Portela Paula, Van Dijck Patrick, Thevelein Johan M, Moreno Silvia
Departamento de Qui;mica Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria, Pabellón 2, 1428, Buenos Aires, Argentina.
FEMS Yeast Res. 2003 Mar;3(1):119-26. doi: 10.1111/j.1567-1364.2003.tb00147.x.
Protein kinase A (PKA) activity was measured in situ in permeabilised Saccharomyces cerevisiae cells in the absence and the presence of cAMP. Four strains genetically predicted to have differential PKA-dependent phenotypes were used: a wild-type strain and a strain containing a bcy1-14 mutation (with almost constitutively active PKA), and the same strains with overexpression of the wild-type or mutant BCY1 gene, respectively. Cells were grown on galactose or glucose. The measured phenotypic characteristics were: trehalose and glycogen levels and the activity of a reporter gene under control of the NTH1 promoter. The 'endogenous' PKA activity (measured in situ in the absence of cAMP) showed the best correlation with the PKA-dependent phenotypes determined in vivo. We propose that this parameter offers a good estimate for the degree of activation of PKA in vivo.
在有无环磷酸腺苷（cAMP）的情况下，对通透化的酿酒酵母细胞中的蛋白激酶A（PKA）活性进行了原位测量。使用了四种经基因预测具有不同PKA依赖性表型的菌株：一个野生型菌株和一个含有bcy1 - 14突变（PKA几乎持续激活）的菌株，以及分别过表达野生型或突变型BCY1基因的相同菌株。细胞在半乳糖或葡萄糖上生长。测量的表型特征包括：海藻糖和糖原水平以及在NTH1启动子控制下的报告基因的活性。“内源性”PKA活性（在无cAMP的情况下原位测量）与体内确定的PKA依赖性表型显示出最佳相关性。我们认为该参数为体内PKA激活程度提供了良好的估计。
