A new means to identify type 3 secreted effectors: functionally interchangeable class IB chaperones recognize a conserved sequence.
A new means to identify type 3 secreted effectors: functionally interchangeable class IB chaperones recognize a conserved sequence.
复制标题
作者：
Costa SC;Schmitz AM;Jahufar FF;Boyd JD;Cho MY;Glicksman MA;Lesser CF
许多革兰氏阴性菌利用专门的分泌系统将蛋白质（效应器）直接注入宿主细胞。关于细菌如何确保它们编码的数千种蛋白质中只有一小部分被识别为分泌系统的底物，人们知之甚少，这限制了通过生物信息学分析对它们的识别。许多这些蛋白质需要伴侣来指导它们的分泌。在这里，使用新描述的蛋白质相互作用平台实验，我们证明了来自一种细菌的3型分泌系统IB类伴侣蛋白直接结合它们自己的效应物以及来自其他物种的效应物。此外，我们观察到包括病原体和内共生体在内的7个物种的IB类同源物的表达，介导了志贺氏菌效应物直接转运到宿主细胞中，这表明IB类伴侣蛋白通常在功能上是可互换的。值得注意的是，IB类伴侣蛋白结合了许多效应物。然而，正如之前提出的，它们不是混杂的；相反，它们识别一个已定义的序列，我们将其称为保守伴侣结合域（CCBD）序列[(lif)1XXX(IV)5XX(IV)8X(N)10]。该序列是第一个确定的氨基酸序列，用于任何种间细菌分泌系统，即直接将蛋白质传递到真核细胞的系统。该序列为鉴定III型分泌系统的底物提供了一种新的手段。事实上，使用CCBD序列的模式搜索算法，我们已经确定了来自内共生体Sodalis舌索菌的前两个可能的效应物。许多革兰氏阴性病原体利用3型分泌系统将数十种效应物输送到宿主细胞中。为了了解这些生物引起疾病的不同方式，有必要确定它们的效应物，其中许多需要分泌伴侣。在这里，我们确定IB类伴侣不是混杂的，正如之前提出的，而是识别一个保守的效应序列。我们证明了基于此定义序列的模式搜索算法可以用于识别以前未知的效应器。此外，我们观察到至少7种细菌的IB类伴侣蛋白在功能上是可互换的。它们不仅结合并介导自身效应物向宿主细胞的传递，而且还结合来自其他细菌的3型底物，这表明阻断伴侣-效应物相互作用的抑制剂可能提供一种有效治疗革兰氏阴性病原体感染的新方法，包括对现有抗生素具有耐药性的生物体。
Many Gram-negative bacteria utilize specialized secretion systems to inject proteins (effectors) directly into host cells. Little is known regarding how bacteria ensure that only small subsets of the thousands of proteins they encode are recognized as substrates of the secretion systems, limiting their identification through bioinformatic analyses. Many of these proteins require chaperones to direct their secretion. Here, using the newly described protein interaction platform assay, we demonstrate that type 3 secretion system class IB chaperones from one bacterium directly bind their own effectors as well as those from other species. In addition, we observe that expression of class IB homologs from seven species, including pathogens and endosymbionts, mediate the translocation of effectors from Shigella directly into host cells, demonstrating that class IB chaperones are often functionally interchangeable. Notably, class IB chaperones bind numerous effectors. However, as previously proposed, they are not promiscuous; rather they recognize a defined sequence that we designate the conserved chaperone-binding domain (CCBD) sequence [(LMIF)1XXX(IV)5XX(IV)8X(N)10]. This sequence is the first defined amino acid sequence to be identified for any interspecies bacterial secretion system, i.e., a system that delivers proteins directly into eukaryotic cells. This sequence provides a new means to identify substrates of type III secretion systems. Indeed, using a pattern search algorithm for the CCBD sequence, we have identified the first two probable effectors from an endosymbiont, Sodalis glossinidius. Many Gram-negative pathogens utilize type 3 secretion systems to deliver tens of effectors into host cells. In order to understand the diverse ways that these organisms cause disease, it is necessary to identify their effectors, many of which require chaperones to be secreted. Here we establish that class IB chaperones are not promiscuous, as previously proposed, but rather recognize a conserved effector sequence. We demonstrate that pattern search algorithms based on this defined sequence can be used to identify previously unknown effectors. Furthermore, we observe that class IB chaperones from at least seven bacterial species are functionally interchangeable. Not only do they bind and mediate the delivery of their own set of effectors into host cells but they also bind to type 3 substrates from other bacteria, suggesting that inhibitors that block chaperone-effector interactions could provide a novel means to effectively treat infections due to Gram-negative pathogens, including organisms resistant to currently available antibiotics.