Chen Yunxin, Liu Li, Wei Qiang, Zhu Hua, Jiang Hong, Tu Xinming, Qin Chuan, Chen Zhiwei
Institute of Laboratory Animal Science, Chinese Academy of Medical Sciences and Peking Union Medical College, No.5, Panjiayuan, Nanli, Chaoyang District, Beijing, China.
Virology. 2008 Nov 10;381(1):89-97. doi: 10.1016/j.virol.2008.08.016. Epub 2008 Sep 17.
Angiotensin converting enzyme 2 (ACE2) is the receptor that severe acute respiratory syndrome coronavirus (SARS-CoV) utilizes for target cell entry and, therefore, plays an important role in SARS pathogenesis. Since Chinese rhesus (rh) macaques do not usually develop SARS after SARS-CoV infection, it has been suggested that rh-ACE2 probably does not support viral entry efficiently. To determine the role of rh-ACE2 in early lung pathogenesis in vivo, we studied eleven Chinese rhesus monkeys experimentally infected with a pathogenic SARS-CoV(PUMC01) strain. Rh-ACE2 genes were amplified from all animals by reverse transcription polymerase chain reaction, and their function was studied in vitro using a pseudovirus entry assay. Many natural non-synonymous (NS) changes were found in rh-ACE2 genes. Compared to human (hu) ACE2, thirty-eight consensus NS changes were found in rh-ACE2. Since these changes do not interact with the receptor binding domain of SARS-CoV, rh-ACE2 in general is as effective as human homolog in supporting viral entry. Rh-ACE2, however, is more polymorphic than hu-ACE2. Additional sporadic NS substitutions in clone Rh11-7 reduced the level of rh-ACE2 protein expression and did not support viral entry effectively. Further mutagenesis analysis showed that a natural mutation Y217N dramatically alters ACE2 expression and entry efficiency. Moreover, introduction of the Y217N mutation into hu-ACE2 caused the down-regulation of expression and reduced viral entry efficiency. These results indicate that the Y217N mutation plays a role in modulating SARS-CoV infection. Our results provide insights for understanding the role of rh-ACE2 in SARS lung pathogenesis in a non-human primate model.
血管紧张素转换酶2（ACE2）是严重急性呼吸综合征冠状病毒（SARS-CoV）用于进入靶细胞的受体，因此在SARS发病机制中起重要作用。由于中国恒河猴在感染SARS-CoV后通常不会患上SARS，因此有人提出恒河猴ACE2（rh-ACE2）可能无法有效地支持病毒进入。为了确定rh-ACE2在体内早期肺部发病机制中的作用，我们研究了11只实验感染致病性SARS-CoV（PUMC01）毒株的中国恒河猴。通过逆转录聚合酶链反应从所有动物中扩增rh-ACE2基因，并使用假病毒进入试验在体外研究其功能。在rh-ACE2基因中发现了许多自然的非同义（NS）变化。与人类（hu）ACE2相比，在rh-ACE2中发现了38个一致的NS变化。由于这些变化不与SARS-CoV的受体结合域相互作用，因此rh-ACE2在支持病毒进入方面通常与人类同源物一样有效。然而，rh-ACE2比hu-ACE2具有更多的多态性。克隆Rh11-7中的额外散发性NS替代降低了rh-ACE2蛋白表达水平，并且不能有效地支持病毒进入。进一步的诱变分析表明，自然突变Y217N显著改变了ACE2的表达和进入效率。此外，将Y217N突变引入hu-ACE2导致表达下调并降低了病毒进入效率。这些结果表明Y217N突变在调节SARS-CoV感染中起作用。我们的结果为理解rh-ACE2在非人类灵长类动物模型中SARS肺部发病机制中的作用提供了见解。
