In vivo measurement of cytosolic and mitochondrial pH using a pH-sensitive GFP derivative in Saccharomyces cerevisiae reveals a relation between intracellular pH and growth
In vivo measurement of cytosolic and mitochondrial pH using a pH-sensitive GFP derivative in Saccharomyces cerevisiae reveals a relation between intracellular pH and growth
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DOI：
10.1099/mic.0.022038-0
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发表时间：
2009-01-01
期刊：
影响因子：
2.8
通讯作者：
Smits, Gertien J.
中科院分区：
文献类型：
--
作者：
Orij, Rick;Postmus, Jarne;Smits, Gertien J.
细胞区室的特定pH值影响几乎所有的生物化学过程，包括酶活性、蛋白质折叠和氧化还原状态。因此，准确，灵敏和特定的细胞内pH值（pH（i））的动态测量在活细胞中的隔室是至关重要的压力反应和适应的理解。我们使用了pH敏感的GFP衍生物“比率pHluorin”在细胞质和线粒体基质中表达的生长酿酒酵母来评估细胞质pH（pH（cyt））和线粒体pH（pH（mit））的变化，以响应营养物质的可用性、呼吸链活性、环境pH的变化和由添加山梨酸诱导的应激。体内测量允许准确测定细胞器特异性pH，确定在葡萄糖上指数生长的细胞中恒定pH（cyt）为7.2和恒定p（Hmit）为7.5。我们发现，p（Hcyt）和p（Hmit）的差异调节碳源和呼吸链抑制剂。在葡萄糖饥饿或山梨酸胁迫下，pH（i）降低与生长停滞一致。此外，在向葡萄糖饥饿培养物中加入葡萄糖后或从山梨酸脉冲中恢复后，pH（i）和生长在恢复中相似地一致。我们提出了pH（i）和细胞能量产生之间的关系，因此pH（i）和生长之间的关系。
The specific pH values of cellular compartments affect virtually all biochemical processes, including enzyme activity, protein folding and redox state. Accurate, sensitive and compartment-specific measurements of intracellular pH (pH(i)) dynamics in living cells are therefore crucial to the understanding of stress response and adaptation. We used the pH-sensitive GFP derivative 'ratiometric pHluorin' expressed in the cytosol and in the mitochondrial matrix of growing Saccharomyces cerevisiae to assess the variation in cytosolic pH (pH(cyt)) and mitochondrial pH (pH(mit)) in response to nutrient availability, respiratory chain activity, shifts in environmental pH and stress induced by addition of sorbic acid. The in vivo measurement allowed accurate determination of organelle-specific pH, determining a constant pH(cyt) of 7.2 and a constant p(Hmit) of 7.5 in cells exponentially growing on glucose. We show that p(Hcyt) and p(Hmit) are differentially regulated by carbon source and respiratory chain inhibitors. Upon glucose starvation or sorbic acid stress, pH(i) decrease coincided with growth stasis. Additionally, pH(i) and growth coincided similarly in recovery after addition of glucose to glucose-starved cultures or after recovery from a sorbic acid pulse. We suggest a relation between pH(i) and cellular energy generation, and therefore a relation between pH(i) and growth.