Differential regulation of gamma-glutamylcysteine synthetase heavy and light subunit gene expression
Differential regulation of gamma-glutamylcysteine synthetase heavy and light subunit gene expression
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DOI：
10.1042/bj3260167
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发表时间：
1997-08-15
影响因子：
4.1
通讯作者：
Lu, SC
中科院分区：
文献类型：
--
作者：
Cai, JX;Huang, ZZ;Lu, SC
γ-谷氨酰半胱氨酸合成酶（GCS）是谷胱甘肽生物合成中的限速酶，由重亚基和轻亚基组成。虽然重亚基是单独的酶活性，轻亚基起着重要的调节作用，使全酶功能更有效，在目前的研究中，我们研究了是否已知的影响基因表达的重亚基的条件也影响轻亚基，和所涉及的机制。用胰岛素和氢化可的松等激素处理培养的大鼠肝细胞，或在低细胞密度下接种肝细胞，仅增加重亚基的稳态mRNA水平。用马来酸二乙酯（DEM）、丁硫酰亚胺（BSO）和叔丁基对苯二酚（TBH）处理，增加两种亚基的稳态mRNA水平和基因转录速率。这些治疗共同具有诱导氧化应激和激活核因子κ B（NF-κ B）的能力。蛋白酶抑制剂7-amino-1-chloro-3-tosylamido-2-heptanone（TLCK）或L-1-tosylamido-2-phenylethyl chloromethyl ketone（TPCK）处理对基础NF-κ B和GCS亚基mRNA水平没有影响，但阻断了DEM、BSO和TBH对NF-κ B的激活以及BSO和TBH对GCS重亚基mRNA水平的升高。另一方面，DEM-，BSO-和TBH-诱导的GCS轻亚基mRNA水平的增加不受TLCK和TPCK的影响。因此，只有重亚基是神经调节和生长敏感的，而这两个亚基都受氧化应激的调节。通过NF-κ B的信号转导仅参与氧化应激介导的重亚基基因表达的变化。
gamma-Glutamylcysteine synthetase (GCS) is the rate-limiting enzyme in the biosynthesis of glutathione and is composed of a heavy and a light subunit. Although the heavy subunit is enzymically active alone, the light subunit plays an important regulatory role by making the holoenzyme function more efficiently, In the current study we examined whether conditions which are known to influence gene expression of the heavy subunit also influence that of the light subunit, and the mechanisms involved. Treatment of cultured rat hepatocytes with hormones such as insulin and hydrocortisone, or plating hepatocytes under low cell density increased the steady-state mRNA level of the heavy subunit only, Treatment with diethyl maleate (DEM), buthionine sulphoximine (BSO) and t-butylhydroquinone (TBH) increased the steady state mRNA level and gene transcription rates of both subunits. These treatments share in common their ability to induce oxidative stress and activate nuclear factor kappa B (NF-kappa B). Treatment with protease inhibitors 7-amino-1-chloro-3-tosylamido-2-heptanone (TLCK) or L-1-tosylamido-2-phenylethyl chloromethyl ketone (TPCK) had no influence on the basal NF-kappa B and GCS subunit mRNA levels, but blocked the activation of NF-kappa B by DEM, BSO and TBH, and the increase in GCS heavy subunit mRNA level by BSO and TBH. On the other hand, the DEM-, BSO-and TBH-induced increase in GCS light-subunit mRNA level was unaffected by TLCK and TPCK. Thus only the heavy subunit is hormonally regulated and growth sensitive, whereas a both subunits are regulated by oxidative stress. Signalling through NF-kappa B is involved only in the oxidative-stress-mediated changes in the heavy subunit gene expression.