Dataset Information

Direct imaging of liquid domains in membranes by cryo-electron tomography.

ABSTRACT: Images of micrometer-scale domains in lipid bilayers have provided the gold standard of model-free evidence to understand the domains' shapes, sizes, and distributions. Corresponding techniques to directly and quantitatively assess smaller (nanoscale and submicron) liquid domains have been limited. Researchers commonly seek to correlate activities of membrane proteins with attributes of the domains in which they reside; doing so hinges on identification and characterization of membrane domains. Although some features of membrane domains can be probed by indirect methods, these methods are often constrained by the limitation that data must be analyzed in the context of models that require multiple assumptions or parameters. Here, we address this challenge by developing and testing two methods of identifying submicron domains in biomimetic membranes. Both methods leverage cryo-electron tomograms of ternary membranes under vitrified, hydrated conditions. The first method is optimized for probe-free applications: Domains are directly distinguished from the surrounding membrane by their thickness. This technique quantitatively and accurately measures area fractions of domains, in excellent agreement with known phase diagrams. The second method is optimized for applications in which a single label is deployed for imaging membranes by both high-resolution cryo-electron tomography and diffraction-limited optical microscopy. For this method, we test a panel of probes, find that a trimeric mCherry label performs best, and specify criteria for developing future high-performance, dual-use probes. These developments have led to direct and quantitative imaging of submicron membrane domains in vitrified, hydrated vesicles.

SUBMITTER: Cornell CE

PROVIDER: S-EPMC7443872 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Direct imaging of liquid domains in membranes by cryo-electron tomography.

Cornell Caitlin E CE Mileant Alexander A Thakkar Niket N Lee Kelly K KK Keller Sarah L SL

Proceedings of the National Academy of Sciences of the United States of America 20200805 33

Images of micrometer-scale domains in lipid bilayers have provided the gold standard of model-free evidence to understand the domains' shapes, sizes, and distributions. Corresponding techniques to directly and quantitatively assess smaller (nanoscale and submicron) liquid domains have been limited. Researchers commonly seek to correlate activities of membrane proteins with attributes of the domains in which they reside; doing so hinges on identification and characterization of membrane domains. ...[more]

PMID: 32759217

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Dataset Information

Direct imaging of liquid domains in membranes by cryo-electron tomography.

Publications

Direct imaging of liquid domains in membranes by cryo-electron tomography.

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