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El silenciamiento de genes transcripcionales inducidos por ARN pequeño interferente en células humanas
Kevin V Morris1, Simon W-L Chan, Steven E Jacobsen
1Department of Medicine 0678, Stein Clinical Research Building, Room 302, University of California-San Diego, La Jolla, CA 92093-0678, USA. kmorris@coh.org
Science (New York, N.Y.)
|August 7, 2004
Resumen
El pequeño ARN interferente (siRNA) puede silenciar la transcripción génica en las células de mamíferos. Este proceso implica la metilación del ADN y requiere la entrada nuclear de siRNA, lo que demuestra mecanismos de silenciamiento de genes conservados.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- La epigenética es la epigenética.
- Interferencia en el ARN Interferencia en el ARN
Sus antecedentes:
- Se sabe que el ARN pequeño interferente (siRNA) y el microRNA regulan la expresión génica en múltiples niveles.
- Los mecanismos de silenciamiento de genes son cruciales para la función y el desarrollo celular.
Objetivo del estudio:
- Investigar el potencial del siRNA dirigido por el promotor para el silenciamiento de genes transcripcionales en células de mamíferos.
- Determinar los mecanismos involucrados en el silenciamiento transcripcional mediado por siRNA, incluida la metilación del ADN y el transporte nuclear.
Principales métodos:
- Se utilizan células de cultivo de tejidos humanos con un factor de alargamiento integrado 1alfa (EF1A) promotor-gen reportero de proteína fluorescente verde.
- Se aplicó siRNA dirigido al promotor para atacar tanto los genes EF1A reporteros como los endógenos.
- Investigó el papel de la metilación del ADN y las vías de transporte nuclear (transporte activo o transducción lentiviral) en el proceso de silenciamiento.
Principales resultados:
- El siRNA dirigido por el promotor inhibió con éxito la transcripción del gen reportero impulsado por el promotor EF1A.
- La transcripción endógena del gen EF1A también fue suprimida por el siRNA.
- El silenciamiento de la transcripción se correlacionó con un aumento de la metilación del ADN de la secuencia objetivo.
- El silenciamiento efectivo requería el transporte nuclear activo de siRNA o la permeabilización de la envoltura nuclear a través de la transducción lentiviral.
Conclusiones:
- El silenciamiento transcripcional dirigido por siRNA es un mecanismo conservado en los mamíferos.
- Esto proporciona una nueva estrategia para inhibir la función génica de los mamíferos a través de modificaciones epigenéticas.
- Los hallazgos destacan la importancia de la localización nuclear para el silenciamiento de genes transcripcionales mediados por siRNA.
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