Resolution doubling using confocal microscopy via analogy with structured illumination microscopy

Hayashi, S.

doi:10.7567/jjap.55.082501

Cited by 19 publications

(9 citation statements)

References 27 publications

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“…For each lipid composition, tubules with both pearled and undulating morphologies were observed, Figure 4a-d. Further, the data were reasonably well fit by a curve in which tubule diameter was proportional to the square root of bending rigidity, in agreement with the predictions of the simulation, compare Figure 4f, Figure 3e, g. Here optical reassignment during spinning disc confocal microscopy, followed by deconvolution was used to increase the optical resolution to better than 150 nm (31).…”

Section: Tubule Diameter Varies With Membrane Bending Rigidity and Sasupporting

confidence: 82%

Membrane bending by protein phase separation

Feng

Alimohamadi

Bakka

et al. 2020

Preprint

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Membrane bending is a ubiquitous cellular process that is required for membrane traffic, cell motility, organelle biogenesis, and cell division. Proteins that bind to membranes using specific structural features, such as wedge-like amphipathic helices and crescentshaped scaffolds, are thought to be the primary drivers of membrane bending. However, many membrane-binding proteins have substantial regions of intrinsic disorder, which lack a stable three-dimensional structure. Interestingly, many of these disordered domains have recently been found to form networks stabilized by weak, multi-valent contacts, leading to assembly of protein liquid phases on membrane surfaces. Here we ask how membrane-associated protein liquids impact membrane curvature. Using synthetic and cell-derived membrane vesicles, we find that liquid-liquid phase separation of membrane-associated disordered proteins creates a substantial compressive stress in the plane of the membrane. This stress drives the membrane to bend away from the protein layer, creating protein-lined membrane tubules. A simple mechanical model of this process accurately predicts the experimentally measured relationship between the rigidity of the membrane and the diameter of the membrane tubules. Discovery of this mechanism, which may be relevant to a broad range of membrane protrusions, illustrates that membrane remodeling is not exclusive to structured scaffolds, but can also be driven by the rapidly emerging class of liquid-like protein networks that assemble at membranes.

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Section: Tubule Diameter Varies With Membrane Bending Rigidity and Sasupporting

confidence: 82%

Membrane bending by protein phase separation

Feng

Alimohamadi

Bakka

et al. 2020

Preprint

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“…The field is already addressing some of these issues such as using reactive oxygen species (ROS) scavenging buffers ( 168 , 169 , 171 ) and also minimizing pre-imaging stress of cells, by limiting overexpression of tag proteins and titration of fluorescent dyes ( 165 , 172 ). Indeed even simple adjustments such as limiting laser intensities ( 173 ), performing microscopy under optimal cell culture conditions ( 165 , 172 ) and the use of far-red excitation wavelengths can all reduce photo-damage ( 168 ), especially when then combined with increased scanning speed ( 138 , 172 , 174 ). These technological limitations will continue to co-develop with organoid research being addressed as and when required.…”

Section: Discussionmentioning

confidence: 99%

Liver Organoids: Recent Developments, Limitations and Potential

et al. 2021

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Liver cell types derived from induced pluripotent stem cells (iPSCs) share the potential to investigate development, toxicity, as well as genetic and infectious disease in ways currently limited by the availability of primary tissue. With the added advantage of patient specificity, which can play a role in all of these areas. Many iPSC differentiation protocols focus on 3 dimensional (3D) or organotypic differentiation, as these offer the advantage of more closely mimicking in vivo systems including; the formation of tissue like architecture and interactions/crosstalk between different cell types. Ultimately such models have the potential to be used clinically and either with or more aptly, in place of animal models. Along with the development of organotypic and micro-tissue models, there will be a need to co-develop imaging technologies to enable their visualization. A variety of liver models termed “organoids” have been reported in the literature ranging from simple spheres or cysts of a single cell type, usually hepatocytes, to those containing multiple cell types combined during the differentiation process such as hepatic stellate cells, endothelial cells, and mesenchymal cells, often leading to an improved hepatic phenotype. These allow specific functions or readouts to be examined such as drug metabolism, protein secretion or an improved phenotype, but because of their relative simplicity they lack the flexibility and general applicability of ex vivo tissue culture. In the liver field these are more often constructed rather than developed together organotypically as seen in other organoid models such as brain, kidney, lung and intestine. Having access to organotypic liver like surrogates containing multiple cell types with in vivo like interactions/architecture, would provide vastly improved models for disease, toxicity and drug development, combining disciplines such as microfluidic chip technology with organoids and ultimately paving the way to new therapies.

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“…4F and3 E and G). Here, optical reassignment during spinning disk confocal microscopy, followed by deconvolution, was used to increase the optical resolution to better than 150 nm (33).…”

Section: Tubule Diameter Varies With Membrane Bending Rigidity and Saltmentioning

confidence: 99%

Membrane bending by protein phase separation

Feng

Alimohamadi

Bakka

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

178

156

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Membrane bending is a ubiquitous cellular process that is required for membrane traffic, cell motility, organelle biogenesis, and cell division. Proteins that bind to membranes using specific structural features, such as wedge-like amphipathic helices and crescent-shaped scaffolds, are thought to be the primary drivers of membrane bending. However, many membrane-binding proteins have substantial regions of intrinsic disorder which lack a stable three-dimensional structure. Interestingly, many of these disordered domains have recently been found to form networks stabilized by weak, multivalent contacts, leading to assembly of protein liquid phases on membrane surfaces. Here we ask how membrane-associated protein liquids impact membrane curvature. We find that protein phase separation on the surfaces of synthetic and cell-derived membrane vesicles creates a substantial compressive stress in the plane of the membrane. This stress drives the membrane to bend inward, creating protein-lined membrane tubules. A simple mechanical model of this process accurately predicts the experimentally measured relationship between the rigidity of the membrane and the diameter of the membrane tubules. Discovery of this mechanism, which may be relevant to a broad range of cellular protrusions, illustrates that membrane remodeling is not exclusive to structured scaffolds but can also be driven by the rapidly emerging class of liquid-like protein networks that assemble at membranes.

show abstract

Resolution doubling using confocal microscopy via analogy with structured illumination microscopy

Cited by 19 publications

References 27 publications

Membrane bending by protein phase separation

Membrane bending by protein phase separation

Liver Organoids: Recent Developments, Limitations and Potential

Membrane bending by protein phase separation

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