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Transformación celular por pp60c-src mutado en el dominio regulador carboxiterminal
Cell
|April 10, 1987
Resumen
La introducción de mutaciones en pollo pp60c-src revela que alterar la tirosina 527 aumenta la actividad de la quinasa y el potencial de transformación. El extremo carboxilo es crucial para la asociación con el antígeno del tumor medio del polioma (mT).
Área de la Ciencia:
- Biología Molecular Biología Molecular
- La oncogénesis es la oncogénesis.
- Función de la proteína quinasa Función de la proteína quinasa
Sus antecedentes:
- El pollo pp60c-src es una tirosina quinasa proto-oncogénica regulada por su región carboxiterminal.
- La fosforilación en la tirosina 527 (Y527) típicamente inhibe la actividad de la quinasa.
- Las mutaciones que afectan a esta región reguladora pueden alterar la función de la src quinasa y la transformación celular.
Objetivo del estudio:
- Investigar las consecuencias funcionales de mutaciones específicas en la región reguladora carboxiterminal del pollo pp60c-src.src.
- Determinar el impacto de estas mutaciones en la actividad de la proteína quinasa, la fosforilación in vivo y la transformación celular.
- Explorar el papel del extremo carboxilo en la interacción con el antígeno del tumor medio del polioma (mT).
Principales métodos:
- Se utilizó la mutagénesis dirigida al sitio para introducir dos mutaciones en el gen pp60c-src del pollo: F527 (Y527F) y Am517 (truncamiento).
- Las proteínas mutantes y de tipo salvaje se analizaron para la fosforilación in vivo en tirosina 416 (Y416).
- Se midió la actividad específica de la proteína quinasa y se realizaron ensayos de formación de foco en células 3T3 de NIH.
- Se evaluó la formación de complejos entre las variantes pp60c-src y el antígeno del tumor medio del polioma (mT).
Principales resultados:
- Tanto las proteínas mutantes pp60c-src F527 y Am517 fueron fosforiladas en Y416 in vivo.
- El mutante F527 exhibió un aumento de 5 a 10 veces en la actividad específica de la quinasa en comparación con el tipo silvestre.
- El mutante Am517 mostró una actividad de quinasa similar a la del tipo salvaje, pero ambos mutantes indujeron la formación de focos con menor frecuencia y focos más pequeños que el mT.
- El tipo salvaje y F527 pp60c-src formaron complejos con mT, mientras que Am517 pp60c-src no lo hizo.
Conclusiones:
- La incapacidad para fosforilar Y527 mejora significativamente la actividad de la proteína quinasa pp60c-src y la capacidad de transformación.
- La transformación inducida por mT implica eventos celulares más allá de la falta de fosforilación de Y527 en pp60c-src.
- La activación de la actividad de la quinasa pp60c-src puede no ser esencial para la transformación mediada por el mutante Am517.
- El extremo carboxilo de pp60c-src es necesario para su asociación con mT.
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