STUDIES ON THE EFFECT OF LYSOSOMOTROPIC AGENTS ON THE RELEASE OF GAL-BETA-1-4GLCNAC ALPHA-2,6-SIALYLTRANSFERASE FROM RAT-LIVER SLICES DURING THE ACUTE-PHASE RESPONSE
STUDIES ON THE EFFECT OF LYSOSOMOTROPIC AGENTS ON THE RELEASE OF GAL-BETA-1-4GLCNAC ALPHA-2,6-SIALYLTRANSFERASE FROM RAT-LIVER SLICES DURING THE ACUTE-PHASE RESPONSE
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DOI：
10.1042/bj2610389
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发表时间：
1989-07-15
影响因子：
4.1
通讯作者：
JAMIESON, JC
中科院分区：
文献类型：
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作者：
LAMMERS, G;JAMIESON, JC
在急性期反应中，Gal.beta.1-4GlcNAcα-2，6-唾液酸基转移酶(CMP-N-乙酰神经氨酸盐：β-半乳糖苷α-2，6-唾液酸基转移酶，EC 2.4.99.1)从大鼠肝脏中释放的机制是由于组织蛋白酶D样蛋白酶的作用，它将跨高尔基膜结合的酶从膜锚上裂解出来；这使得含有催化位点的酶的大部分逃逸到细胞外空间[Lammers和Jamieson(1988)Biochem]。J.256,623-631]。唾液酸转移酶的释放在pH值为5.6时最有效，这表明整个细胞从高尔基体释放唾液酸转移酶依赖于维持肝细胞跨高尔基体区的酸性环境。高尔基膜包含一个质子泵，可以维持这些隔室中的酸性pH[Glickman，Croen，Kelly和Al-Awquati(1983)J.Cell Biol]。97,1303-1308；Yamashiro，Tycko和Maxfield(1984)Cell(Cambridege，Mass.)37,789-800；Zhang和Schneider(1983)Biochem。生物群落。[中英文摘要]Re.Commun.114,620-625；安德森和帕萨克(1985)细胞(马萨诸塞州坎布里奇)40,635-643]。溶酶促进剂，如NH4Cl、氯喹和甲胺，可以穿透细胞的酸性隔间，如高尔基复合体，提高pH，从而影响蛋白水解性切割事件。本文用全细胞系统研究了溶酶促进剂对肝细胞释放唾液酸转移酶的影响。切片是从对照组大鼠和遭受急性期反应的大鼠制备的，其中唾液酸转移酶的释放显著增加[Lammers和Jamieson(1988)Biochem]。J.256,623-631；Kaplan，Woloski，Hellman和Jamieson(1983)J.Biol。化学。258、11505-11509]。在50 mM氯化铵、50 mM甲胺或1 mM氯喹存在下，唾液酸转移酶的释放几乎完全消失。去掉溶酶促进剂后，唾液酸转移酶释放的抑制作用被逆转，说明在所用条件下，这些促溶酶剂对细胞没有细胞毒性。大鼠α1-酸性糖蛋白的分泌不受高尔基复合体中蛋白水解物的影响，也没有发现溶酶体促进剂的存在对其有实质性的影响。结果表明，唾液酸转移酶催化部位的蛋白水解性切割是一个受高尔基体内pH显著影响的过程。
The mechanism of release of Gal.beta.1-4GlcNAc .alpha.-2,6-sialyltransferase (CMP-N-acetylneuraminate:.beta.-galactoside .alpha.-2,6-sialyltransferase, EC 2.4.99.1) from rat liver during the acute-phase response is due to the action of a cathepsin D-like proteinase that cleaves the trans-Golgi membrane-bound enzyme from a membrane anchor; this allows a major portion of the enzyme containing the catalytic site to escape into the extracellular space [Lammers and Jamieson (1988) Biochem. J. 256, 623-631]. The release of sialytransferase was most effective at pH 5.6, suggesting that release of sialytransferase from the Golgi in whole cells is dependent on maintaining an acidic environment in the trans-Golgi compartment of the hepatocyte. Golgi membranes contain a proton pump that maintains the acidic pH in these compartments [Glickman, Croen, Kelly and Al-Awquati (1983) J. Cell Biol. 97, 1303-1308; Yamashiro, Tycko and Maxfield (1984) Cell (Cambridege, Mass.) 37, 789-800; Zhang and Schneider (1983) Biochem. Biophys. Res. Commun. 114, 620-625; Anderson and Pathak (1985) Cell (Cambridge, Mass.) 40, 635-643]. Lysosomotropic agents, such as NH4Cl, chloroquine and methylamine can penetrate acidic compartments of the cell, such as the Golgi complex, raise the pH, and thus affect proteolytic cleavage events. The present paper describes the effect of lysosomotropic agents on the release of sialytransferase from the hepatocyte using liver slices as a whole-cell system. Slices were prepared from control rats and rats suffering from the acute-phase response, where release of sialytransferase is increased substantially [Lammers and Jamieson (1988) Biochem. J. 256, 623-631; Kaplan, Woloski, Hellman and Jamieson (1983) J. Biol. Chem. 258, 11505-11509]. Release of sialytransferase was almost abolished in presence of 50 mM-NH4Cl, 50 mM-methylamine or 1 mM-chloroquine. Inhibition of release of sialyltransferase was reversed when the lysosomotropic agents were removed from the medium, showing that these agents are not cytotoxic to the cells under the conditions used. The secretion of rat .alpha.1-acid glycoprotein, which is not subject to proteolytic processing in the Golgi complex, was not found to be substantially affected by the presence of lysosomotropic agents. The results suggest that proteolytic cleavage of the catalytic site of sialytransferase is a process that is significantly affected by the intra-Golgi pH.