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Reconstructing the 4D space of intracellular lipid phase separation and lipid droplet biogenesis

Project description

Advanced cryo-electron microscopy offers a molecular level view of lipid droplet biogenesis

Lipid droplets are cellular organelles that regulate the storage and hydrolysis of neutral lipids and are central for cellular metabolism. Funded by the Marie Skłodowska-Curie Actions programme, the 4D lipid phase project aims to expand our understanding of lipid droplet biogenesis at a molecular level. To this end, it will develop novel cryo-correlative light and electron microscopy methods to construct 3D structural models that represent the timeline of lipid droplet biogenesis in human cells. Imaging of the early stages of lipid droplet formation at unprecedented resolution will provide new insight into the molecular mechanisms of intracellular lipid storage.

Objective

Lipid droplets (LDs) are neutral lipid (NL) storage organelles, central for cellular metabolism. Current models of LD biogenesis postulate that synthesis of NLs in the ER bilayer favors their phase separation, followed by bending of the bilayer and budding of the LD into the cytoplasm, coordinated by a specific, not fully characterized, molecular machinery. Understanding of LD biogenesis through direct observations at molecular resolution is critically lacking. A major obstacle is that conventional electron microscopy techniques, lending the required resolving power, do not preserve the detailed membrane architecture of the ER and the fine structure of the macromolecular machinery driving LD biogenesis.

I propose an ambitious cryo-correlative light and electron microscopy (CLEM) study to construct 3D structural models representing the timeline of LD biogenesis in human cells. Using a combination of 4 key proteins tagged with fluorescent markers in cells stimulated towards LD biogenesis, I will stage and image all early events. As nascent LD intermediates are a priori nano-scale structures dispersed within the ER, accurate cryo-CLEM is crucial. I will address this technical challenge with two strategies: cryo-superresolution CLEM and genetically-coded multimeric nanoparticles as markers for cryo-EM. This project harnesses state-of-the-art methods to synergize physical chemistry, structural and cell biology. The high resolution and pristine structural preservation exclusively attainable with in situ cryo-EM will provide first direct observations of early LD assembly, allowing construction of comprehensive 4D models of intracellular lipid phase separation and LD biogenesis.

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MSCA-IF - Marie Skłodowska-Curie Individual Fellowships (IF)

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Call for proposal

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(opens in new window) H2020-MSCA-IF-2020

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Coordinator

EUROPEAN MOLECULAR BIOLOGY LABORATORY
Net EU contribution

Net EU financial contribution. The sum of money that the participant receives, deducted by the EU contribution to its linked third party. It considers the distribution of the EU financial contribution between direct beneficiaries of the project and other types of participants, like third-party participants.

€ 162 806,40
Address
Meyerhofstrasse 1
69117 Heidelberg
Germany

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Region
Baden-Württemberg Karlsruhe Heidelberg, Stadtkreis
Activity type
Research Organisations
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Total cost

The total costs incurred by this organisation to participate in the project, including direct and indirect costs. This amount is a subset of the overall project budget.

€ 162 806,40
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