VirA of Agrobacterium tumefaciens is an intradimer transphosphorylase and can actively block vir gene expression in the absence of phenolic signals
VirA of Agrobacterium tumefaciens is an intradimer transphosphorylase and can actively block vir gene expression in the absence of phenolic signals
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DOI：
10.1111/j.1365-2958.2004.04057.x
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发表时间：
2004-06-01
影响因子：
3.6
通讯作者：
Winans, SC
中科院分区：
文献类型：
--
作者：
Brencic, A;Xia, Q;Winans, SC
病毒-病毒双组分系统调节30个基因的VIR调节子，以响应宿主释放的化学信号。VirA是一种跨膜的同源二聚体组氨酸蛋白激酶。在这里，我们证明了两个基本的Vira残基，His-474和Gly-657的突变可以通过形成混合的异二聚体来补充，这表明Vira二聚体的每个亚基都能转磷酸化相反的亚基。病毒包含一个能抑制激酶活性的受体结构域。我们使用强迫异二聚体系统来证明病毒二聚体的两个接收域独立起作用，并且每个接收域都抑制相反亚基的磷受体亚域。我们还证明，共表达组成性Vira突变体和野生型Vira的分二倍体菌株的VIR基因表达水平远远低于仅表达构成等位基因的单倍体菌株。野生型VirA可以在没有酚类信号的情况下主动阻断VIR基因的表达，这一事实表明它可能具有磷酸化病毒磷酸酶活性。VirA的接收域是这一活性所必需的，而激活域的H474和G657残基不是必需的。共表达一个组成性的VirA等位基因和一个激酶失活但擅长抑制活性的等位基因的大二倍体菌株表现出强烈的诱导VIR基因表达，表明抑制活性受环境信号的调节。
The VirA-VirG two-component system regulates the 30-gene vir regulon in response to host-released chemical signals. VirA is a homodimeric membrane-spanning histidine protein kinase. Here, we show that mutations in two essential VirA residues, His-474 and Gly-657, can be complemented by the formation of mixed heterodimers, indicating that each subunit of a VirA dimer transphosphorylates the opposite subunit. VirA contains a receiver domain that inhibits kinase activity. We use the forced heterodimer system to show that the two receiver domains of a VirA dimer act independently and that each inhibits the phosphoacceptor subdomain of the opposite subunit. We also demonstrate that merodiploid strains co-expressing constitutive VirA mutants and wild-type VirA show levels of vir gene expression far lower than haploid strains expressing just the constitutive alleles. The fact that wild-type VirA can actively block vir gene expression in the absence of phenolic signals suggests that it might have a phospho-VirG phosphatase activity. The receiver domain of VirA is essential for this activity, whereas residues H474 and G657 of the kinase domain are not required. Merodiploid strains co-expressing a constitutive VirA allele and an allele that is kinase inactive but proficient in the inhibitory activity show strongly inducible vir gene expression, indicating that the inhibitory activity is modulated by environmental signals.