxoplasma egress from its host cell by artificially raising cGMP or calcium levels to activate PKG or CDPKs, respectively. We show that both these inducers trigger near identical signalling pathways and provide evidence for a positive feedback loop involving CDPK3. We measure phospho- and lipid signalling in parasites treated with the Ca2+ ionophore A23187 in a sub-minute timecourse and show CDPK3-dependent regulation of diacylglycerol levels and increased phosphorylation of four phosphodiesterases (PDEs), suggesting their function in the feedback loop. Disruption of CDPK3 leads to elevated cAMP levels and inhibition of PKA signalling rescues the egress defect of {Delta}CDPK3 parasites treated with A23187. Biochemical analysis of the four PDEs identifies PDE2 as the only cAMP-specific PDE among these candidates, while the other PDEs are cGMP specific, two of which are inhibited by the predicted PDE inhibitor BIPPO. Conditional deletion of the four PDEs supports an important, but non-essential role for PDE1 and PDE2 in growth, with PDE2 controlling A23187-mediated egress. In summary we uncover a positive feedback loop that enhances signalling during egress and links several signalling pathways together.


title: Distinctive nuclear zone for RAD51-mediated homologous recombinational DNA repair

abstract: Genome-based functions are inseparable from the dynamic higher-order architecture of the cell nucleus. In this context, the repair of DNA damage is coordinated by precise spatiotemporal controls that target and regulate the repair machinery required to maintain genome integrity. However, the mechanisms that pair damaged DNA with intact template for repair by homologous recombination (HR) without illegitimate recombination remain unclear. This report highlights the intimate relationship between nuclear architecture and HR in mammalian cells. RAD51, the key recombinase of HR, forms spherical foci in S/G2 phases spontaneously. Using super-resolution microscopy, we show that following induction of DNA double-strand breaks RAD51 foci at damaged sites elongate to bridge between intact and damaged sister chromatids; this assembly occurs within bundle-shaped distinctive nuclear zones, requires interactions of RAD51 with various factors, and precedes ATP-dependent events involved the recombination of intact and damaged DNA. We observed a time-dependent transfer of single-stranded DNA overhangs, generated during HR, into such zones. Our observations suggest that RAD51-mediated homologous pairing during HR takes place within the distinctive nuclear zones to execute appropriate recombination.


title: The evolutionary history of human spindle genes includes back-and-forth gene flow with Neandertals

abstract: Proteins associated with the spindle apparatus, a cytoskeletal structure that ensures the proper segregation of chromosomes during cell division, experienced an unusual number of amino acid substitutions in modern humans after the split from the ancestors of Neandertals and Denisovans. Here, we analyze the history of these substitutions and show that some of the genes in which they occur may have been targets of positive selection. We also find that the two changes in the kinetochore scaffold 1 (KNL1) protein, previously believed to be specific to modern humans, were present in some Neandertals. We show that the KNL1 gene of these Neandertals shared a common ancestor with present-day Africans about 200,000 years ago due to gene flow from the ancestors (or relatives) of modern humans into Neandertals. Subsequently, some non-Africans inherited this modern human-like gene variant from Neandertals, but none inherited the ancestral gene variants. These results add to the growing evidence of early contacts between modern humans and archaic groups in Eurasia and illustrate the intricate relationships among these groups.


title: Genome-wide analyses reveal a strong association between LEPR gene variants and body fat reserves in ewes

abstract: Among the adaptive capacities of animals, the management of energetic body reserves (BR) through the BR mobilization and accretion processes (BR dynamics, BRD) has become an increasingly valuable attribute for livestock sustainability, allowing animals to cope with more variable environments. BRD has previously been reported to be heritable in ruminants. In the present study, we conducted genome-wide studies (GWAS) in sheep to determine genetic variants associated with BRD. BR levels and BR changes over time were obtained through body condition score measurements at eight physiological stages throughout each productive cycle in Romane ewes (n=1034) and were used as phenotypes for GWAS. After quality controls and imputation, 48,513 single nucleotide polymorphisms (SNP) were included in the GWAS. Among the QTLs identified, a major QTL associated with BR levels during pregnancy and lactation was identified on chromosome 1. In this region, several significant SNPs mapped to the leptin receptor gene (LEPR), among which one SNP mapped to the coding sequence. The point mutation induces the p.P1019S substitution in the cytoplasmic domain, close to tyrosine phosphorylation sites. The frequency of the SNP associated with increased BR levels was 32%, and the LEPR genotype explained up to 5% of the variance of the trait. These results provide strong evidence for involvement of LEPR in the regulation of BRD in sheep and highlight it as a major candidate for improving adaptive capacities.


title: Pancreatic cancer ductal cell of origin drives CD73-dependent generation of immunosuppressive adenosine

abstract:
The microenvironment that surrounds pancreatic ductal adenocarcinoma (PDAC) is profoundly desmoplastic and immunosuppressive. Understanding initial triggers of immunosuppression during the process of pancreatic tumorigenesis would aid in establishing novel targets for effective prevention and therapy. Here, we interrogate the differential molecular mechanisms dependent on cell of origin and pathology subtype that determine immunosuppression during PDAC initiation and in established tumors. Transcriptomic analysis of cell of origin dependent-epithelial gene signatures revealed that Nt5e/CD73, a cell surface enzyme that is the pacemaker for extracellular adenosine generation, is one of the top 10% of genes over-expressed in murine tumors arising from ductal pancreatic epithelium as opposed to those rising from acinar cells. These findings were confirmed by Imaging Mass Cytometry and High-Performance Liquid Chromatography. Our data indicate that ductal activation of oncogenic mutant Kras results in loss of PTEN and elevated AKT signaling which ultimately releases CD73 suppression. Delivery of CD73 small molecule inhibitors through various delivery routes reduced tumor development and growth in genetically engineered and syngeneic mouse models. Analysis in human PDAC subtypes indicates that high Nt5e in murine ductal PDAC models overlaps with high NT5E in human PDAC Squamous and Basal Subtypes, considered to have the highest immunosuppression and worst prognosis. These findings highlight a molecular trigger of the immunosuppressive PDAC microenvironment which is dependent on ductal cell of origin, linking biology with pathological subtype classification, critical components to personalized approaches for PDAC prevention and immunotherapeutic intervention.

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category: cancer biology


title: Endocytic Trafficking Promotes Vacuolar Enlargements for Fast Cell Expansion Rates in Plants

abstract: The vacuole has a space-filling function, allowing a particularly rapid plant cell expansion with very little increase in cytosolic content (Lofke et al., 2015; Scheuring et al., 2016; Dunser et al., 2019). Despite its importance for cell size determination in plants, very little is known about the mechanisms that define vacuolar size. Here we show that the cellular and vacuolar size expansions are coordinated. By developing a pharmacological tool, we enabled the investigation of membrane delivery to the vacuole during cellular expansion. Counterintuitively, our data reveal that endocytic trafficking from the plasma membrane to the vacuole is enhanced in the course of rapid root cell expansion. While this "compromise" mechanism may theoretically at first decelerate cell surface enlargements, it fuels vacuolar expansion and, thereby, ensures the coordinated augmentation of vacuolar occupancy in dynamically expanding plant cells.

category: plant biology


title: Structure of Dunaliella Photosystem II reveals conformational flexibility of stacked and unstacked supercomplexes

abstract: Photosystem II (PSII) generates an oxidant whose redox potential is high enough to enable water oxidation1,2, a substrate so abundant that it assures a practically unlimited electron source for life on earth3. Our knowledge on the mechanism of water photooxidation was greatly advanced by high-resolution structures of prokaryotic PSII4-6. Here we show high-resolution structures of eukaryotic PSII from the green algae Dunaliella salina at two distinct conformations. The conformers are also present in stacked PSII, exhibiting flexibility that is relevant to the grana formation in chloroplasts of the green lineage. CP29, one of PSII associated light harvesting antennae, plays a major role in distinguishing the two conformations of the supercomplex. We also show that the stacked PSII dimer, a form suggested to support the organization of thylakoid membranes7,8, can appear in many different orientations providing a flexible stacking mechanism for the arrangement of grana stacks in thylakoids. Our findings provide a structural basis for the heterogenous nature of the eukaryotic PSII on multiple levels.


title: Shared components of heritability across genetically correlated traits

abstract: Most disease-associated genetic variants are pleiotropic, affecting multiple genetically correlated traits. Their pleiotropic associations can be mechanistically informative: if many variants have similar patterns of association, they may act via similar pleiotropic mechanisms, forming a shared component of heritability. We developed Pleiotropic Decomposition Regression (PDR) to identify shared components and their underlying genetic variants. We validated PDR on simulated data and identified limitations of existing methods in recovering the true components. We applied PDR to three clusters of 5-6 traits genetically correlated with coronary disease, asthma, and type II diabetes respectively, producing biologically interpretable components. For CAD, PDR identified components related to BMI, hypertension and cholesterol, and it clarified the relationship among these highly correlated risk factors. We assigned variants to components, calculated their posterior-mean effect sizes, and performed out-of-sample validation. Our posterior-mean effect sizes pool statistical power across traits and substantially boost the correlation (r2) between true and estimated effect sizes compared with the original summary statistics: by 94% and 70% for asthma and T2D out of sample, and by a predicted 300% for CAD.


title: Distinct roles of the Na+ binding sites in the allosteric coupling mechanism of the glutamate transporter homolog, GltPh.

abstract: Glutamate transporters carry out the concentrative uptake of glutamate by harnessing the ionic gradients present across cellular membranes. A central step in the transport mechanism is the coupled binding of Na+ and substrate. The sodium coupled Asp transporter, GltPh is an archaeal homolog of glutamate transporters that has been extensively used to probe the transport mechanism. Previous studies have shown that hairpin-2 (HP2) functions as the extracellular gate for the aspartate binding site and plays a key role in the coupled binding of sodium and aspartate to GltPh. The binding sites for three Na+ ions (Na1-3) have been identified in GltPh but the specific roles of the individual Na+ sites in the binding process has not been elucidated. In this study, we developed assays to probe Na+ binding to the Na1 and Na3 sites and to monitor the conformational switch in the NMDGT motif. We used these assays along with a fluorescence assay to monitor HP2 movement and EPR spectroscopy to show that Na+ binding to the Na3 site is required for the NMDGT conformational switch while Na+ binding to the Na1 site is responsible for the partial opening of HP2. Complete opening of HP2 requires the conformational switch of the NMDGT motif and therefore Na+ binding to both the Na1 and the Na3 sites. Based on our studies we also propose an alternate pathway for the coupled binding of Na+ and Asp.


title: A Conformational Transition of Von Willebrand Factor's D'D3 Domain Primes It For Multimerization

abstract: Von Willebrand factor (VWF) is a multimeric plasma glycoprotein that is critically involved in hemostasis. Biosynthesis of long VWF concatemers in the endoplasmic reticulum and the (trans-)Golgi is still not fully understood. We use the single-molecule force spectroscopy technique magnetic tweezers to analyze a previously hypothesized conformational change in the DD3 domain crucial for VWF multimerization. We find that the interface formed by submodules C8-3, TIL3, and E3 wrapping around VWD3 can open and expose two previously buried cysteines that are known to be vital for multimerization. By characterizing the conformational change at varying levels of force, we are able to quantify the kinetics of the transition and the stability of the interface. We find a pronounced destabilization of the interface upon lowering the pH from 7.4 to 6.2 and 5.5. This is consistent with initiation of the conformational change that enables VWF multimerization at the DD3 domain by a decrease in pH in the trans-Golgi network and Weibel-Palade bodies. Furthermore, we find a stabilization of the interface in the presence of coagulation factor VIII (FVIII), providing evidence for a previously hypothesized binding site in submodule C8-3. Our findings highlight the critical role of the DD3 domain in VWF biosynthesis and function and we anticipate our methodology to be applicable to study other, similar conformational changes in VWF and beyond.


title: A Binary Matrix Method to Enumerate, Hierarchically Order and Structurally Classify Peptide Aggregation

abstract:
Protein aggregation is a common and complex phenomenon in biological processes, yet a robust analysis of this aggregation process remains elusive. The commonly used methods such as center-of-mass to center-of-mass (COM-COM) distance, the radius of gyration (Rg), hydrogen bonding (HB) and solvent accessible surface area (SASA) do not quantify the aggregation accurately. Herein, a new and robust method that uses an aggregation matrix (AM) approach to investigate peptide aggregation in a MD simulation trajectory is presented. A nxn two-dimensional aggregation matrix (AM) is created by using the inter-peptide C-C cut-off distances which are binarily encoded (0 or 1). These aggregation matrices are analyzed to enumerate, hierarchically order and structurally classify the aggregates. Comparison of the present AM method suggests that it is superior to the HB method since it can incorporate non-specific interactions and Rg, COM-COM methods since the cut-off distance is independent of the length of the peptide. More importantly, the present method can structurally classify the peptide aggregates, which the conventional Rg, COM- COM and HB methods fail. The unique selling point of this method is its ability to structurally classify peptide aggregates using two-dimensional matrices.

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