Pub Date : 2024-10-15Epub Date: 2024-09-14DOI: 10.1016/j.bpj.2024.09.005
Sam Walcott, Sean Sun, Edward P Debold, Walter Herzog
{"title":"In defense of Huxley.","authors":"Sam Walcott, Sean Sun, Edward P Debold, Walter Herzog","doi":"10.1016/j.bpj.2024.09.005","DOIUrl":"10.1016/j.bpj.2024.09.005","url":null,"abstract":"","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142280071","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Fluorescence microscopy, which employs fluorescent tags to label and observe cellular structures and their dynamics, is a powerful tool for life sciences. However, due to the spectral overlap between different dyes, a limited number of structures can be separately labeled and imaged for live-cell applications. In addition, the conventional sequential channel imaging procedure is quite time consuming, as it needs to switch either different lasers or filters. Here, we propose a novel double-structure network (DBSN) that consists of multiple connected models, which can extract six distinct subcellular structures from three raw images with only two separate fluorescent labels. DBSN combines the intensity-balance model to compensate for uneven fluorescent labels for different structures and the structure-separation model to extract multiple different structures with the same fluorescent labels. Therefore, DBSN breaks the bottleneck of the existing technologies and holds immense potential applications in the field of cell biology.
{"title":"Deep learning permits imaging of multiple structures with the same fluorophores.","authors":"Luhong Jin, Jingfang Liu, Heng Zhang, Yunqi Zhu, Haixu Yang, Jianhang Wang, Luhao Zhang, Cuifang Kuang, Baohua Ji, Ju Zhang, Xu Liu, Yingke Xu","doi":"10.1016/j.bpj.2024.09.001","DOIUrl":"10.1016/j.bpj.2024.09.001","url":null,"abstract":"<p><p>Fluorescence microscopy, which employs fluorescent tags to label and observe cellular structures and their dynamics, is a powerful tool for life sciences. However, due to the spectral overlap between different dyes, a limited number of structures can be separately labeled and imaged for live-cell applications. In addition, the conventional sequential channel imaging procedure is quite time consuming, as it needs to switch either different lasers or filters. Here, we propose a novel double-structure network (DBSN) that consists of multiple connected models, which can extract six distinct subcellular structures from three raw images with only two separate fluorescent labels. DBSN combines the intensity-balance model to compensate for uneven fluorescent labels for different structures and the structure-separation model to extract multiple different structures with the same fluorescent labels. Therefore, DBSN breaks the bottleneck of the existing technologies and holds immense potential applications in the field of cell biology.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142131745","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-14DOI: 10.1016/j.bpj.2024.10.009
Russell K W Spencer,Isaac Santos-Perez,Anna V Shnyrova,Marcus Müller
The division of a cellular compartment culminates with the scission of a highly constricted membrane neck. Scission requires lipid rearrangements, topology changes, and transient formation of non-bilayer intermediate structures driven by curvature stress. Often, a side effect of this stress is pore formation that may lead to content leakage and thus breaching of the membrane barrier function. In single membrane systems, leakage is avoided through the formation of a hemifusion (HF) intermediate, whose structure is still a subject of debate. The consequences of curvature stress have not been explored in double-membrane systems, such as the mitochondrion. Here we combine experimental and theoretical approaches to study neck constriction and scission driven by tension in biomimetic lipid systems, namely single- and double-membrane nanotubes (sNTs and dNTs), respectively. In sNTs, constriction by high tension gives rise to a metastable HF intermediate (seen as stalk or worm-like micelle), whereas poration is universally slower in a simple neck. In dNTs, high membrane tension causes sequential rupture of each membrane. In contrast, low tension leads to the hemifusion of both membranes, which may lead to a leaky fusion pathway, or may progress to further fusion of the two membranes along a number of transformation pathways. These findings provide a new mechanistic basis for fundamental cellular processes.
{"title":"Fission of Double-Membrane Tubes under Tension.","authors":"Russell K W Spencer,Isaac Santos-Perez,Anna V Shnyrova,Marcus Müller","doi":"10.1016/j.bpj.2024.10.009","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.009","url":null,"abstract":"The division of a cellular compartment culminates with the scission of a highly constricted membrane neck. Scission requires lipid rearrangements, topology changes, and transient formation of non-bilayer intermediate structures driven by curvature stress. Often, a side effect of this stress is pore formation that may lead to content leakage and thus breaching of the membrane barrier function. In single membrane systems, leakage is avoided through the formation of a hemifusion (HF) intermediate, whose structure is still a subject of debate. The consequences of curvature stress have not been explored in double-membrane systems, such as the mitochondrion. Here we combine experimental and theoretical approaches to study neck constriction and scission driven by tension in biomimetic lipid systems, namely single- and double-membrane nanotubes (sNTs and dNTs), respectively. In sNTs, constriction by high tension gives rise to a metastable HF intermediate (seen as stalk or worm-like micelle), whereas poration is universally slower in a simple neck. In dNTs, high membrane tension causes sequential rupture of each membrane. In contrast, low tension leads to the hemifusion of both membranes, which may lead to a leaky fusion pathway, or may progress to further fusion of the two membranes along a number of transformation pathways. These findings provide a new mechanistic basis for fundamental cellular processes.","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142443669","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-14DOI: 10.1016/j.bpj.2024.10.010
Ethan A Perets,Ty Santiago,Jens Neu,Elsa C Y Yan
{"title":"Water-protein Interactions as a Driver of Phase Separation, Biology, and Disease.","authors":"Ethan A Perets,Ty Santiago,Jens Neu,Elsa C Y Yan","doi":"10.1016/j.bpj.2024.10.010","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.010","url":null,"abstract":"","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142439455","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lipid compositional asymmetry across the leaflets of the plasma membrane is a ubiquitous feature in eukaryotic cells. How this asymmetry is maintained is thought to be primarily controlled by active transport of lipids between leaflets. This strategy is facilitated by the fact that long tail phospholipids and sphingolipids diffuse through the lipid bilayer slowly - taking many hours or days. However, a lipid like cholesterol - which is the most abundant lipid in the plasma membrane of animal cells - has been harder to pin-point in terms of its favored side. In the present work we show that when a saturated lipid is added to a mix of the unsaturated lipid palmitoyl-oleoyl-phosphatidylcholine (POPC) and cholesterol, both cholesterol and the long tail phospholipids organize asymmetrically across the membrane's leaflets naturally. In these extruded unilamellar vesicles, most cholesterol as well as the saturated lipid - dipalmitoylphosphatidylcholine (DPPC) or sphingomyelin (SM) - segregated to the inner leaflet while POPC preferentially localized in the outer leaflet. This asymmetric arrangement generated a slight phospholipid number imbalance favoring the outer leaflet and thus opposite to where cholesterol and the saturated lipids preferentially partitioned. These results were obtained using Magic Angle Spinning (MAS) NMR in combination with Small Angle Neutron Scattering (SANS) using isotope labeling to differentiate lipid species. We suggest that sidedness in membranes can be driven by thermodynamic processes. In addition, our MAS NMR results show that the lower bound for cholesterol's flip-flop half-time at 45°C is 10ms, which is at least two orders of magnitude slower than current MD simulations predict. This result stands in stark contrast to previous work that suggested that cholesterol's flip-flop half-time at 37°C has an upper bound of 10ms.
{"title":"Unexpected asymmetric distribution of cholesterol and phospholipids in equilibrium model membranes.","authors":"Yuli Zhu, Lionel Porcar, Thirupathi Ravula, Krishna C Batchu, Tera Lavoie, Ying Liu, Ursula Perez-Salas","doi":"10.1016/j.bpj.2024.10.004","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.004","url":null,"abstract":"<p><p>Lipid compositional asymmetry across the leaflets of the plasma membrane is a ubiquitous feature in eukaryotic cells. How this asymmetry is maintained is thought to be primarily controlled by active transport of lipids between leaflets. This strategy is facilitated by the fact that long tail phospholipids and sphingolipids diffuse through the lipid bilayer slowly - taking many hours or days. However, a lipid like cholesterol - which is the most abundant lipid in the plasma membrane of animal cells - has been harder to pin-point in terms of its favored side. In the present work we show that when a saturated lipid is added to a mix of the unsaturated lipid palmitoyl-oleoyl-phosphatidylcholine (POPC) and cholesterol, both cholesterol and the long tail phospholipids organize asymmetrically across the membrane's leaflets naturally. In these extruded unilamellar vesicles, most cholesterol as well as the saturated lipid - dipalmitoylphosphatidylcholine (DPPC) or sphingomyelin (SM) - segregated to the inner leaflet while POPC preferentially localized in the outer leaflet. This asymmetric arrangement generated a slight phospholipid number imbalance favoring the outer leaflet and thus opposite to where cholesterol and the saturated lipids preferentially partitioned. These results were obtained using Magic Angle Spinning (MAS) NMR in combination with Small Angle Neutron Scattering (SANS) using isotope labeling to differentiate lipid species. We suggest that sidedness in membranes can be driven by thermodynamic processes. In addition, our MAS NMR results show that the lower bound for cholesterol's flip-flop half-time at 45°C is 10ms, which is at least two orders of magnitude slower than current MD simulations predict. This result stands in stark contrast to previous work that suggested that cholesterol's flip-flop half-time at 37°C has an upper bound of 10ms.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142399238","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-09DOI: 10.1016/j.bpj.2024.10.005
Nels Schimek, Thomas R Wood, David A C Beck, Michael McKenna, Ali Toghani, Elizabeth Nance
Multiple particle tracking (MPT) is a microscopy technique capable of simultaneously tracking hundreds to thousands of nanoparticles in a biological sample and has been used extensively to characterize biological microenvironments, including the brain extracellular space (ECS). Machine learning techniques have been applied to MPT datasets to predict the diffusion mode of nanoparticle trajectories as well as more complex biological variables, such age biological age. In this study, we develop a machine learning pipeline to predict and investigate changes to the brain ECS due to injury using supervised classification and feature importance calculations. We first validate the pipeline on three related but distinct MPT datasets from the living brain ECS - age differences, region differences, and enzymatic degradation of ECS structure. We predict three ages with 86% accuracy, three regions with 90% accuracy, and healthy versus enzyme-treated tissue with 69% accuracy. Since injury across groups is normally compared with traditional statistical approaches, we first used linear mixed effects models to compare features between healthy control conditions and injury induced by two different oxygen glucose deprivation [1] exposure times. We then used machine learning to predict injury state using MPT features. We show that the pipeline predicts between the healthy control, 0.5-hour OGD treatment, and 1.5-hour OGD treatment with 59% accuracy in the cortex and 66% in the striatum and identifies nonlinear relationships between trajectory features that were not evident from traditional linear models. Our work demonstrates that machine learning applied to MPT data is effective across multiple experimental conditions and can find unique biologically relevant features of nanoparticle diffusion.
{"title":"High fidelity predictions of diffusion in the brain microenvironment.","authors":"Nels Schimek, Thomas R Wood, David A C Beck, Michael McKenna, Ali Toghani, Elizabeth Nance","doi":"10.1016/j.bpj.2024.10.005","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.005","url":null,"abstract":"<p><p>Multiple particle tracking (MPT) is a microscopy technique capable of simultaneously tracking hundreds to thousands of nanoparticles in a biological sample and has been used extensively to characterize biological microenvironments, including the brain extracellular space (ECS). Machine learning techniques have been applied to MPT datasets to predict the diffusion mode of nanoparticle trajectories as well as more complex biological variables, such age biological age. In this study, we develop a machine learning pipeline to predict and investigate changes to the brain ECS due to injury using supervised classification and feature importance calculations. We first validate the pipeline on three related but distinct MPT datasets from the living brain ECS - age differences, region differences, and enzymatic degradation of ECS structure. We predict three ages with 86% accuracy, three regions with 90% accuracy, and healthy versus enzyme-treated tissue with 69% accuracy. Since injury across groups is normally compared with traditional statistical approaches, we first used linear mixed effects models to compare features between healthy control conditions and injury induced by two different oxygen glucose deprivation [1] exposure times. We then used machine learning to predict injury state using MPT features. We show that the pipeline predicts between the healthy control, 0.5-hour OGD treatment, and 1.5-hour OGD treatment with 59% accuracy in the cortex and 66% in the striatum and identifies nonlinear relationships between trajectory features that were not evident from traditional linear models. Our work demonstrates that machine learning applied to MPT data is effective across multiple experimental conditions and can find unique biologically relevant features of nanoparticle diffusion.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142399236","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-09DOI: 10.1016/j.bpj.2024.10.006
Irene Sagrafena, Maxim Morin, Georgios Paraskevopoulos, Emelie J Nilsson, Iva Hrdinová, Andrej Kováčik, Sebastian Björklund, Kateřina Vávrová
Lipid membranes play a crucial role in regulating the body's water balance by adjusting their properties in response to hydration. The intercellular lipid matrix of the stratum corneum (SC), the outermost skin layer, serves as the body's primary defense against environmental factors. Osmolytes, including urocanic acid (UCA) and glycerol, are key components of the natural moisturizing factor that help the SC resist osmotic stress from dry environments. This study examines the effects of UCA and glycerol (each at 5 mol%) on isolated human SC lipids. For this, different techniques were employed, offering complementary information of the system's multiscale characteristics, including humidity-scanning quartz crystal microbalance with dissipation monitoring, infrared spectroscopy, X-ray diffraction, electrical impedance spectroscopy, and studies of water loss and permeability. Our results show that UCA increases water sorption and makes lipid films more liquid-like at high relative humidity, without significantly altering the lipid lamellar structure, chain order, or orthorhombic chain packing. Lipid films containing UCA exhibited higher water loss, significantly higher model drug permeability, and kinetically faster changes in electrical properties upon contact with aqueous solution compared to control lipids. These observations suggest that UCA reduces lipid cohesion in regions other than the acyl chain-rich leaflets, which may impact SC desquamation. In contrast, glycerol did not influence the hydration or permeability of the SC lipid matrix. However, it increased the proportion of orthorhombic domains at high humidities and slowed the kinetics of the hydration process, as evidenced by slower changes in the dielectric properties of the lipid film. These findings suggest that glycerol enhances lipid cohesion rather than increasing water uptake, which is typically the expected function of humectants. Consequently, UCA and glycerol appear to have distinct roles in maintaining epidermal homeostasis.
脂质膜可根据水合作用调整自身特性,在调节人体水分平衡方面发挥着至关重要的作用。皮肤最外层角质层(SC)的细胞间脂质基质是人体抵御环境因素的主要屏障。包括尿囊酸(UCA)和甘油在内的渗透溶解物是天然保湿因子的关键成分,可帮助角质层抵御干燥环境带来的渗透压力。本研究探讨了 UCA 和甘油(各为 5 摩尔%)对分离的人体 SC 脂质的影响。为此,我们采用了不同的技术,包括湿度扫描石英晶体微天平(带耗散监测)、红外光谱、X 射线衍射、电阻抗光谱以及失水和渗透性研究,从而为系统的多尺度特性提供互补信息。我们的研究结果表明,UCA 增加了水的吸附性,并使脂膜在高相对湿度下更像液体,而不会明显改变脂质的层状结构、链序或正交链堆积。与对照组脂质相比,含有 UCA 的脂质膜在与水溶液接触时会表现出更高的失水率、更高的模型药物渗透性和更快的电特性变化。这些观察结果表明,UCA 降低了酰基链丰富的小叶以外区域的脂质内聚力,这可能会影响 SC 脱膜。相比之下,甘油并不影响 SC 脂质基质的水合作用或渗透性。然而,甘油在高湿度条件下增加了正交菱形结构域的比例,并减缓了水合过程的动力学速度,这从脂膜介电性质的缓慢变化可以看出。这些研究结果表明,甘油能增强脂质的内聚力,而不是增加吸水性,后者通常是保湿剂的预期功能。因此,UCA 和甘油在维持表皮平衡方面似乎具有不同的作用。
{"title":"Structure and function of skin barrier lipids: Effects of hydration and natural moisturizers in vitro.","authors":"Irene Sagrafena, Maxim Morin, Georgios Paraskevopoulos, Emelie J Nilsson, Iva Hrdinová, Andrej Kováčik, Sebastian Björklund, Kateřina Vávrová","doi":"10.1016/j.bpj.2024.10.006","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.006","url":null,"abstract":"<p><p>Lipid membranes play a crucial role in regulating the body's water balance by adjusting their properties in response to hydration. The intercellular lipid matrix of the stratum corneum (SC), the outermost skin layer, serves as the body's primary defense against environmental factors. Osmolytes, including urocanic acid (UCA) and glycerol, are key components of the natural moisturizing factor that help the SC resist osmotic stress from dry environments. This study examines the effects of UCA and glycerol (each at 5 mol%) on isolated human SC lipids. For this, different techniques were employed, offering complementary information of the system's multiscale characteristics, including humidity-scanning quartz crystal microbalance with dissipation monitoring, infrared spectroscopy, X-ray diffraction, electrical impedance spectroscopy, and studies of water loss and permeability. Our results show that UCA increases water sorption and makes lipid films more liquid-like at high relative humidity, without significantly altering the lipid lamellar structure, chain order, or orthorhombic chain packing. Lipid films containing UCA exhibited higher water loss, significantly higher model drug permeability, and kinetically faster changes in electrical properties upon contact with aqueous solution compared to control lipids. These observations suggest that UCA reduces lipid cohesion in regions other than the acyl chain-rich leaflets, which may impact SC desquamation. In contrast, glycerol did not influence the hydration or permeability of the SC lipid matrix. However, it increased the proportion of orthorhombic domains at high humidities and slowed the kinetics of the hydration process, as evidenced by slower changes in the dielectric properties of the lipid film. These findings suggest that glycerol enhances lipid cohesion rather than increasing water uptake, which is typically the expected function of humectants. Consequently, UCA and glycerol appear to have distinct roles in maintaining epidermal homeostasis.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142399237","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-08DOI: 10.1016/j.bpj.2024.10.003
David Denberg, Xiaoxuan Zhang, Tomer Stern, Eric Wieschaus, Krishna Garikipati, Stanislav Y Shvartsman
Gastrulation is a critical process during embryonic development that transforms a single-layered blastula into a multi-layered embryo with distinct germ layers, which eventually give rise to all the tissues and organs of the organism. Studies across species have uncovered the mechanisms underlying the building blocks of gastrulation movements, such as localized in-plane and out-of-plane epithelial deformations. The next challenge is to understand dynamics on the scale of the embryo: this requires quantifying strain tensors, which rigorously describe the differences between the deformed configurations taken on by local clusters of cells at time instants of observation and their reference configuration at an initial time. We present a systematic strategy for computing such tensors from the local dynamics of cell clusters, which are chosen across the embryo from several regions whose morphogenetic fate is central to viable gastrulation. As an application of our approach, we demonstrate a strategy of identifying distinct Drosophila morphological domains using strain tensors.
{"title":"Computing Whole Embryo Strain Maps During Gastrulation.","authors":"David Denberg, Xiaoxuan Zhang, Tomer Stern, Eric Wieschaus, Krishna Garikipati, Stanislav Y Shvartsman","doi":"10.1016/j.bpj.2024.10.003","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.10.003","url":null,"abstract":"<p><p>Gastrulation is a critical process during embryonic development that transforms a single-layered blastula into a multi-layered embryo with distinct germ layers, which eventually give rise to all the tissues and organs of the organism. Studies across species have uncovered the mechanisms underlying the building blocks of gastrulation movements, such as localized in-plane and out-of-plane epithelial deformations. The next challenge is to understand dynamics on the scale of the embryo: this requires quantifying strain tensors, which rigorously describe the differences between the deformed configurations taken on by local clusters of cells at time instants of observation and their reference configuration at an initial time. We present a systematic strategy for computing such tensors from the local dynamics of cell clusters, which are chosen across the embryo from several regions whose morphogenetic fate is central to viable gastrulation. As an application of our approach, we demonstrate a strategy of identifying distinct Drosophila morphological domains using strain tensors.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142387631","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-04DOI: 10.1016/j.bpj.2024.10.001
Luca Codutti, John P Kirkpatrick, Susanne Zur Lage, Teresa Carlomagno
DEAD-box helicases use ATP to unwind short double-stranded RNA (dsRNA). The helicase core consists of two discrete domains, termed RecA_N and RecA_C. The nucleotide binding site is harbored in RecA_N, while both RecA_N and RecA_C are involved in RNA recognition and ATP hydrolysis. In the absence of nucleotides or RNA, RecA_N and RecA_C do not interact ("open" form of the enzyme). In the presence of both RNA and ATP the two domains come together ("closed" form), building the composite RNA binding site and stimulating ATP hydrolysis. Because of the different roles and thermodynamic properties of the ADP-bound and ATP-bound states in the catalytic cycle, the conformations of DEAD-box helicases in complex with ATP and ADP are assumed to be different. However, the available crystal structures do not recapitulate these supposed differences and show identical conformations of DEAD-box helicases independent of the identity of the bound nucleotide. Here, we use NMR to demonstrate that the conformations of the ATP- and ADP-bound forms of the DEAD-box helicase Vasa are indeed different, contrary to the results from x-ray crystallography. These differences do not relate to the populations of the open and closed forms, but are intrinsic to the RecA_N domain. NMR chemical shift analysis reveals the regions of RecA_N where the average conformations of Vasa-ADP and Vasa-ATP are most different and indicates that these differences may contribute to modulating the affinity of the two nucleotide-bound complexes for RNA substrates.
DEAD-box 螺旋酶利用 ATP 解旋短双链 RNA(dsRNA)。螺旋酶核心由两个不同的结构域组成,分别称为 RecA_N 和 RecA_C。核苷酸结合位点位于 RecA_N,而 RecA_N 和 RecA_C 都参与 RNA 识别和 ATP 水解。在没有核苷酸或 RNA 的情况下,RecA_N 和 RecA_C 不发生相互作用(酶的 "开放 "形式)。在有 RNA 和 ATP 的情况下,这两个结构域会结合在一起("封闭 "形式),形成复合 RNA 结合位点并促进 ATP 的水解。由于 ADP 结合态和 ATP 结合态在催化循环中的作用和热力学性质不同,因此假定 DEAD-box 螺旋酶与 ATP 和 ADP 复合物的构象也不同。然而,现有的晶体结构并没有再现这些假定的差异,而是显示了 DEAD-box 螺旋酶的相同构象,与结合核苷酸的身份无关。在这里,我们利用核磁共振技术证明了与 ATP 和 ADP 结合的 DEAD-box 螺旋酶 Vasa 的构象确实不同,这与 X 射线晶体学的结果相反。这些差异与开放型和封闭型的群体无关,而是 RecA_N 结构域的固有差异。核磁共振化学位移分析揭示了 RecA_N 中 Vasa-ADP 和 Vasa-ATP 平均构象差异最大的区域,并表明这些差异可能有助于调节这两种核苷酸结合复合物对 RNA 底物的亲和力。
{"title":"Long-range conformational changes in the nucleotide-bound states of the DEAD-box helicase Vasa.","authors":"Luca Codutti, John P Kirkpatrick, Susanne Zur Lage, Teresa Carlomagno","doi":"10.1016/j.bpj.2024.10.001","DOIUrl":"10.1016/j.bpj.2024.10.001","url":null,"abstract":"<p><p>DEAD-box helicases use ATP to unwind short double-stranded RNA (dsRNA). The helicase core consists of two discrete domains, termed RecA_N and RecA_C. The nucleotide binding site is harbored in RecA_N, while both RecA_N and RecA_C are involved in RNA recognition and ATP hydrolysis. In the absence of nucleotides or RNA, RecA_N and RecA_C do not interact (\"open\" form of the enzyme). In the presence of both RNA and ATP the two domains come together (\"closed\" form), building the composite RNA binding site and stimulating ATP hydrolysis. Because of the different roles and thermodynamic properties of the ADP-bound and ATP-bound states in the catalytic cycle, the conformations of DEAD-box helicases in complex with ATP and ADP are assumed to be different. However, the available crystal structures do not recapitulate these supposed differences and show identical conformations of DEAD-box helicases independent of the identity of the bound nucleotide. Here, we use NMR to demonstrate that the conformations of the ATP- and ADP-bound forms of the DEAD-box helicase Vasa are indeed different, contrary to the results from x-ray crystallography. These differences do not relate to the populations of the open and closed forms, but are intrinsic to the RecA_N domain. NMR chemical shift analysis reveals the regions of RecA_N where the average conformations of Vasa-ADP and Vasa-ATP are most different and indicates that these differences may contribute to modulating the affinity of the two nucleotide-bound complexes for RNA substrates.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142375032","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-04DOI: 10.1016/j.bpj.2024.09.029
Sophia P Hirakis, Thomas M Bartol, Ludovic Autin, Rommie E Amaro, Terrence J Sejnowski
We present the first-ever, fully-discrete, stochastic model of triggered cardiac calcium dynamics. Using anatomically accurate subcellular cardiac myocyte geometries, we simulate the molecular players involved in calcium handling using high-resolution stochastic and explicit-particle methods at the level of an individual cardiac dyadic junction. Integrating data from multiple experimental sources, the model not only replicates the findings of traditional in silico studies and complements in vitro experimental data, but also reveals new insights into the molecular mechanisms driving cardiac dysfunction under stress and disease conditions. We improve upon older, non-discrete models using the same realistic geometry by incorporating molecular mechanisms for spontaneous, as well as triggered Calcium-Induced Calcium Release (CICR). Action potentials are used to activate L-type calcium channels (LTCCs), triggering CICR through Ryanodine receptors (RyR) on the surface of the sarcoplasmic reticulum. These improvements allow for the specific focus on the couplon: the structure-function relationship between LTCC and RyR. We investigate the electrophysical effects of normal and diseased action potentials on CICR and interrogate the effects of dyadic junction deformation through detubulation and orphaning of RyR. Our work demonstrates the importance of the electrophysical integrity of the CRU on CICR fidelity, giving insights into the molecular basis of heart disease. Finally, we provide a unique, detailed, molecular view of the CICR process using advanced rendering techniques. This easy-to-use model comes complete with tutorials and all necessary software for use and analysis so as to maximize usability and reproducibility. Our work focuses on quantifying, qualifying, and visualizing the behavior of the molecular species that underlie the function and dysfunction of subcellular cardiomyocyte systems.
{"title":"Electrophysical cardiac remodeling at the molecular level: insights into Ryanodine Receptor activation and calcium-induced calcium release from a stochastic explicit-particle model.","authors":"Sophia P Hirakis, Thomas M Bartol, Ludovic Autin, Rommie E Amaro, Terrence J Sejnowski","doi":"10.1016/j.bpj.2024.09.029","DOIUrl":"https://doi.org/10.1016/j.bpj.2024.09.029","url":null,"abstract":"<p><p>We present the first-ever, fully-discrete, stochastic model of triggered cardiac calcium dynamics. Using anatomically accurate subcellular cardiac myocyte geometries, we simulate the molecular players involved in calcium handling using high-resolution stochastic and explicit-particle methods at the level of an individual cardiac dyadic junction. Integrating data from multiple experimental sources, the model not only replicates the findings of traditional in silico studies and complements in vitro experimental data, but also reveals new insights into the molecular mechanisms driving cardiac dysfunction under stress and disease conditions. We improve upon older, non-discrete models using the same realistic geometry by incorporating molecular mechanisms for spontaneous, as well as triggered Calcium-Induced Calcium Release (CICR). Action potentials are used to activate L-type calcium channels (LTCCs), triggering CICR through Ryanodine receptors (RyR) on the surface of the sarcoplasmic reticulum. These improvements allow for the specific focus on the couplon: the structure-function relationship between LTCC and RyR. We investigate the electrophysical effects of normal and diseased action potentials on CICR and interrogate the effects of dyadic junction deformation through detubulation and orphaning of RyR. Our work demonstrates the importance of the electrophysical integrity of the CRU on CICR fidelity, giving insights into the molecular basis of heart disease. Finally, we provide a unique, detailed, molecular view of the CICR process using advanced rendering techniques. This easy-to-use model comes complete with tutorials and all necessary software for use and analysis so as to maximize usability and reproducibility. Our work focuses on quantifying, qualifying, and visualizing the behavior of the molecular species that underlie the function and dysfunction of subcellular cardiomyocyte systems.</p>","PeriodicalId":8922,"journal":{"name":"Biophysical journal","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142378995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}