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The following table provides information about the project.
WEIZMANN INSTITUTE OF SCIENCE
|Coordinator Country||Israel [IL]|
|Total cost||1˙500˙000 €|
|EC max contribution||1˙500˙000 € (100%)|
1. H2020-EU.1.1. (EXCELLENT SCIENCE - European Research Council (ERC))
|Duration (year-month-day)||from 2019-11-01 to 2024-10-31|
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|1||WEIZMANN INSTITUTE OF SCIENCE||IL (REHOVOT)||coordinator||1˙500˙000.00|
Cell-to-cell fusion is a ubiquitous phenomenon essential for the physiological function of numerous tissues. A striking example is the fusion of myoblasts to form multinucleated myofibers during skeletal muscle development and regeneration. During myoblast fusion, membrane architecture must be radically remodeled. Yet, how membrane remodeling occurs on the molecular level is poorly understood as, until now, there was no approach available for visualizing dynamic changes in the cellular ultrastructure and the organization of the fusion machinery in situ. To fill this gap, we have developed correlative light and 3D electron microscopy (CLEM) methods that allow us to identify fluorescent signals within EM samples with high sensitivity and subsequently localize the source of these signals with high precision. In this proposal, we will apply these methods in combination with live-cell imaging, biochemistry and cryo-electron tomography (ET) to deliver fundamental knowledge about the mechanism of myoblast fusion. Our specific aims are: Aim 1: To resolve the molecular and ultrastructural events underlying cell fusion, by revealing how plasma membrane architecture is remodeled at sites of fusion using 3D EM. Aim 2: To dissect the mechanism driving membrane remodeling during fusion, by visualizing how the fusion machinery assembles at sites of fusion and how its assembly is mirrored by changes in membrane shape, using biochemistry and live-cell imaging. Aim 3: To determine the structure of the fusion machinery in situ, by using cryo-ET and subtomogram averaging. Our synergetic experimental strategy will generate a quantitative, dynamic high-resolution view of the fusogenic synapse of vertebrate muscle, revealing how the fusion machinery remodels the plasma membrane at sites of fusion. These data are vital for deriving a biophysical model of myoblast fusion, understanding the general mechanism of cell fusion, and developing strategies to treat primary muscle diseases.
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