Loss of Vacuolar H+-ATPase (V-ATPase) Activity in Yeast Generates an Iron Deprivation Signal That Is Moderated by Induction of the Peroxiredoxin TSA2
Loss of Vacuolar H+-ATPase (V-ATPase) Activity in Yeast Generates an Iron Deprivation Signal That Is Moderated by Induction of the Peroxiredoxin TSA2
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DOI：
10.1074/jbc.m112.419259
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发表时间：
2013-04-19
影响因子：
4.8
通讯作者：
Kane, Patricia M.
中科院分区：
文献类型：
--
作者：
Diab, Heba I.;Kane, Patricia M.
液泡H+- atp酶（v - atp酶）使胞内细胞器酸化并有助于调节整个细胞pH。酵母vma突变体缺乏v - atp酶活性，允许探索所有真核生物共同的细胞pH，铁和氧化还原稳态之间的联系。先前对vma突变体的微阵列研究表明，在Aft1p（铁调控子）的控制下，多个铁摄取基因上调，只有一个抗氧化基因，即过氧化物还氧蛋白TSA2 （Milgrom， E., Diab， H., Middleton， F., and Kane， P. M.(2007)）。酵母液泡质子易位atp酶活性的丧失会导致慢性氧化应激。生物。化学。282,7125-7136)。构建了荧光生物传感器，将GFP置于aft1依赖启动子（P-FIT2-GFP）或TSA2启动子（P-TSA2-GFP）的转录控制下，以监测转录信号。在vma2 Delta突变体中，这两个生物传感器都被上调，康那霉素A对V-ATPase的急性抑制诱导了这两个启动子的协同上调。P-TSA2-GFP诱导依赖于yap1p，表明氧化应激信号。细胞总铁测量表明，尽管铁调控上调，但vma2 δ突变体是铁充满的。醋酸上调了野生型细胞中P-FIT2-GFP的表达，表明失去pH控制有助于突变体中的铁缺乏信号。铁的补充显著降低了P-FIT2-GFP的表达，令人惊讶的是，将P-TSA2-GFP恢复到野生型水平。tsa2 δ突变诱导Aft1p和P-FIT2-GFP表达的核定位。这些数据表明，Tsa2p作为aft1p驱动转录的负调节因子具有新的功能，在V-ATPase突变体中被诱导以限制铁调控子的转录。这代表了一种新的机制，架起了与pH稳态密切相关的抗氧化和铁调节途径。
Vacuolar H+-ATPases (V-ATPases) acidify intracellular organelles and help to regulate overall cellular pH. Yeast vma mutants lack V-ATPase activity and allow exploration of connections between cellular pH, iron, and redox homeostasis common to all eukaryotes. A previous microarray study in a vma mutant demonstrated up-regulation of multiple iron uptake genes under control of Aft1p (the iron regulon) and only one antioxidant gene, the peroxiredoxin TSA2 (Milgrom, E., Diab, H., Middleton, F., and Kane, P. M. (2007) Loss of vacuolar proton-translocating ATPase activity in yeast results in chronic oxidative stress. J. Biol. Chem. 282, 7125-7136). Fluorescent biosensors placing GFP under transcriptional control of either an Aft1-dependent promoter (P-FIT2-GFP) or the TSA2 promoter (P-TSA2-GFP) were constructed to monitor transcriptional signaling. Both biosensors were up-regulated in the vma2 Delta mutant, and acute V-ATPase inhibition with concanamycin A induced coordinate up-regulation from both promoters. P-TSA2-GFP induction was Yap1p-dependent, indicating an oxidative stress signal. Total cell iron measurements indicate that the vma2 Delta mutant is iron-replete, despite up-regulation of the iron regulon. Acetic acid up-regulated P-FIT2-GFP expression in wildtype cells, suggesting that loss of pH control contributes to an iron deficiency signal in the mutant. Iron supplementation significantly decreased P-FIT2-GFP expression and, surprisingly, restored P-TSA2-GFP to wild-type levels. A tsa2 Delta mutation induced both nuclear localization of Aft1p and P-FIT2-GFP expression. The data suggest a novel function for Tsa2p as a negative regulator of Aft1p-driven transcription, which is induced in V-ATPase mutants to limit transcription of the iron regulon. This represents a new mechanism bridging the antioxidant and iron-regulatory pathways that is intimately linked to pH homeostasis.