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Reconstitution of Nucleosomes with Differentially Isotope-labeled Sister Histones
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Complejo activado RSC-nucleosoma y forma alterada persistente del nucleosoma
1Department of Structural Biology, Stanford University School of Medicine, California 94305, USA.
Cell
|July 23, 1998
Resumen
El remodelador de cromatina RSC altera los nucleosomas de una manera dependiente del ATP, aumentando su susceptibilidad a las nucleasas mientras retiene las histonas. Esta remodelación de la cromatina es reversible, destacando el RSC.
Área de la Ciencia:
- Biología de la cromatina Biología de la cromatina
- Los mecanismos moleculares de la reparación del ADN.
- Epigenética y regulación génica.
Sus antecedentes:
- El complejo RSC es un remodelador crucial de la cromatina involucrado en la accesibilidad del ADN.
- Comprender la interacción de RSC con los nucleosomas es clave para aclarar su papel en la dinámica de la cromatina.
Objetivo del estudio:
- Investigar las consecuencias funcionales de la unión de RSC a los nucleosomas.
- Para caracterizar los cambios conformacionales dependientes de ATP inducidos por RSC en los nucleosomas.
- Para determinar la estabilidad y reversibilidad de las alteraciones nucleosómicas mediadas por RSC.
Principales métodos:
- Análisis de desplazamiento de gel para evaluar las afinidades de unión RSC-nucleosoma.
- Ensayos enzimáticos que utilizan endo- y exonucleasas para evaluar la susceptibilidad del nucleosoma.
- Análisis de la movilidad electroforética y la tasa de sedimentación para caracterizar los cambios estructurales del nucleosoma.
- Ensayos de estabilidad dependientes de la fuerza iónica.
Principales resultados:
- El RSC se une a los nucleosomas y al ADN desnudo con afinidades similares.
- La unión de ATP induce un cambio conformacional en el complejo RSC-nucleosoma, aumentando la susceptibilidad de la nucleasa.
- El complemento histónico se conserva completamente durante todo el proceso de remodelación.
- La forma alterada del nucleosoma es inestable a altas fuerzas iónicas, pero puede ser revertida al nucleosoma original por RSC y ATP.
Conclusiones:
- La unión de RSC y la hidrólisis de ATP inducen una alteración transitoria, pero significativa, en la estructura del nucleosoma.
- Este evento de remodelación mejora la accesibilidad del ADN para la reparación o maquinaria de transcripción.
- La naturaleza reversible de la reacción sugiere un papel dinámico para el RSC en la regulación de la cromatina.
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Nucleosomes, paradoxically, perform two opposite functions simultaneously. On the one hand, their primary aim is to protect the delicate DNA strands from physical damage and help achieve a higher compaction ratio. On the other hand, they must allow polymerase enzymes to access histone-bound DNA during...
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