ACTIVATION OF TREHALASE DURING GROWTH INDUCTION BY NITROGEN-SOURCES IN THE YEAST SACCHAROMYCES-CEREVISIAE DEPENDS ON THE FREE CATALYTIC SUBUNITS OF CAMP-DEPENDENT PROTEIN-KINASE, BUT NOT ON FUNCTIONAL RAS PROTEINS
ACTIVATION OF TREHALASE DURING GROWTH INDUCTION BY NITROGEN-SOURCES IN THE YEAST SACCHAROMYCES-CEREVISIAE DEPENDS ON THE FREE CATALYTIC SUBUNITS OF CAMP-DEPENDENT PROTEIN-KINASE, BUT NOT ON FUNCTIONAL RAS PROTEINS
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DOI：
10.1002/yea.320100807
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发表时间：
1994-08-01
期刊：
影响因子：
2.6
通讯作者：
THEVELEIN, JM
中科院分区：
文献类型：
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作者：
DURNEZ, P;PERNAMBUCO, MB;THEVELEIN, JM
在存在可发酵碳源的情况下，向酿酒酵母的葡萄糖抑制、氮饥饿的 G0 细胞中添加氮源会诱导生长，并在几分钟内导致海藻糖酶活性增加五倍，与蛋白质合成无关。氮激活的海藻糖酶可以通过碱性磷酸酶处理在体外失活，这支持了该激活是由磷酸化触发的观点。仅含有三个 TPK 基因（编码 cAMP 依赖性蛋白激酶催化亚基）之一的酵母菌株表现出不同程度的氮诱导海藻糖酶激活。有效性的顺序与之前报道的葡萄糖去抑制的酵母细胞中葡萄糖诱导的海藻糖酶激活的顺序不同。通过部分失活剩余 TPK 基因中的点突变进一步降低 TPK 编码的催化亚基活性，进一步减少氮诱导的海藻糖酶激活，而同一菌株中 BCY1 基因（编码调节亚基）的缺失导致激活程度增加。在这样的 tpk(w1) bcy1 菌株中删除 RAS 基因没有效果。这些结果与单独通过游离催化亚基介导氮诱导的海藻糖酶活化一致。他们支持了我们之前的结论，即cAMP在这个氮诱导的激活过程中并不充当第二信使，以及我们的建议，即一种新的氮诱导信号通路在蛋白激酶A的游离催化亚基水平上与cAMP通路整合，Western印迹实验表明海藻糖酶激活程度的差异并不是由于海藻糖酶表达的差异所致。另一方面，我们不能完全排除蛋白激酶A影响氮诱导的激活机制本身，而不是直接作用于海藻糖酶。然而，任何此类替代解释都需要除了蛋白激酶 A 的公认调节之外，还存在一种额外但未知的海藻糖酶激活机制。
Addition of a nitrogen-source to glucose-repressed, nitrogen-starved G0 cells of the yeast Saccharomyces cerevisiae in the presence of a fermentable carbon source induces growth and causes within a few minutes a five-fold, protein-synthesis-independent increase in the activity of trehalase. Nitrogen-activated trehalase could be deactivated in vitro by alkaline phosphatase treatment, supporting the idea that the activation is triggered by phosphorylation. Yeast strains containing only one of the three TPK genes (which encode the catalytic subunit of cAMP-dependent protein kinase) showed different degrees of nitrogen-induced trehalase activation. The order of effectiveness was different from that previously reported for glucose-induced activation of trehalase in glucose-derepressed yeast cells. Further reduction of TPK-encoded catalytic subunit activity by partially inactivating point mutations in the remaining TPK gene further diminished nitrogen-induced trehalase activation, while deletion of the BCY1 gene (which encodes the regulatory subunit) in the same strains resulted in an increase in the extent of activation. Deletion of the RAS genes in such a tpk(w1) bcy1 strain had no effect. These results are consistent with mediation of nitrogen-induced trehalase activation by the free catalytic subunits alone. They support our previous conclusion that cAMP does not act as second messenger in this nitrogen-induced activation process and our suggestion that a novel nitrogen-induced signaling pathway integrates with the cAMP pathway at the level of the free catalytic subunits of protein kinase A, Western blot experiments showed that the differences in the extent of trehalase activation were not due to differences in trehalase expression. On the other hand, we cannot completely exclude that protein kinase A influences the nitrogen-induced activation mechanism itself rather than acting directly on trehalase. However, any such alternative explanation requires the existence of an additional, yet unknown, mechanism for activation of trehalase besides the well-established regulation by protein kinase A.