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Clonado y caracterización de CAP, el gen de S. cerevisiae que codifica la proteína 70 kd asociada a la adenilil
J Field1, A Vojtek, R Ballester
1Cold Spring Harbor Laboratory, New York 11724.
Cell
|April 20, 1990
Resumen
La proteína asociada a la ciclasa (CAP) es esencial para la señalización de la adenilil ciclasa sensible al RAS en la levadura. Los mutantes que carecen de CAP también exhiben defectos de crecimiento y cambios morfológicos, lo que sugiere funciones celulares más amplias.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- Biología celular Biología celular.
- La bioquímica es la bioquímica.
Sus antecedentes:
- La adenilil ciclasa en Saccharomyces cerevisiae es una enzima de varias subunidades.
- La vía RAS/adenilil ciclasa regula las respuestas celulares a diversos estímulos.
Objetivo del estudio:
- Clonar y caracterizar la subunidad de la proteína asociada a la ciclasa (CAP, por sus siglas en inglés) de la adenilil ciclasa de levadura.
- Investigar el papel de CAP en la señalización mediada por RAS y otros procesos celulares.
Principales métodos:
- Clonado de CAP cDNA utilizando una biblioteca de expresión de levadura en E. coli.
- Cribado con antisera contra el CAP purificado.
- Análisis de la actividad de la adenilil ciclasa en las membranas de levadura con deficiencia de CAP.
- Análisis fenotípico de las mutaciones CAP.
Principales resultados:
- Se aisló un ADNc que codifica una proteína CAP de 526 aminoácidos; no mostró homología con secuencias conocidas.
- La levadura que carece de CAP exhibió una disminución de la actividad de la adenilil ciclasa estimulada por RAS2 in vitro.
- Los mutantes deficientes de CAP mostraron sensibilidad a la temperatura, incapacidad para crecer en un medio rico, sensibilidad al hambre de nitrógeno y morfología celular alterada.
Conclusiones:
- CAP es un componente crucial de la vía de señalización de la adenilil ciclasa en levaduras que responde a las RAS.
- CAP juega papeles más allá de la señalización RAS, impactando el crecimiento, la morfología y la respuesta al estrés.
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