Hiraoka Yoshinori, Yoshida Kazuhiro, Ohno Mikiko, Matsuoka Tatsuhiko, Kita Toru, Nishi Eiichiro
Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, 54 Shogoin-Kawahara-Cho, Sakyo-Ku, Kyoto 606-8507, Japan.
Biochem Biophys Res Commun. 2008 May 23;370(1):154-8. doi: 10.1016/j.bbrc.2008.03.050. Epub 2008 Mar 18.
Tumor necrosis factor-alpha (TNF-alpha) is released from cells by proteolytic cleavage of a membrane-anchored precursor. The TNF-alpha-converting enzyme (TACE/ADAM17) is the major sheddase for ectodomain shedding of TNF-alpha. At present, however, it is poorly understood how its catalytic activity is regulated. Here, we show that nardilysin (N-arginine dibasic convertase; NRDc) enhanced TNF-alpha shedding. In a cell-based shedding assay, expression of NRDc synergistically enhanced TACE-induced TNF-alpha shedding. A peptide cleavage assay in vitro showed that recombinant NRDc enhances the cleavage of TNF-alpha induced by TACE. Notably, co-incubation of NRDc completely reversed the inhibitory effect of a physiological concentration of salt on TACE's activity in vitro. Overexpression of NRDc in TACE-deficient fibroblasts resulted in an increase in the amount of TNF-alpha released. Co-expression of NRDc with ADAM10 promoted the release compared with the sole expression of ADAM10. These results suggested that NRDc enhances TNF-alpha shedding through activation of not only TACE but ADAM10. Our results indicate the involvement of NRDc in ectodomain shedding of TNF-alpha, which may be a novel target for anti-inflammatory therapies.
肿瘤坏死因子-α（TNF-α）通过膜锚定前体的蛋白水解切割从细胞中释放出来。TNF-α转化酶（TACE/ADAM17）是TNF-α胞外域脱落的主要剪切酶。然而，目前对其催化活性如何调节知之甚少。在此，我们表明nardilysin（N-精氨酸二肽基转化酶；NRDc）增强了TNF-α的脱落。在基于细胞的脱落试验中，NRDc的表达协同增强了TACE诱导的TNF-α脱落。体外肽切割试验表明，重组NRDc增强了TACE诱导的TNF-α切割。值得注意的是，NRDc的共孵育完全逆转了生理浓度盐对TACE体外活性的抑制作用。在TACE缺陷的成纤维细胞中过表达NRDc导致释放的TNF-α量增加。与单独表达ADAM10相比，NRDc与ADAM10共表达促进了释放。这些结果表明，NRDc不仅通过激活TACE而且通过激活ADAM10来增强TNF-α的脱落。我们的结果表明NRDc参与了TNF-α的胞外域脱落，这可能是抗炎治疗的一个新靶点。
