The function and properties of the Azf1 transcriptional regulator change with growth conditions in Saccharomyces cerevisiae
The function and properties of the Azf1 transcriptional regulator change with growth conditions in Saccharomyces cerevisiae
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DOI：
10.1128/ec.5.2.313-320.2006
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发表时间：
2006-02-01
期刊：
影响因子：
--
通讯作者：
Heideman, W
中科院分区：
文献类型：
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作者：
Slattery, MG;Liko, D;Heideman, W
Azf 1在葡萄糖中生长的酿酒酵母细胞中激活CLN 3转录。有趣的是，其他研究表明，Azf 1在葡萄糖生长的细胞中几乎检测不到。微阵列实验表明，Azf 1激活不同碳源中的非重叠基因组：在葡萄糖中，Azf 1激活碳和能量代谢基因，在甘油-乳酸中，Azf 1激活细胞壁维持所需的基因。与在azf 1 Delta突变体中观察到的细胞壁维持基因表达降低一致，我们在非发酵培养基中于37 ℃观察到azf 1 Delta细胞中存在显著的生长缺陷。细胞壁完整性检测，如对calcofluor白色、十二烷基硫酸钠或咖啡因的敏感性，证实了azf 1 Delta突变体在不可发酵培养基中的细胞壁缺陷。凝胶位移实验表明，Azf 1与序列为AAAAGAAA（A（4）GA（3））的DNA元件结合，该序列在Azf 1敏感基因的启动子中富集，并通过全基因组研究预测。这表明，许多Azf 1依赖的转录本可能直接受Azf 1结合的调节。我们发现，在葡萄糖生长的细胞中的Azf 1蛋白水平与在甘油-乳酸盐中生长的细胞中的Azf 1水平相当;然而，这只能用最小化蛋白水解的细胞提取程序来证明。葡萄糖产生的条件下，不稳定的Azf 1蛋白，这可能反映了葡萄糖诱导的变化Azf 1的三级或四级结构的发现。
Azf1 activates CLN3 transcription in Saccharonzyces cerevisiae cells growing in glucose. Paradoxically, other studies have shown Azf1 be almost undetectable in glucose-grown cells. Microarray experiments showed that Azf1 activates nonoverlapping gene sets in different carbon sources: in glucose, Azf1 activates carbon and energy metabolism genes, and in glycerol-lactate, Azf1 activates genes needed for cell wall maintenance. Consistent with the decreased expression of cell wall maintenance genes observed with azf1 Delta, mutants, we observed a marked growth defect in the azf1 Delta, cells at 37 degrees C in nonfermentable medium. Cell wall integrity assays, such as sensitivity to calcofluor white, sodium dodecyl sulfate, or caffeine, confirmed cell wall defects in azf1 Delta mutants in nonfermentable medium. Gel shift experiments show that Azf1 binds to DNA elements with the sequence AAAAGAAA (A(4)GA(3)), a motif enriched in the promoters of Azf1-sensitive genes and predicted by whole-genome studies. This suggests that many of the Azf1-dependent transcripts may be regulated directly by Azf1 binding. We found that the levels of Azf1 protein in glucose-grown cells were comparable to Azf1 levels in cells grown in glycerol-lactate; however, this could only be demonstrated with a cell extraction procedure that minimizes proteolysis. Glucose produces conditions that destabilize the Azf1 protein, a finding that may reflect a glucose-induced change in Azf1 tertiary or quaternary structure.