Pub Date : 2023-11-14DOI: 10.1101/2023.11.14.567030
Francesca G Smith, John P Goertz, Molly M Stevens, Thomas E Ouldridge
Strand displacement reactions underlie dynamic nucleic acid nanotechnology. The kinetic and thermodynamic features of DNA-based displacement reactions are well understood and well predicted by current computational models. By contrast, understanding of RNA/DNA hybrid strand displacement kinetics is limited, restricting the design of increasingly complex RNA/DNA hybrid reaction networks with more tightly regulated dynamics. Given the importance of RNA as a diagnostic biomarker, and its critical role in intracellular processes, this shortfall is particularly limiting for the development of strand displacement-based therapeutics and diagnostics. Herein, we characterise 22 RNA/DNA hybrid strand displacement systems, systematically varying several common design parameters including toehold length and branch migration domain length. We observe the differences in stability between RNA-DNA hybrids and DNA-DNA duplexes have large effects on strand displacement rates, with rates for equivalent sequences differing by up to 3 orders of magnitude. Crucially, however, this effect is strongly sequence-dependent, with RNA invaders strongly favoured in a system with RNA strands of high purine content, and disfavoured in a system when the RNA strands have low purine content. These results lay the groundwork for more general design principles, allowing for creation of de novo reaction networks with novel complexity while maintaining predictable reaction kinetics.
{"title":"Strong sequence dependence in RNA/DNA hybrid strand displacement kinetics","authors":"Francesca G Smith, John P Goertz, Molly M Stevens, Thomas E Ouldridge","doi":"10.1101/2023.11.14.567030","DOIUrl":"https://doi.org/10.1101/2023.11.14.567030","url":null,"abstract":"Strand displacement reactions underlie dynamic nucleic acid nanotechnology. The kinetic and thermodynamic features of DNA-based displacement reactions are well understood and well predicted by current computational models. By contrast, understanding of RNA/DNA hybrid strand displacement kinetics is limited, restricting the design of increasingly complex RNA/DNA hybrid reaction networks with more tightly regulated dynamics. Given the importance of RNA as a diagnostic biomarker, and its critical role in intracellular processes, this shortfall is particularly limiting for the development of strand displacement-based therapeutics and diagnostics. Herein, we characterise 22 RNA/DNA hybrid strand displacement systems, systematically varying several common design parameters including toehold length and branch migration domain length. We observe the differences in stability between RNA-DNA hybrids and DNA-DNA duplexes have large effects on strand displacement rates, with rates for equivalent sequences differing by up to 3 orders of magnitude. Crucially, however, this effect is strongly sequence-dependent, with RNA invaders strongly favoured in a system with RNA strands of high purine content, and disfavoured in a system when the RNA strands have low purine content. These results lay the groundwork for more general design principles, allowing for creation of de novo reaction networks with novel complexity while maintaining predictable reaction kinetics.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"48 14","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992190","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Implementing integrated pest management programs to limit agricultural pest damage requires an understanding of the interactions between the environmental variability and population demographic processes. However, identifying key environmental drivers of spatiotemporal pest population dynamics remains challenging as numerous candidate factors can operate at a range of scales, from the field (e.g. agricultural practices) to the regional scale (e.g. weather variability). In such a context, data-driven approaches applied to pre-existing data may allow identifying patterns, correlations, and trends that may not be apparent through more restricted hypothesis-driven studies. The resulting insights can lead to the generation of novel hypotheses and inform future experimental work focusing on a limited and relevant set of environmental predictors. In this study, we developed an ecoinformatics approach to unravel the multi-scale environmental conditions that lead to the early re-infestation of mango orchards by a major pest in Senegal, the oriental fruit fly Bactrocera dorsalis (BD). We gathered abundance data from a three-year monitoring conducted in 69 mango orchards as well as environmental data (i.e. orchard management, landscape structure and weather variability) across a range of spatial scales. We then developed a flexible analysis pipeline centred on a recent machine learning algorithm (GPBoost), which allows the combination of gradient boosting and mixed-effects models or Gaussian processes, to hierarchize the effects of multi-scale environmental variables on the timing of annual BD population growth in orchards. We found that physical factors (temperature, humidity), and to some extent landscape features, were the main drivers of the spatio-temporal variability of the onset of population growth in orchards. These results suggest that favourable microclimate conditions could provide refuges for small BD populations that could survive, with little or no reproduction, during the mango off-season and, then, recolonize neighbouring orchards at the beginning of the next mango season. Confirmation of such a hypothesis could help to prioritize surveillance and preventive control actions in refuge areas.
{"title":"Hierarchizing multi-scale environmental effects on agricultural pest population dynamics: a case study on the annual onset of Bactrocera dorsalis population growth in Senegalese orchards","authors":"Cecile Caumette, Paterne Diatta, Sylvain Piry, Marie-Pierre Chapuis, Emile Faye, Fabio Sigrist, Olivier Martin, Julien Papaix, Thierry Brevault, Karine Berthier","doi":"10.1101/2023.11.10.566583","DOIUrl":"https://doi.org/10.1101/2023.11.10.566583","url":null,"abstract":"Implementing integrated pest management programs to limit agricultural pest damage requires an understanding of the interactions between the environmental variability and population demographic processes. However, identifying key environmental drivers of spatiotemporal pest population dynamics remains challenging as numerous candidate factors can operate at a range of scales, from the field (e.g. agricultural practices) to the regional scale (e.g. weather variability). In such a context, data-driven approaches applied to pre-existing data may allow identifying patterns, correlations, and trends that may not be apparent through more restricted hypothesis-driven studies. The resulting insights can lead to the generation of novel hypotheses and inform future experimental work focusing on a limited and relevant set of environmental predictors. In this study, we developed an ecoinformatics approach to unravel the multi-scale environmental conditions that lead to the early re-infestation of mango orchards by a major pest in Senegal, the oriental fruit fly Bactrocera dorsalis (BD). We gathered abundance data from a three-year monitoring conducted in 69 mango orchards as well as environmental data (i.e. orchard management, landscape structure and weather variability) across a range of spatial scales. We then developed a flexible analysis pipeline centred on a recent machine learning algorithm (GPBoost), which allows the combination of gradient boosting and mixed-effects models or Gaussian processes, to hierarchize the effects of multi-scale environmental variables on the timing of annual BD population growth in orchards. We found that physical factors (temperature, humidity), and to some extent landscape features, were the main drivers of the spatio-temporal variability of the onset of population growth in orchards. These results suggest that favourable microclimate conditions could provide refuges for small BD populations that could survive, with little or no reproduction, during the mango off-season and, then, recolonize neighbouring orchards at the beginning of the next mango season. Confirmation of such a hypothesis could help to prioritize surveillance and preventive control actions in refuge areas.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"44 13","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992320","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.14.567055
Linda X Phan, Aaron P Owji, Tingting Yang, Jason Crain, Mark Sansom, Stephen J Tucker
Mechanisms of anion permeation within ion channels and nanopores remain poorly understood. Recent cryo-electron microscopy structures of the human bestrophin 1 chloride channel (hBest1) provide an opportunity to evaluate ion interactions predicted by molecular dynamics (MD) simulations against experimental observations. We implement the fully polarizable forcefield AMOEBA in MD simulations of open and partially-open states of the hBest1. The AMOEBA forcefield models multipole moments up to the quadrupole; therefore, it captures induced dipole and anion-π interactions. By including polarization we demonstrate the key role that aromatic residues play in ion permeation and the functional advantages of pore asymmetry within the highly conserved hydrophobic neck of the pore. We establish that these only arise when electronic polarization is included in the molecular models. We also show that Cl − permeation in this region can be achieved through hydrophobic solvation concomitant with partial ion dehydration, which is compensated for by the formation of contacts with the edge of the phenylalanine ring. Furthermore, we demonstrate how polarizable simulations can help determine the identity of ion-like densities within high-resolution cryo-EM structures. Crucially, neglecting polarization in simulation of these systems results in the localization of Cl − at positions that do not correspond with their experimentally resolved location. Overall, our results demonstrate the importance of including electronic polarization in realistic and physically accurate models of biological systems.
{"title":"Electronic Polarizability Tunes the Function of the Human Bestrophin 1 Cl<sup>−</sup>Channel","authors":"Linda X Phan, Aaron P Owji, Tingting Yang, Jason Crain, Mark Sansom, Stephen J Tucker","doi":"10.1101/2023.11.14.567055","DOIUrl":"https://doi.org/10.1101/2023.11.14.567055","url":null,"abstract":"Mechanisms of anion permeation within ion channels and nanopores remain poorly understood. Recent cryo-electron microscopy structures of the human bestrophin 1 chloride channel (hBest1) provide an opportunity to evaluate ion interactions predicted by molecular dynamics (MD) simulations against experimental observations. We implement the fully polarizable forcefield AMOEBA in MD simulations of open and partially-open states of the hBest1. The AMOEBA forcefield models multipole moments up to the quadrupole; therefore, it captures induced dipole and anion-π interactions. By including polarization we demonstrate the key role that aromatic residues play in ion permeation and the functional advantages of pore asymmetry within the highly conserved hydrophobic neck of the pore. We establish that these only arise when electronic polarization is included in the molecular models. We also show that Cl − permeation in this region can be achieved through hydrophobic solvation concomitant with partial ion dehydration, which is compensated for by the formation of contacts with the edge of the phenylalanine ring. Furthermore, we demonstrate how polarizable simulations can help determine the identity of ion-like densities within high-resolution cryo-EM structures. Crucially, neglecting polarization in simulation of these systems results in the localization of Cl − at positions that do not correspond with their experimentally resolved location. Overall, our results demonstrate the importance of including electronic polarization in realistic and physically accurate models of biological systems.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"40 6","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992526","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.10.566611
Everett A Abhainn, Devin L Shirley, Robert K Stanley, Tatum Scarpato, Jennifer L Koch, Jeanne Romero-Severson
In North America, the ubiquitous planting of a comparatively small number of Fraxinus (ash) cultivars in both urban and rural environments over the last 70-80 years may have permitted extensive gene flow into naturally regenerated stands. In the light of multiple biotic threats to the North American Fraxinus, an assessment of the extent of gene flow from ash cultivars and the current state of genetic diversity in F. pennsylvanica (green ash), one of the most widely distributed species, is needed to inform seed collection strategies for the preservation of genetic diversity range-wide. We used 16 EST-SSR markers to genotype 1291 trees from 48 naturally regenerated populations of green ash across the native range, 19 F. pennsylvanica cultivars and one F. americana (white ash) cultivar. We detected first generation cultivar parentage with high confidence in 172 individuals in 34 of the 48 populations and extensive cultivar parentage (23-50%) in eight populations. The incidence of cultivar parentage was negatively associated with allele richness (R2 = 0.151, p = 0.006). The high frequency of cultivar propagule dispersal in our study suggests that a significant proportion of the standing genetic variation in local populations may not be of local origin, a result that has serious implications for the study of adaptive variation and the conservation of the Fraxinus gene pool.
在北美，在过去的70-80年里，在城市和农村环境中普遍种植了相对较少数量的白蜡属(白蜡)品种，这可能使大量的基因流入自然再生的林分。鉴于北美白蜡属植物面临的多重生物威胁，有必要对白蜡属植物中分布最广泛的白蜡属植物之一宾夕法尼亚白蜡属植物的遗传多样性现状和白蜡属栽培品种的基因流动程度进行评估，为保护白蜡属植物的遗传多样性提供信息。利用16个EST-SSR标记对48个自然再生的白蜡树、19个宾夕法尼亚白蜡树和1个美洲白蜡树进行了基因型分析。在48个群体中，34个群体中有172个个体的第一代亲缘关系具有较高的置信度，在8个群体中有广泛的亲缘关系(23-50%)。品种亲本发生率与等位基因丰富度呈负相关(R2 = 0.151, p = 0.006)。本研究中品种繁殖体传播的高频率表明，当地种群中存在的很大一部分遗传变异可能不是来自当地，这一结果对研究曲霉的适应性变异和基因库的保护具有重要意义。
{"title":"Gene flow from Fraxinus cultivars into natural stands of Fraxinus pennsylvanica occurs range-wide, is regionally extensive, and is associated with a loss of allele richness","authors":"Everett A Abhainn, Devin L Shirley, Robert K Stanley, Tatum Scarpato, Jennifer L Koch, Jeanne Romero-Severson","doi":"10.1101/2023.11.10.566611","DOIUrl":"https://doi.org/10.1101/2023.11.10.566611","url":null,"abstract":"In North America, the ubiquitous planting of a comparatively small number of Fraxinus (ash) cultivars in both urban and rural environments over the last 70-80 years may have permitted extensive gene flow into naturally regenerated stands. In the light of multiple biotic threats to the North American Fraxinus, an assessment of the extent of gene flow from ash cultivars and the current state of genetic diversity in F. pennsylvanica (green ash), one of the most widely distributed species, is needed to inform seed collection strategies for the preservation of genetic diversity range-wide. We used 16 EST-SSR markers to genotype 1291 trees from 48 naturally regenerated populations of green ash across the native range, 19 F. pennsylvanica cultivars and one F. americana (white ash) cultivar. We detected first generation cultivar parentage with high confidence in 172 individuals in 34 of the 48 populations and extensive cultivar parentage (23-50%) in eight populations. The incidence of cultivar parentage was negatively associated with allele richness (R2 = 0.151, p = 0.006). The high frequency of cultivar propagule dispersal in our study suggests that a significant proportion of the standing genetic variation in local populations may not be of local origin, a result that has serious implications for the study of adaptive variation and the conservation of the Fraxinus gene pool.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"34 15","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992677","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.10.566533
Nicolas Virlet, João Paulo Pennacchi, Pouria Sadeghi-Tehran, Tom Ashfield, Douglas Orr, Elizabete Carmo-Silva, Malcolm Hawkesford
With the development of the digital phenotyping, repeated measurements of agronomic traits over time are easily accessible, notably for morphological and phenological traits. However high throughput methods for estimating physiological traits such as photosynthesis are lacking. This study demonstrates the links of fluorescence and reflectance imaging with photosynthetic traits. Two wheat cultivars were grown in pots in a controlled environment. Photosynthesis was characterised by gas-exchange and biochemical analysis at five time points, from booting to 21 days post anthesis. On the same days imaging was performed on the same pots, at leaf and plant scale, using indoor and outdoor phenotyping platforms, respectively. Five image variables (Fv/Fm and NDVI at the whole plant level and Fv/Fm, Φ(II)532 and Φ(NPQ)1077 at the leaf scale) were compared to variables from A-Ci and A-Par curves, biochemical analysis, and fluorescence instruments. The results suggested that the image variables are robust estimators of photosynthetic traits, as long as senescence is driving the variability. Despite contrasting cultivar behaviour, linear regression models which account for the cultivar and the interaction effects, further improved the modelling of photosynthesis indicators. Finally, the results highlight the challenge of discriminating functional to cosmetic stay green genotypes using digital imaging.
{"title":"A multiscale approach to investigate fluorescence and NDVI imaging as proxy of photosynthetic traits in wheat","authors":"Nicolas Virlet, João Paulo Pennacchi, Pouria Sadeghi-Tehran, Tom Ashfield, Douglas Orr, Elizabete Carmo-Silva, Malcolm Hawkesford","doi":"10.1101/2023.11.10.566533","DOIUrl":"https://doi.org/10.1101/2023.11.10.566533","url":null,"abstract":"With the development of the digital phenotyping, repeated measurements of agronomic traits over time are easily accessible, notably for morphological and phenological traits. However high throughput methods for estimating physiological traits such as photosynthesis are lacking. This study demonstrates the links of fluorescence and reflectance imaging with photosynthetic traits. Two wheat cultivars were grown in pots in a controlled environment. Photosynthesis was characterised by gas-exchange and biochemical analysis at five time points, from booting to 21 days post anthesis. On the same days imaging was performed on the same pots, at leaf and plant scale, using indoor and outdoor phenotyping platforms, respectively. Five image variables (Fv/Fm and NDVI at the whole plant level and Fv/Fm, Φ(II)532 and Φ(NPQ)1077 at the leaf scale) were compared to variables from A-Ci and A-Par curves, biochemical analysis, and fluorescence instruments. The results suggested that the image variables are robust estimators of photosynthetic traits, as long as senescence is driving the variability. Despite contrasting cultivar behaviour, linear regression models which account for the cultivar and the interaction effects, further improved the modelling of photosynthesis indicators. Finally, the results highlight the challenge of discriminating functional to cosmetic stay green genotypes using digital imaging.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"37 5","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992985","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.13.566806
Nandan Haloi, Emelia Karlsson, Rebecca J. Howard, Erik Lindahl
Ligand-gated ion channels propagate electrochemical signals in the nervous system by opening ion-selective pores in response to neurotransmitter release. A diverse set of allosteric modulators including neurosteroids, anesthetics, and lipids modulate their function in myriad ways, suggesting a complex conformational landscape. However, structures of ligand-bound ion-channel complexes can be difficult to capture by experimental techniques like cryogenic electron microscopy, particularly when binding is dynamic or transient. Here, we used computational methods to identify a possible bound state of a modulatory stimulant derivative (4-bromoamphetamine) in a cryptic vestibular pocket of a mammalian serotonin-3A receptor. Starting from an experimental activated structure containing a closed pocket, we first applied a molecular dynamics (MD) simulations-based goal-oriented adaptive sampling method to identify possible open-pocket conformations. To find plausible ligand-binding poses, we performed ensemble docking of the newly identified modulator, and reweighted docking scores by the Boltzmann energy function derived from Markov state model analysis of our trajectories. We then performed replicates of unbiased MD simulations of representative complexes in two forcefields to estimate ligand stability, and screened the most stable complexes for accessibility to the aqueous environment. For one relatively stable and accessible site, mutations predicted to disrupt ligand binding were validated by electrophysiology recordings in Xenopus laevis oocytes, and provided a mechanistic rationale for allosteric stabilization of an activated state. Given the pharmaceutical relevance of serotonin-3 receptors in emesis, pain, psychiatric and gastrointestinal diseases, characterizing relatively unexplored modulatory sites in these proteins could open valuable avenues to understanding conformational cycling and designing state-dependent drugs.
{"title":"Discovering cryptic pocket opening and ligand binding in a vestibular site of the 5-HT3A receptor","authors":"Nandan Haloi, Emelia Karlsson, Rebecca J. Howard, Erik Lindahl","doi":"10.1101/2023.11.13.566806","DOIUrl":"https://doi.org/10.1101/2023.11.13.566806","url":null,"abstract":"Ligand-gated ion channels propagate electrochemical signals in the nervous system by opening ion-selective pores in response to neurotransmitter release. A diverse set of allosteric modulators including neurosteroids, anesthetics, and lipids modulate their function in myriad ways, suggesting a complex conformational landscape. However, structures of ligand-bound ion-channel complexes can be difficult to capture by experimental techniques like cryogenic electron microscopy, particularly when binding is dynamic or transient. Here, we used computational methods to identify a possible bound state of a modulatory stimulant derivative (4-bromoamphetamine) in a cryptic vestibular pocket of a mammalian serotonin-3A receptor. Starting from an experimental activated structure containing a closed pocket, we first applied a molecular dynamics (MD) simulations-based goal-oriented adaptive sampling method to identify possible open-pocket conformations. To find plausible ligand-binding poses, we performed ensemble docking of the newly identified modulator, and reweighted docking scores by the Boltzmann energy function derived from Markov state model analysis of our trajectories. We then performed replicates of unbiased MD simulations of representative complexes in two forcefields to estimate ligand stability, and screened the most stable complexes for accessibility to the aqueous environment. For one relatively stable and accessible site, mutations predicted to disrupt ligand binding were validated by electrophysiology recordings in Xenopus laevis oocytes, and provided a mechanistic rationale for allosteric stabilization of an activated state. Given the pharmaceutical relevance of serotonin-3 receptors in emesis, pain, psychiatric and gastrointestinal diseases, characterizing relatively unexplored modulatory sites in these proteins could open valuable avenues to understanding conformational cycling and designing state-dependent drugs.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"36 19","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134992989","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.10.566553
Andrew R Morris, Erwin K Gudenschwager Basso, Miguel A Gutierrez-Monreal, Rawad Daniel Arja, Firas H Kobeissy, Christopher G Janus, Kevin KW Wang, Jiepei Zhu, Andrew C Liu
Chronic sleep/wake disturbances are strongly associated with traumatic brain injury (TBI) in patients and are being increasingly recognized. However, the underlying mechanisms are largely understudied and there is an urgent need for animal models of lifelong sleep/wake disturbances. The objective of this study was to develop a chronic TBI rodent model and investigate the lifelong chronic effect of TBI on sleep/wake behavior. We performed repetitive midline fluid percussion injury (rmFPI) in four months old mice and monitored their sleep/wake behavior using the non-invasive PiezoSleep system. The sleep/wake states were recorded before injury (baseline) and then monthly thereafter. We found that TBI mice displayed a significant decrease in sleep duration in both the light and dark phases, beginning at three months post-TBI and continuing throughout the study. Consistent with the sleep phenotype, these TBI mice showed circadian locomotor activity phenotypes and exhibited reduced anxiety-like behavior. TBI mice also gained less weight, and had less lean mass and total body water content, compared to sham controls. Furthermore, TBI mice showed extensive brain tissue loss and increased GFAP and IBA1 levels in the hypothalamus and the vicinity of the injury, indicative of chronic neuropathology. In summary, our study identified a critical time window of TBI pathology and associated circadian and sleep/wake phenotypes. Future studies should leverage this mouse model to investigate the molecular mechanisms underlying the chronic sleep/wake phenotypes following TBI early in life.
{"title":"Sleep Disruption in a Mouse Model of Chronic Traumatic Brain Injury","authors":"Andrew R Morris, Erwin K Gudenschwager Basso, Miguel A Gutierrez-Monreal, Rawad Daniel Arja, Firas H Kobeissy, Christopher G Janus, Kevin KW Wang, Jiepei Zhu, Andrew C Liu","doi":"10.1101/2023.11.10.566553","DOIUrl":"https://doi.org/10.1101/2023.11.10.566553","url":null,"abstract":"Chronic sleep/wake disturbances are strongly associated with traumatic brain injury (TBI) in patients and are being increasingly recognized. However, the underlying mechanisms are largely understudied and there is an urgent need for animal models of lifelong sleep/wake disturbances. The objective of this study was to develop a chronic TBI rodent model and investigate the lifelong chronic effect of TBI on sleep/wake behavior. We performed repetitive midline fluid percussion injury (rmFPI) in four months old mice and monitored their sleep/wake behavior using the non-invasive PiezoSleep system. The sleep/wake states were recorded before injury (baseline) and then monthly thereafter. We found that TBI mice displayed a significant decrease in sleep duration in both the light and dark phases, beginning at three months post-TBI and continuing throughout the study. Consistent with the sleep phenotype, these TBI mice showed circadian locomotor activity phenotypes and exhibited reduced anxiety-like behavior. TBI mice also gained less weight, and had less lean mass and total body water content, compared to sham controls. Furthermore, TBI mice showed extensive brain tissue loss and increased GFAP and IBA1 levels in the hypothalamus and the vicinity of the injury, indicative of chronic neuropathology. In summary, our study identified a critical time window of TBI pathology and associated circadian and sleep/wake phenotypes. Future studies should leverage this mouse model to investigate the molecular mechanisms underlying the chronic sleep/wake phenotypes following TBI early in life.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"33 7","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134993584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.13.566960
Jacob Lewis Watts, Graham J Dow, Thomas N Buckley, Chris D Muir
Photosynthesis is co-limited by multiple factors depending on the plant and its environment. These include biochemical rate limitations, internal and external water potentials, temperature, irradiance, and carbon dioxide (CO2). Amphistomatous leaves have stomata on both abaxial and adaxial leaf surfaces. This feature is considered an adaptation to alleviate CO2 diffusion limitations in productive environments where other factors are not limiting as the diffusion path length from stomate to chloroplast is effectively halved. Plants can also reduce CO2 limitations through other aspects of optimal stomatal anatomy: stomatal density, distribution, patterning, and size. A number of studies have demonstrated that stomata are overdispersed on a single leaf surface; however, much less is known about stomatal anatomy in amphistomatous leaves, especially the coordination between leaf surfaces, despite their prevelance in nature and near ubiquity among crop species. Here we use novel spatial statistics based on simulations and photosynthesis modeling to test hypotheses about how amphistomatous plants may optimize CO2 limitations in the model angiosperm Arabidopsis thaliana grown in different light environments. We find that 1) stomata are overdispersed, but not ideally dispersed, on both leaf surfaces across all light treatments; 2) abaxial and adaxial leaf surface patterning are independent; and 3) the theoretical improvements to photosynthesis from abaxial-adaxial stomatal coordination are miniscule (≪ 1%) across the range of feasible parameter space. However, we also find that 4) stomatal size is correlated with the mesophyll volume that it supplies with CO2, suggesting that plants may optimize CO2 diffusion limitations through alternative pathways other than ideal, uniform stomatal spacing. We discuss the developmental, physical, and evolutionary constraits which may prohibit plants from reaching the theoretical adaptive peak of uniform stomatal spacing and inter surface stomatal coordination. These findings contribute to our understanding of variation in the anatomy of amphistomatous leaves.
{"title":"Does stomatal patterning in amphistomatous leaves minimize the CO2 diffusion path length within leaves?","authors":"Jacob Lewis Watts, Graham J Dow, Thomas N Buckley, Chris D Muir","doi":"10.1101/2023.11.13.566960","DOIUrl":"https://doi.org/10.1101/2023.11.13.566960","url":null,"abstract":"Photosynthesis is co-limited by multiple factors depending on the plant and its environment. These include biochemical rate limitations, internal and external water potentials, temperature, irradiance, and carbon dioxide (CO2). Amphistomatous leaves have stomata on both abaxial and adaxial leaf surfaces. This feature is considered an adaptation to alleviate CO2 diffusion limitations in productive environments where other factors are not limiting as the diffusion path length from stomate to chloroplast is effectively halved. Plants can also reduce CO2 limitations through other aspects of optimal stomatal anatomy: stomatal density, distribution, patterning, and size. A number of studies have demonstrated that stomata are overdispersed on a single leaf surface; however, much less is known about stomatal anatomy in amphistomatous leaves, especially the coordination between leaf surfaces, despite their prevelance in nature and near ubiquity among crop species. Here we use novel spatial statistics based on simulations and photosynthesis modeling to test hypotheses about how amphistomatous plants may optimize CO2 limitations in the model angiosperm Arabidopsis thaliana grown in different light environments. We find that 1) stomata are overdispersed, but not ideally dispersed, on both leaf surfaces across all light treatments; 2) abaxial and adaxial leaf surface patterning are independent; and 3) the theoretical improvements to photosynthesis from abaxial-adaxial stomatal coordination are miniscule (≪ 1%) across the range of feasible parameter space. However, we also find that 4) stomatal size is correlated with the mesophyll volume that it supplies with CO2, suggesting that plants may optimize CO2 diffusion limitations through alternative pathways other than ideal, uniform stomatal spacing. We discuss the developmental, physical, and evolutionary constraits which may prohibit plants from reaching the theoretical adaptive peak of uniform stomatal spacing and inter surface stomatal coordination. These findings contribute to our understanding of variation in the anatomy of amphistomatous leaves.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"33 18","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134993736","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.13.566904
Vanessa Lopes-Rodrigues, Samuel A. Nyantakyi, Xueqing Lun, Jianbo Zhang, Ajeena Ramanujuan, Shuhailah Salim, Michael Saleeb, Donna L. Senger, Carlos F. Ibanez
Receptor transmembrane domains (TMDs) are crucially involved in relaying ligand information from extracellular to intracellular spaces and represent attractive targets for small molecule manipulation of receptor function. Screening a library of over 8,000 drug-like compounds with an assay based on the TMD of death receptor p75NTR, we identified a novel small molecule capable of inhibiting p75NTR-mediated migration of human melanoma cells. Employing medicinal chemistry, a more potent derivative termed Np75-4A22 was identified that blocks nerve growth factor (NGF)-mediated melanoma invasion at submicromolar concentrations. Mechanistically, Np75-4A22 was found, at least in part, to function by antagonizing NGF-mediated recruitment of the actin-bundling protein fascin to p75NTR and its association with the actin cytoskeleton. Importantly, preclinical assessment of Np75-4A22 showed high oral bioavailability, low toxicity, and significant inhibition of melanoma lung metastases in a highly metastatic mouse model. These results support further development of this approach as an alternative or complementary strategy for patients that do not respond to conventional chemotherapy or immune checkpoint inhibitors.
{"title":"Impaired migration and metastatic spread of human melanoma by a novel small molecule targeting the transmembrane domain of death receptor p75NTR","authors":"Vanessa Lopes-Rodrigues, Samuel A. Nyantakyi, Xueqing Lun, Jianbo Zhang, Ajeena Ramanujuan, Shuhailah Salim, Michael Saleeb, Donna L. Senger, Carlos F. Ibanez","doi":"10.1101/2023.11.13.566904","DOIUrl":"https://doi.org/10.1101/2023.11.13.566904","url":null,"abstract":"Receptor transmembrane domains (TMDs) are crucially involved in relaying ligand information from extracellular to intracellular spaces and represent attractive targets for small molecule manipulation of receptor function. Screening a library of over 8,000 drug-like compounds with an assay based on the TMD of death receptor p75NTR, we identified a novel small molecule capable of inhibiting p75NTR-mediated migration of human melanoma cells. Employing medicinal chemistry, a more potent derivative termed Np75-4A22 was identified that blocks nerve growth factor (NGF)-mediated melanoma invasion at submicromolar concentrations. Mechanistically, Np75-4A22 was found, at least in part, to function by antagonizing NGF-mediated recruitment of the actin-bundling protein fascin to p75NTR and its association with the actin cytoskeleton. Importantly, preclinical assessment of Np75-4A22 showed high oral bioavailability, low toxicity, and significant inhibition of melanoma lung metastases in a highly metastatic mouse model. These results support further development of this approach as an alternative or complementary strategy for patients that do not respond to conventional chemotherapy or immune checkpoint inhibitors.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"56 6","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134901678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-11-14DOI: 10.1101/2023.11.14.566925
Gabriela Zurawska, Zuzanna Sas, Aneta Jonczy, Patryk Slusarczyk, Raghunandan Mahadeva, Marta Chwalek, Maria Kulecka, Izabela Rumienczyk, Morgane Moulin, Kamil Jastrzebski, Michal Mikula, Anders Etzerodt, Marta Miaczynska, Tomasz P. Rygiel, Katarzyna Mleczko-Sanecka
Mild hemolysis of senescent erythrocytes occurs physiologically in the spleen, resulting in hemoglobin (Hb) release, whereas pathologic erythrocyte rupture characterizes several diseases. Iron recycling from Hb and Hb detoxification have been attributed to the sequestration of Hb-haptoglobin complexes by macrophages. However, we found the existence of additional efficient Hb clearance routes in mice. We identified liver sinusoidal endothelial cells (LSECs) as the primary cells responsible for Hb sequestration, a process that involves macropinocytosis and operates independently of the Hb-haptoglobin receptor CD163. LSECs expressed heme oxygenase 1 and hepcidin-controlled ferroportin and were the most efficient cellular scavengers of Hb at doses below and above the haptoglobin binding capacity. Erythrocyte transfusion assays further demonstrated that while splenic red pulp macrophages are adept at erytrophagocytosis, liver Kupffer cells and LSECs mainly clear erythrocyte ghosts and Hb, respectively, transported from the spleen via the portal circulation. High-dose Hb injections in mice resulted in transient hepatic iron retention and early activation of the gene encoding heme oxygenase 1 (Hmox1) in LSECs. This response was associated with the transcriptional induction of the iron-sensing angiokine Bmp6, culminating in hepcidin-mediated transient serum hypoferremia. Injection of Hb and iron citrate elicited distinct transcriptional signatures in LSECs, and the Bmp6 induction was phenocopied by erythrocyte lysis upon phenylhydrazine. Collectively, we propose that LSECs provide a key mechanism for Hb clearance, a function that establishes the spleen-liver axis for physiological iron recycling from Hb and contributes to heme detoxification during hemolysis, coupled with the induction of the BMP6-hepcidin axis, ultimately restoring iron homeostasis.
{"title":"Liver sinusoidal endothelial cells constitute a major route for hemoglobin clearance","authors":"Gabriela Zurawska, Zuzanna Sas, Aneta Jonczy, Patryk Slusarczyk, Raghunandan Mahadeva, Marta Chwalek, Maria Kulecka, Izabela Rumienczyk, Morgane Moulin, Kamil Jastrzebski, Michal Mikula, Anders Etzerodt, Marta Miaczynska, Tomasz P. Rygiel, Katarzyna Mleczko-Sanecka","doi":"10.1101/2023.11.14.566925","DOIUrl":"https://doi.org/10.1101/2023.11.14.566925","url":null,"abstract":"Mild hemolysis of senescent erythrocytes occurs physiologically in the spleen, resulting in hemoglobin (Hb) release, whereas pathologic erythrocyte rupture characterizes several diseases. Iron recycling from Hb and Hb detoxification have been attributed to the sequestration of Hb-haptoglobin complexes by macrophages. However, we found the existence of additional efficient Hb clearance routes in mice. We identified liver sinusoidal endothelial cells (LSECs) as the primary cells responsible for Hb sequestration, a process that involves macropinocytosis and operates independently of the Hb-haptoglobin receptor CD163. LSECs expressed heme oxygenase 1 and hepcidin-controlled ferroportin and were the most efficient cellular scavengers of Hb at doses below and above the haptoglobin binding capacity. Erythrocyte transfusion assays further demonstrated that while splenic red pulp macrophages are adept at erytrophagocytosis, liver Kupffer cells and LSECs mainly clear erythrocyte ghosts and Hb, respectively, transported from the spleen via the portal circulation. High-dose Hb injections in mice resulted in transient hepatic iron retention and early activation of the gene encoding heme oxygenase 1 (Hmox1) in LSECs. This response was associated with the transcriptional induction of the iron-sensing angiokine Bmp6, culminating in hepcidin-mediated transient serum hypoferremia. Injection of Hb and iron citrate elicited distinct transcriptional signatures in LSECs, and the Bmp6 induction was phenocopied by erythrocyte lysis upon phenylhydrazine. Collectively, we propose that LSECs provide a key mechanism for Hb clearance, a function that establishes the spleen-liver axis for physiological iron recycling from Hb and contributes to heme detoxification during hemolysis, coupled with the induction of the BMP6-hepcidin axis, ultimately restoring iron homeostasis.","PeriodicalId":486943,"journal":{"name":"bioRxiv (Cold Spring Harbor Laboratory)","volume":"29 12","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134954549","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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