Lee C G, Jenkins N A, Gilbert D J, Copeland N G, O'Brien W E
Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030, USA.
Immunogenetics. 1995;41(5):263-70. doi: 10.1007/BF00172150.
The expression of many genes is altered upon the activation of macrophages by bacterial LPS. These genes play a crucial role in the orchestration of various responses to protect the host against infection. A novel 2.3 kilobase (kb) cDNA, designated IRG1, was obtained from a cDNA library prepared with RNA isolated from RAW 264.7 following lipopolysaccharide stimulation. Sequence analysis of the clone revealed no identity to any known genes but showed the presence of many potential phosphorylation sites suggesting that IRG1 protein product may be regulated at this level. Furthermore, IRG1 contains the motif for glycosaminoglycan attachment site, implying that IRG1 may be a proteoglycan. By interspecific back-cross analysis, Irg1 was mapped to mouse chromosome 14 linked to Tyrp2 and Rap2a. The IRG1 message appears 1.5 h following LPS exposure and its induction was not dependent on new protein synthesis. In fact, cycloheximide induced the expression of IRG1, suggesting that a protein repressor prevents the expression of IRG1 when uninduced. The role of the protein kinase A pathway in regulating the induction of IRG1 by LPS is questionable, because although forskolin inhibited its induction, neither dibutyrl-cAMP nor 8-(4-chlorophenylthio)-cAMP had much effect on its expression. In contrast, activation of protein kinase C potentiated the LPS response. Chelation of extracellular calcium inhibited IRG1 4 h after LPS induction, while increasing intracellular calcium had little effect on the levels of the IRG1 transcript. Inhibiting tyrosine phosphorylation abrogated the induction of IRG1 by LPS. Hence, the induction of IRG1 by LPS is mediated by tyrosine kinase and protein kinase C pathway.
细菌脂多糖激活巨噬细胞后，许多基因的表达会发生改变。这些基因在协调各种保护宿主免受感染的反应中起着关键作用。从脂多糖刺激后的RAW 264.7细胞中分离的RNA构建的cDNA文库中，获得了一个新的2.3千碱基（kb）的cDNA，命名为IRG1。对该克隆的序列分析表明，它与任何已知基因均无同源性，但显示存在许多潜在的磷酸化位点，这表明IRG1蛋白产物可能在此水平受到调控。此外，IRG1含有糖胺聚糖附着位点的基序，这意味着IRG1可能是一种蛋白聚糖。通过种间回交分析，将Irg1定位到与Tyrp2和Rap2a连锁的小鼠14号染色体上。脂多糖暴露后1.5小时出现IRG1信息，其诱导不依赖于新的蛋白质合成。事实上，放线菌酮可诱导IRG1的表达，这表明在未诱导时，一种蛋白质阻遏物会阻止IRG1的表达。蛋白激酶A途径在调节脂多糖对IRG1的诱导作用方面存在疑问，因为尽管福司可林抑制其诱导，但二丁酰环磷腺苷和8-（4-氯苯硫基）-环磷腺苷对其表达均无太大影响。相反，蛋白激酶C的激活增强了脂多糖反应。脂多糖诱导4小时后，细胞外钙的螯合抑制了IRG1，而细胞内钙的增加对IRG1转录本水平影响不大。抑制酪氨酸磷酸化可消除脂多糖对IRG1的诱导。因此，脂多糖对IRG1的诱导是由酪氨酸激酶和蛋白激酶C途径介导的。
