A novel mechanism for the inhibition of hyaluronan biosynthesis by 4-methylumbelliferone
A novel mechanism for the inhibition of hyaluronan biosynthesis by 4-methylumbelliferone
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DOI：
10.1074/jbc.m405918200
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发表时间：
2004-08-06
影响因子：
4.8
通讯作者：
Itano, N
中科院分区：
文献类型：
--
作者：
Kakizaki, I;Kojima, K;Itano, N
透明质酸（HA）生物合成的特异性抑制剂可以是预防癌症侵袭和转移的有价值的治疗剂。我们以前已经发现，4-甲基伞形酮（MU）抑制HA合成的人皮肤成纤维细胞和C组链球菌。在本文中，在哺乳动物细胞中的抑制机制进行了研究，使用大鼠3 Y1成纤维细胞稳定表达HA合成酶（HAS）2。转染子暴露于抑制剂导致HA生物合成和基质形成的显着减少。HAS转录本的评价和无细胞HA合成的分析证明了MU对HAS活性的转录后抑制。最有趣的是，HAS活性的转录后抑制也被观察到使用对硝基苯酚，一个众所周知的UDP-葡糖醛酸转移酶（UGT）的底物。我们研究了是否抑制施加的葡萄糖醛酸化的MU使用高压液相色谱法和TLC分析。MU-葡萄糖醛酸（GlcUA）的产生与HAS转染子中HA合成的抑制一致。在对照细胞中也检测到相似水平的MU-GlcUA，表明葡萄糖醛酸化由内源性UGT介导。升高的UGT水平显著增强MU的抑制作用。相反，当向无细胞HA合成系统中加入过量的UDP-GlcUA时，MU的抑制作用减弱至对照水平。我们提出了一种新的机制，MU介导的抑制HA的合成，涉及葡萄糖醛酸化的MU内源性UGT导致的UDP-GlcUA耗尽。
Specific inhibitors of hyaluronan (HA) biosynthesis can be valuable therapeutic agents to prevent cancer invasion and metastasis. We have found previously that 4-methylumbelliferone (MU) inhibits HA synthesis in human skin fibroblasts and in group C Streptococcus. In this paper, the inhibition mechanism in mammalian cells was investigated using rat 3Y1 fibroblasts stably expressing HA synthase (HAS) 2. Exposure of the transfectants to the inhibitor resulted in significant reduction of HA biosynthesis and matrix formation. The evaluation of HAS transcripts and analysis of cell-free HA synthesis demonstrated the post-transcriptional suppression of HAS activity by MU. Most interesting, the post-transcriptional suppression of HAS activity was also observed using p-nitrophenol, a well known substrate for UDP-glucuronyltransferases (UGT). We investigated whether the inhibition was exerted by the glucuronidation of MU using both high pressure liquid chromatography and TLC analyses. The production of MU-glucuronic acid (GlcUA) was consistent with the inhibition of HA synthesis in HAS transfectants. MU-GlcUA was also detected at a similar level in control cells, suggesting that the glucuronidation was mediated by an endogenous UGT. Elevated levels of UGT significantly enhanced the inhibitory effects of MU. In contrast, the inhibition by MU was diminished to the control level when an excess of UDP-GlcUA was added to the cell-free HA synthesis system. We propose a novel mechanism for the MU-mediated inhibition of HA synthesis involving the glucuronidation of MU by endogenous UGT resulting in a depletion of UDP-GlcUA.