An understanding of how viruses interact with their receptors is vital as this step is a major determinant of host susceptibility and disease. The enterovirus coxsackievirus A9 (CVA9) is an important pathogen responsible for respiratory infections, myocarditis, infections of the central nervous system, chronic dilated cardiomyopathy and possibly type I diabetes. CVA9 harbours an integrin- recognition motif, RGD (Arg-Gly-Asp), in the capsid protein VP1 and this motif is believed to be primarily responsible for binding to integrins αvβ6 and/or αvβ3 during cell entry. Despite the consistent conservation of RGD-flanking amino acids in multiple RGD-containing picornaviruses, the significance of these amino acids to cell tropism has not been thoroughly investigated. In this study we used 10 CVA9 mutants and a panel of cells to analyse cell tropism. We showed that CVA9 infection proceeds by either an RGD- dependent or an apparently RGD- independent pathway. Differences in the amino acid found at the +1 position of the RGD motif affect the cell tropism of CVA9 when an RGD- dependent pathway is used. Naturally occurring CVA9 isolates have either the sequence RGDM and RGDL and we found that the corresponding viruses in our panel infected cells most efficiently. There was also a strong selection pressure for RGDL in adaptation experiments. However, there was also an unexpected selection of an RGDL variant in an apparently RGD- independent cell line. There was also no simple relationship between infection of cells and expression of integrins αvβ3 and αvβ6. The results obtained have greatly improved our understanding of how CVA9 infects cells. This will be useful in the design of antivirus drugs and also gives a framework for the modification of CVA9 or other RGD containing picornaviruses for specific targeting of cancer cells for oncolytic therapy.

了解病毒如何与其受体相互作用至关重要，因为这一步骤是宿主易感性和疾病的主要决定因素。肠道病毒柯萨奇病毒 A9 (CVA9) 是导致呼吸道感染、心肌炎、中枢神经系统感染、慢性扩张型心肌病和可能的 I 型糖尿病的重要病原体。CVA9 在衣壳蛋白 VP1 中含有整合素识别基序 RGD (Arg-Gly-Asp)，据信该基序主要负责在细胞进入过程中与整合素 αvβ6 和/或 αvβ3 结合。尽管在多个含有 RGD 的小核糖核酸病毒中 RGD 侧翼氨基酸的保守性是一致的，但这些氨基酸对细胞嗜性的重要性尚未得到彻底研究。在这项研究中，我们使用了 10 个 CVA9 突变体和一组细胞来分析细胞嗜性。我们发现 CVA9 感染通过依赖 RGD 或明显不依赖 RGD 的途径进行。当使用 RGD 依赖性途径时，在 RGD 基序 +1 位置发现的氨基酸差异会影响 CVA9 的细胞嗜性。天然存在的 CVA9 分离株具有 RGDM 和 RGDL 序列，我们发现我们小组中的相应病毒最有效地感染细胞。在适应实验中，RGDL 也有很强的选择压力。然而，在一个明显不依赖于 RGD 的细胞系中，也有一个意想不到的 RGDL 变体选择。细胞感染与整合素αvβ3和αvβ6的表达之间也没有简单的关系。获得的结果大大提高了我们对 CVA9 如何感染细胞的理解。