Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells
Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells
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DOI：
10.1128/jvi.00234-15
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发表时间：
2015-07-01
影响因子：
5.4
通讯作者：
Schelhaas, Mario
中科院分区：
文献类型：
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作者：
Cerqueira, Carla;Ventayol, Pilar Samperio;Schelhaas, Mario
人乳头瘤病毒进入宿主细胞是一个复杂的过程。它涉及在细胞表面的构象变化，受体转换，内化的一种新的内吞机制，在核内体中的脱壳，运输的亚病毒复合体的高尔基体复合体，和有丝分裂期间的核进入。在这里，我们解决了如何稳定的人乳头瘤病毒16（HPV 16）的衣壳接触可能会被逆转，以允许病毒基因组的外壳。使用生物化学和细胞生物学分析，我们确定，主要衣壳蛋白L1进行蛋白水解裂解过程中进入。除了在从核内体中的亚病毒复合物中去除衣壳后可能发生的α-组织蛋白酶介导的蛋白水解外，还观察到L1的至少两个进一步的蛋白水解裂解，其中一个与核内体的低pH环境无关。这种分裂发生在细胞外。进一步的分析表明，负责的蛋白酶是分泌的胰蛋白酶样丝氨酸蛋白酶激肽释放酶-8（KLK 8），参与表皮稳态和伤口愈合。感染所需的裂解是由病毒颗粒与硫酸乙酰肝素蛋白聚糖的预先相互作用促进的。KLK 8介导的切割对于暴露次要衣壳蛋白L2的重要表位的进一步构象变化至关重要。与介导L2暴露的亲环蛋白和弗林蛋白酶无关，KLK 8介导的L1切割可能促进了进入位于衣壳腔中的L2，并可能使其脱膜。由于HPV 6和HPV 18也需要KLK 8才能进入，因此我们认为KLK 8依赖性进入步骤是保守的。重要信息我们对进入的HPV 16（宫颈癌的病原体）的蛋白水解加工的分析表明，衣壳在细胞外被一种在伤口愈合期间活跃的丝氨酸蛋白酶切割，并且这种切割对于感染至关重要。L1的裂解是至少四种结构改变之一，这些结构改变使病毒在细胞外进行受体转换、内化和可能的脱壳。这一步骤对HPV 6和HPV 18也很重要，这可能表明它在乳头瘤病毒中是保守的。这项研究推进了对HPV 16最初如何感染细胞的理解，加强了创伤促进表皮组织感染的概念，并可能有助于开发抗病毒措施。
The entry of human papillomaviruses into host cells is a complex process. It involves conformational changes at the cell surface, receptor switching, internalization by a novel endocytic mechanism, uncoating in endosomes, trafficking of a subviral complex to the Golgi complex, and nuclear entry during mitosis. Here, we addressed how the stabilizing contacts in the capsid of human papillomavirus 16 (HPV16) may be reversed to allow uncoating of the viral genome. Using biochemical and cell-biological analyses, we determined that the major capsid protein L1 underwent proteolytic cleavage during entry. In addition to a dispensable cathepsin-mediated proteolysis that occurred likely after removal of capsomers from the subviral complex in endosomes, at least two further proteolytic cleavages of L1 were observed, one of which was independent of the low-pH environment of endosomes. This cleavage occurred extracellularly. Further analysis showed that the responsible protease was the secreted trypsin-like serine protease kallikrein-8 (KLK8) involved in epidermal homeostasis and wound healing. Required for infection, the cleavage was facilitated by prior interaction of viral particles with heparan sulfate proteoglycans. KLK8-mediated cleavage was crucial for further conformational changes exposing an important epitope of the minor capsid protein L2. Occurring independently of cyclophilins and of furin that mediate L2 exposure, KLK8-mediated cleavage of L1 likely facilitated access to L2, located in the capsid lumen, and potentially uncoating. Since HPV6 and HPV18 also required KLK8 for entry, we propose that the KLK8-dependent entry step is conserved.IMPORTANCEOur analysis of the proteolytic processing of incoming HPV16, an etiological agent of cervical cancer, demonstrated that the capsid is cleaved extracellularly by a serine protease active during wound healing and that this cleavage was crucial for infection. The cleavage of L1 is one of at least four structural alterations that prime the virus extracellularly for receptor switching, internalization, and possibly uncoating. This step was also important for HPV6 and HPV18, which may suggest that it is conserved among the papillomaviruses. This study advances the understanding of how HPV16 initially infects cells, strengthens the notion that wounding facilitates infection of epidermal tissue, and may help the development of antiviral measures.