RESUMEN
We demonstrate that the hormone-binding domain (HBD) of the human estrogen receptor (ER) can function as an autonomous regulatory domain in the budding yeast, Saccharomyces cerevisiae. As in mammalian cells, the HBD can subject the activity of a heterologous protein, which is fused to it, to hormonal control. Thus, a chimeric transcriptional activator consisting of (i) the DNA-binding domain of GAL4, (ii) the ER HBD, and (iii) the activation domain of viral protein 16 (VP16) stimulates both episomal and integrated reporter genes exclusively in the presence of steroid hormone. Steroids being gratuitous signals for yeast, this fusion protein is a convenient tool for highly regulated production of proteins of interest. Notably, it can be exploited to activate the commonly used galactose-inducible expression vectors without switching the carbon source.
Asunto(s)
Proteínas Fúngicas/metabolismo , Regulación Fúngica de la Expresión Génica , Receptores de Estrógenos/fisiología , Proteínas Recombinantes de Fusión/genética , Transactivadores/metabolismo , Activación Transcripcional , ADN Recombinante , Proteínas de Unión al ADN/metabolismo , Inducción Enzimática , Proteínas Fúngicas/genética , Genes Fúngicos , Genes Virales , Células HeLa , Humanos , Plásmidos , Regiones Promotoras Genéticas , Receptores de Estrógenos/genética , Proteínas Recombinantes de Fusión/fisiología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Transactivadores/genéticaRESUMEN
Analysis of the structure and function of a protein such as the epidermal growth factor receptor is facilitated by the use of antibodies directed against discrete portions of the protein. Here, we describe the characterization and use of antibodies directed against synthetic peptides corresponding to specific portions of the epidermal growth factor receptor and/or v-erbB protein. In particular, one useful antiserum has allowed us to compare the protein kinase activities of the epidermal growth factor receptor and the v-erbB proteins and to conclude that the v-erbB protein is a protein-tyrosine specific kinase as is its homologue the avian epidermal growth factor receptor.