The β subunit of yeast AMP-activated protein kinase directs substrate specificity in response to alkaline stress.
The β subunit of yeast AMP-activated protein kinase directs substrate specificity in response to alkaline stress.
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DOI：
10.1016/j.cellsig.2016.08.016
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发表时间：
2016-12
影响因子：
4.8
通讯作者：
Schmidt MC
中科院分区：
文献类型：
--
作者：
Chandrashekarappa DG;McCartney RR;O'Donnell AF;Schmidt MC
酿酒酵母表达Snf 1激酶的三种同种型，不同之处在于β亚基存在，Gal 83、Sip 1或Sip 2。在这里，我们调查的丰度，激活，定位和信号特异性的三个Snf 1亚型。这些亚型的相对丰度进行了评估，通过定量免疫印迹使用两种不同的蛋白质提取方法和荧光显微镜。含Gal 83的同种型在所有测定中最丰富，而Sip 1和Sip 2同种型的丰度通常被低估，尤其是在玻璃珠提取中。早期评估Snf 1亚型功能的研究利用基因缺失作为抑制特异性亚型的手段。在这里，我们使用Gal 83和Sip 2的点突变和Sip 1的17个氨基酸的C-末端截短，以使其特异性同种型不影响其丰度或与其他亚基的关联。低葡萄糖和碱性应力的影响进行了检查两个Snf 1磷酸化底物，Mig 1和Mig 2蛋白。三种亚型中的任何一种都能够磷酸化Mig 1以响应葡萄糖应激。相比之下，Gal 83同种型的Snf 1是必要的和足够的Mig 2蛋白的磷酸化，在响应碱性胁迫。碱性应激导致所有三种同种型的激活，但只有Gal 83同种型易位到细胞核并磷酸化Mig 2。SAK 1基因的缺失阻断了Gal 83的核转位和向Mig 2的信号传导。这些数据强烈支持Snf 1信号传导特异性是由不同Snf 1亚型的定位介导的观点。
Saccharomyces cerevisiae express three isoforms of Snf1 kinase that differ by which β subunit is present, Gal83, Sip1 or Sip2. Here we investigate the abundance, activation, localization and signaling specificity of the three Snf1 isoforms. The relative abundance of these isoforms was assessed by quantitative immunoblotting using two different protein extraction methods and by fluorescence microscopy. The Gal83 containing isoform is the most abundant in all assays while the abundance of the Sip1 and Sip2 isoforms is typically underestimated especially in glass-bead extractions. Earlier studies to assess Snf1 isoform function utilized gene deletions as a means to inactivate specific isoforms. Here we use point mutations in Gal83 and Sip2 and a 17 amino acid C-terminal truncation of Sip1 to inactivate specific isoforms without affecting their abundance or association with the other subunits. The effect of low glucose and alkaline stresses was examined for two Snf1 phosphorylation substrates, the Mig1 and Mig2 proteins. Any of the three isoforms was capable of phosphorylating Mig1 in response to glucose stress. In contrast, the Gal83 isoform of Snf 1 was both necessary and sufficient for the phosphorylation of the Mig2 protein in response to alkaline stress. Alkaline stress led to the activation of all three isoforms yet only the Gal83 isoform translocates to the nucleus and phosphorylates Mig2. Deletion of the SAK1 gene blocked nuclear translocation of Gal83 and signaling to Mig2. These data strongly support the idea that Snf1 signaling specificity is mediated by localization of the different Snf1 isoforms.
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腺嘌呤核苷酸和糖原调节 AMPK 的结构基础。
影响因子：
44.1
作者：
Li X;Wang L;Zhou XE;Ke J;de Waal PW;Gu X;Tan MH;Wang D;Wu D;Xu HE;Melcher K
通讯作者：
Melcher K
影响因子：
5.3
作者：
Hedbacker, K;Hong, SP;Carlson, M
通讯作者：
Carlson, M
影响因子：
4.8
作者：
Nath, N;McCartney, RR;Schmidt, MC
通讯作者：
Schmidt, MC
DOI：
10.1126/science.1215327
发表时间：
2012-05-18
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Hawley SA;Fullerton MD;Ross FA;Schertzer JD;Chevtzoff C;Walker KJ;Peggie MW;Zibrova D;Green KA;Mustard KJ;Kemp BE;Sakamoto K;Steinberg GR;Hardie DG
通讯作者：
Hardie DG
影响因子：
4.8
作者：
Iseli, TJ;Walter, M;Stapleton, D
通讯作者：
Stapleton, D