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Quantitation and Analysis of the Formation of HO-Endonuclease Stimulated Chromosomal Translocations by Single-Strand Annealing in Saccharomyces cerevisiae
Published on: September 23, 2011
La remodelación de la cromatina en un sitio de ruptura de doble hebra de ADN en Saccharomyces cerevisiae
Toyoko Tsukuda1, Alastair B Fleming, Jac A Nickoloff
1Department of Molecular Genetics and Microbiology, University of New Mexico School of Medicine, 915 Camino de Salud Albuquerque, New Mexico 87131, USA.
Nature
|November 18, 2005
Resumen
Las rupturas de la doble hebra del ADN desencadenan modificaciones de histonas y pérdida de nucleosomas, cruciales para la estabilidad del genoma. El complejo MRX y el remodelador INO80 orquestan estos cambios en la cromatina, facilitando el reclutamiento de proteínas de reparación del ADN.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- Genética La genética.
- Biología celular Biología celular.
Sus antecedentes:
- Las rupturas de doble hebra de ADN (DSB) amenazan la estabilidad del genoma y se reparan a través de la unión de extremos no homólogos y la recombinación homóloga.
- Las alteraciones de la cromatina después de los DSB son críticas para una reparación eficiente, que involucra modificaciones de histonas como la fosforilación H2A.X.
Objetivo del estudio:
- Para investigar los cambios estructurales de la cromatina cerca de los DSB en Saccharomyces cerevisiae.
- Para aclarar los roles de los complejos de proteínas específicas en la mediación de estas alteraciones de la cromatina y su impacto en la reparación del ADN.
Principales métodos:
- Análisis de las modificaciones histónicas (fosforilación H2A) y la pérdida de nucleosomas utilizando la digestión de la nucleasa del micrococo.
- Investigando el requisito del sensor de daño al ADN MRX y el complejo de remodelación de nucleosomas INO80 para la pérdida de histonas.
- Evaluación de la dinámica de reclutamiento de la proteína de reparación Rad51.
Principales resultados:
- La formación de DSB induce la fosforilación de H2A, seguida de la pérdida de H2B y H3, aumentando la accesibilidad de la cromatina.
- La pérdida de histona y la fosforilación H2A ocurren independientemente y requieren el complejo MRX e INO80.
- La ausencia de pérdida de histonas retrasa el reclutamiento de Rad51, lo que indica que la remodelación del nucleosoma mediada por MRX regula el acceso al factor de recombinación homólogo.
Conclusiones:
- El complejo MRX coordina distintas vías de modificación de la cromatina: desplazamiento de los nucleosomas para el reclutamiento de proteínas de recombinación homólogas y fosforilación H2A para la señalización de puntos de control.
- La remodelación del nucleosoma dependiente de MRX es esencial para el acceso eficiente de los factores de reparación a los DSB, destacando un paso regulador clave en la recombinación homóloga.
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