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Rapid Analysis of Circadian Phenotypes in Arabidopsis Protoplasts Transfected with a Luminescent Clock Reporter
Published on: September 17, 2016
Identificación de RACK1 y de la proteína quinasa Calpha como componentes integrales del reloj circadiano de los
Maria S Robles1, Cyril Boyault, Darko Knutti
1Department of Neurobiology, Harvard Medical School, Boston, MA 02115, USA.
Science (New York, N.Y.)
|January 23, 2010
Resumen
El reloj circadiano.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- Cronobiología cronobiología.
- La bioquímica es la bioquímica.
Sus antecedentes:
- El reloj circadiano de los mamíferos se basa en un circuito de retroalimentación negativa que involucra al factor de transcripción CLOCK-BMAL1.
- Comprender los mecanismos moleculares de este bucle de retroalimentación es crucial para descifrar la regulación del ritmo circadiano.
Objetivo del estudio:
- Para investigar los complejos proteicos asociados con BMAL1 durante la fase de retroalimentación circadiana.
- Para aclarar el papel de los receptores para la cinasa C-1 activada (RACK1) y la proteína kinasa C-alfa (PKCalpha) en el reloj circadiano de los mamíferos.
Principales métodos:
- Análisis de los complejos de proteínas nucleares de ratón que contienen BMAL1.1.
- Estudios de sobreexpresión y agotamiento de RACK1 y PKCalpha en fibroblastos.
- Ensayos de fosforilación in vitro para evaluar la modificación de BMAL1 por PKCalpha.
Principales resultados:
- RACK1 y PKCalpha fueron identificados de manera circadiana dentro de un complejo nuclear BMAL1.
- La sobreexpresión de RACK1 y PKCalpha inhibió la actividad transcripcional de CLOCK-BMAL1.
- RACK1 mejoró la fosforilación mediada por PKCalpha de BMAL1 in vitro.
- El agotamiento de RACK1 o PKCalpha dio lugar a un período circadiano acortado en los fibroblastos.
Conclusiones:
- La vía de señalización de la proteína quinasa C (PKC) está regulada dinámicamente dentro del reloj circadiano.
- RACK1 y PKCalpha son componentes integrales del bucle de retroalimentación circadiana de los mamíferos, que influyen en las oscilaciones del reloj.
- La vía de la PKC es activada rítmicamente por procesos celulares internos, no sólo por estímulos externos, contribuyendo a la ritmicidad circadiana.
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