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SHIELD

Financiado

To Shield, or not to Shield: deciphering DNA repair pathway choice after CRISPR-...

To Shield, or not to Shield: deciphering DNA repair pathway choice after CRISPR-Cas cleavage Genome editing has triggered a revolution with stark implications in life science. Recently, a new gene-editing technique CRISPR-Cas has become dominant in laboratory conditions. The first step in a targeted genome modification re... ver más

31/08/2024

UCPH

231K€

Presupuesto del proyecto: 231K€

Líder del proyecto

KOBENHAVNS UNIVERSITET No se ha especificado una descripción o un objeto social para esta compañía.

TRL 4-5

Ver 1 Participantes

Financiación concedida El organismo HORIZON EUROPE notifico la concesión del proyecto el día 2024-08-31

Línea de financiación objetivo El proyecto se financió a través de la siguiente ayuda:

HORIZON-MSCA-2021-PF-01-01: MSCA Postdoctoral Fellowships 2021

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UCPH

230.77K€ | Lider

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Duración del proyecto: 27 meses Fecha Inicio: 2022-05-25
Fecha Fin: 2024-08-31

Líder del proyecto

KOBENHAVNS UNIVERSITET

TRL 4-5

Presupuesto del proyecto 231K€

Descripción del proyecto Genome editing has triggered a revolution with stark implications in life science. Recently, a new gene-editing technique CRISPR-Cas has become dominant in laboratory conditions. The first step in a targeted genome modification requires the CRISPR-Cas nuclease to generate a specific DNA double-strand break (DSB). Eukaryotic cells repair the DSBs by the fast but potentially mutagenic Non-Homologous End Joining (NHEJ), and by the accurate Homology Directed Repair (HDR) pathways. Gene editing techniques exploit the HDR to modify the DNA to the desired sequence. Usually, it is the NHEJ that repairs the DSBs. That presents a threat of introducing mutations and limits gene editing efficiency and use in medical applications. Moreover, even after entering the HDR pathway, an unclear mechanism involving protein 53BP1 and Shieldin complex blocks the pathway from progressing. My goal is to decipher the 53BP1-Shieldin induced block and then assess Shieldin pathway as a possible therapeutic target to increase the frequency of HDR after CRISPR-Cas cleavage. The objectives include: 1. Reconstitution of the Shieldin pathway in mammalian HEK293 and insect Hi5 cells 2. Structural characterization of the protein assembly at DSBs by electron microscopy and mass spectrometry 3. Biochemical analysis of the relation between DSB repair and Shieldin pathway In general, the findings can be applied to enhance any gene-editing technique involving a generation of DSBs. Importantly, the results will provide a base for a long-awaited revolution in personalized medicine. The host laboratory of Prof. Montoya already focuses on improving the properties of the Cas nucleases by means of structural biology and the Shieldin project fits right in their research portfolio. Developing the project will boost my career as new skills in structural biology, including protein electron microscopy, will complement my previous expertise in protein-protein interaction and crosslinking mass spectrometry studies.

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