Kuroyanagi Miwa, Yamada Kenji, Hatsugai Noriyuki, Kondo Maki, Nishimura Mikio, Hara-Nishimura Ikuko
Graduate School of Science, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
J Biol Chem. 2005 Sep 23;280(38):32914-20. doi: 10.1074/jbc.M504476200. Epub 2005 Jul 25.
Some compatible pathogens secrete toxins to induce host cell death and promote their growth. The toxin-induced cell death is a pathogen strategy for infection. To clarify the executioner of the toxin-induced cell death, we examined a fungal toxin (fumonisin B1 (FB1))-induced cell death of Arabidopsis plants. FB1-induced cell death was accompanied with disruption of vacuolar membrane followed by lesion formation. The features of FB1-induced cell death were completely abolished in the Arabidopsis vacuolar processing enzyme (VPE)-null mutant, which lacks all four VPE genes of the genome. Interestingly, an inhibitor of caspase-1 abolished FB1-induced lesion formation, as did a VPE inhibitor. The VPE-null mutant had no detectable activities of caspase-1 or VPE in the FB1-treated leaves, although wild-type leaves had the caspase-1 and VPE activities, both of which were inhibited by a caspase-1 inhibitor. gammaVPE is the most essential among the four VPE homologues for FB1-induced cell death in Arabidopsis leaves. Recombinant gammaVPE recognized a VPE substrate with Km = 30.3 microm and a caspase-1 substrate with Km = 44.2 microm, which is comparable with the values for mammalian caspase-1. The gammaVPE precursor was self-catalytically converted into the mature form exhibiting caspase-1 activity. These in vivo and in vitro analyses demonstrate that gammaVPE is the proteinase that exhibits a caspase-1 activity. We show that VPE exhibiting a caspase-1 activity is a key molecule in toxin-induced cell death. Our findings suggest that a susceptible response of toxin-induced cell death is caused by the VPE-mediated vacuolar mechanism similar to a resistance response of hypersensitive cell death (Hatsugai, N., Kuroyanagi, M., Yamada, K., Meshi, T., Tsuda, S., Kondo, M., Nishimura, M., and Hara-Nishimura, I. (2004) Science 305, 855-858).
一些共生病原体分泌毒素以诱导宿主细胞死亡并促进自身生长。毒素诱导的细胞死亡是病原体的一种感染策略。为了阐明毒素诱导细胞死亡的执行者，我们研究了一种真菌毒素（伏马菌素B1（FB1））诱导的拟南芥植株细胞死亡。FB1诱导的细胞死亡伴随着液泡膜破裂，随后形成病斑。在拟南芥液泡加工酶（VPE）缺失突变体中，FB1诱导的细胞死亡特征完全消失，该突变体缺乏基因组中的所有四个VPE基因。有趣的是，半胱天冬酶-1抑制剂消除了FB1诱导的病斑形成，VPE抑制剂也有同样的效果。在经FB1处理的叶片中，VPE缺失突变体未检测到半胱天冬酶-1或VPE的活性，而野生型叶片具有半胱天冬酶-1和VPE活性，这两种活性均被半胱天冬酶-1抑制剂抑制。γVPE是拟南芥叶片中FB1诱导细胞死亡的四个VPE同源物中最关键的一个。重组γVPE识别一种Km = 30.3微摩尔的VPE底物和一种Km = 44.2微摩尔的半胱天冬酶-1底物，这与哺乳动物半胱天冬酶-1的值相当。γVPE前体通过自我催化转化为具有半胱天冬酶-1活性的成熟形式。这些体内和体外分析表明，γVPE是具有半胱天冬酶-1活性的蛋白酶。我们表明，具有半胱天冬酶-1活性的VPE是毒素诱导细胞死亡中的关键分子。我们的数据表明，毒素诱导细胞死亡的敏感反应是由VPE介导的液泡机制引起的，类似于过敏细胞死亡的抗性反应（初贝直、黑柳正明、山田健、梅西、津田诚、近藤真、西村正、原西村郁子（2004年）《科学》305卷，855 - 858页）。
