Helicobacter pylori type IV secretion apparatus exploits beta1 integrin in a novel RGD-independent manner.
Helicobacter pylori type IV secretion apparatus exploits beta1 integrin in a novel RGD-independent manner.
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DOI：
10.1371/journal.ppat.1000684
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发表时间：
2009-12
期刊：
影响因子：
6.7
通讯作者：
Haas R
中科院分区：
文献类型：
--
作者：
Jiménez-Soto LF;Kutter S;Sewald X;Ertl C;Weiss E;Kapp U;Rohde M;Pirch T;Jung K;Retta SF;Terradot L;Fischer W;Haas R
幽门螺杆菌（Hp）细胞毒素相关基因A （CagA）效应蛋白通过cag- IV型分泌系统（T4SS）易位进入宿主细胞是包括胃癌在内的严重胃疾病的主要危险因素。然而，转运的机制和宿主细胞对该事件的要求尚不清楚。T4SS由横跨内外膜的Cag蛋白复合物和位于表面的菌毛组成。此前有报道称，CagL与α5β1整合素的典型整合素/配体型相互作用对CagA易位至关重要。在这里，我们报道了t4ss -pilus相关成分cgy和效应蛋白CagA与宿主细胞β1整合素受体的特异性结合。表面等离子体共振测量显示，与rgd依赖性整合素/纤维连接蛋白相互作用（KD为15 nM）相比，CagA与α5β1整合素的结合相当强（解离常数，KD为0.15 nM）。对于CagA易位，β1整合素亚基的细胞外部分是必需的，但不是其细胞质结构域，也不是通过整合素连接激酶的下游信号传导。一组针对各种已定义的β1整合素表位（如PSI、I-like、EGF或β-尾结构域）的β1整合素特异性单克隆抗体无法干扰CagA易位。然而，一种特异性抗体（9EG7）稳定了β1整合素异源二聚体的开放活性构象，有效地阻断了CagA易位。我们的数据支持一种新的模型，其中，cage - t4ss通过rgd独立的相互作用利用β1整合素受体，包括从开放（延伸）到封闭（弯曲）构象的构象转换，从而启动效应蛋白易位。整合素是存在于几乎所有类型细胞中的单一跨膜蛋白。它们由α和β亚基组成，它们共同形成配体结合袋，能够与细胞外基质蛋白相互作用。整合素配体上最著名的结合域是RGD结构域。许多细菌和病毒病原体利用这个配体结合结构域与宿主细胞上的整合素相互作用。幽门螺杆菌是一种与胃部疾病相关的常见细菌病原体，最近也被列入了这一名单。与胃病理相关的幽门螺杆菌最重要的因素之一是CagA蛋白。该蛋白通过Cag IV型分泌系统（Cag - t4ss）直接注入宿主细胞。先前的研究表明，cag-T4SS需要整合素才能将CagA注射（易位）到细胞中。我们提供的证据表明，三种蛋白质，CagA， CagI和CagY，以不依赖于rgd的方式与整合素相互作用。此外，我们的数据指出，Cag装置需要β1整合素异源二聚体的物理能力才能从活性/延伸构象转变为封闭/弯曲构象。这种新型的整合素相互作用开辟了病原体利用细胞受体的新途径。
Translocation of the Helicobacter pylori (Hp) cytotoxin-associated gene A (CagA) effector protein via the cag-Type IV Secretion System (T4SS) into host cells is a major risk factor for severe gastric diseases, including gastric cancer. However, the mechanism of translocation and the requirements from the host cell for that event are not well understood. The T4SS consists of inner- and outer membrane-spanning Cag protein complexes and a surface-located pilus. Previously an arginine-glycine-aspartate (RGD)-dependent typical integrin/ligand type interaction of CagL with α5β1 integrin was reported to be essential for CagA translocation. Here we report a specific binding of the T4SS-pilus-associated components CagY and the effector protein CagA to the host cell β1 Integrin receptor. Surface plasmon resonance measurements revealed that CagA binding to α5β1 integrin is rather strong (dissociation constant, KD of 0.15 nM), in comparison to the reported RGD-dependent integrin/fibronectin interaction (KD of 15 nM). For CagA translocation the extracellular part of the β1 integrin subunit is necessary, but not its cytoplasmic domain, nor downstream signalling via integrin-linked kinase. A set of β1 integrin-specific monoclonal antibodies directed against various defined β1 integrin epitopes, such as the PSI, the I-like, the EGF or the β-tail domain, were unable to interfere with CagA translocation. However, a specific antibody (9EG7), which stabilises the open active conformation of β1 integrin heterodimers, efficiently blocked CagA translocation. Our data support a novel model in which the cag-T4SS exploits the β1 integrin receptor by an RGD-independent interaction that involves a conformational switch from the open (extended) to the closed (bent) conformation, to initiate effector protein translocation. Integrins are single transmembrane proteins present on almost all types of cells. They are composed of an α and a β subunit, which together form the ligand binding pocket, able to interact with extracellular matrix proteins. The best known binding domain on integrin ligands is the RGD domain. Many bacterial, but also viral pathogens exploit this ligand-binding domain to interact with integrins on the host cell. Helicobacter pylori, a common bacterial pathogen associated with gastric diseases, was recently added to this list. One of H. pylori's most important factors associated with gastric pathologies is the CagA protein. This protein is directly injected into host cells through the Cag Type IV Secretion System (cag-T4SS). Previous studies demonstrated that the cag-T4SS requires integrins for the injection (translocation) of CagA into cells. We provide evidence that three proteins, CagA, CagI and CagY, interact with integrins in an RGD-independent way. Additionally, our data point out that the Cag apparatus needs the physical capacity of a β1 integrin heterodimer to change from an active/extended conformation to a closed/bent conformation. This novel kind of integrin interaction opens a new way in which pathogens can use receptors on cells.
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