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Dissecting gamma-TuRC composition and activity by Single Molecule Pull-down

Objective

In this project I will use Single Molecule Pull-down (SiMPull) for studying activity and regulation of human gamma-tubulin
ring complexes (gamma-TuRCs). Gamma-TuRCs are the main nucleators of microtubule (MT) polymerization. Additionally, gamma-TuRCs may play a role in modulating MT dynamics. Regulation of gamma-TuRC activity is key to organizing the dynamic MT arrays needed for essential processes in various cell types. Progress in understanding gamma-TuRC regulation at the molecular level is currently hampered by a lack of information about subunit stoichiometries and interactions, and by technical difficulties that have prevented reconstitution of gamma-TuRCs in vitro. I will tackle these challenges by using SiMPull to immunoprecipitate and immobilize gamma-TuRCs to a glass surface directly from cell extracts collected at different cell cycle stages, and analyze individual gamma-TuRCs using high resolution microscopy. I will express tagged gamma-TuRC subunits in cells to isolate wildtype and mutant gamma-TuRCs. Using fluorescent protein tags and antibodies I will visualize and quantify components in individual gamma-TuRCs and, by incubation in pure tubulin and GTP, determine how gamma-TuRC composition is related to its ability to nucleate MTs and modulate MT dynamics. Together, the proposed research will expand the field’s toolbox, thereby allowing new, molecular level insight into gamma-TuRC activity and its regulation.

Fields of science (EuroSciVoc)

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Topic(s)

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Funding Scheme

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MSCA-IF-EF-ST - Standard EF

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Call for proposal

Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.

(opens in new window) H2020-MSCA-IF-2015

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Coordinator

FUNDACIO INSTITUT DE RECERCA BIOMEDICA (IRB BARCELONA)
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.

€ 158 121,60
Address
CARRER BALDIRI REIXAC 10-12 PARC SCIENTIFIC DE BARCELONA
08028 Barcelona
Spain

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Region
Este Cataluña Barcelona
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.

€ 158 121,60
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