血管紧张素转换酶2（angiotensin-converting enzyme 2, ACE2）是新型冠状病毒（severe acute respiratory syndrome coronavirus 2, SARS-CoV-2）进入细胞的受体，其表达水平与SARS-CoV-2的感染密切相关。多项研究报道ACE2可以调控细胞增殖，然而，目前关于细胞增殖状态如何影响ACE2的表达水平还知之甚少。在药物再利用研究中，参与细胞周期调控的CDK4/6抑制剂palbociclib是治疗COVID-19（coronavirus disease 2019）的最高评分的药物之一。本研究旨在探讨palbociclib和其他细胞周期阻滞化合物对ACE2表达和后续病毒感染的调节作用，以为治疗COVID-19提供依据。本实验使用了四种方法来实现多个细胞系的细胞周期阻滞。在转录水平上，使用RT-qPCR检测ACE2 mRNA的水平变化，进一步利用RT-qPCR、western blot、双荧光素酶报告基因实验探讨生信分析预测的STAT3、c-Jun转录因子对于ACE2的转录调控。在蛋白水平上，利用western blot、免疫荧光、流式细胞术、生信分析等方法对ACE2的蛋白降解途径进行了深入的研究。最后，在Huh-7和Vero E6两种细胞中利用SARS-CoV-2假病毒感染模型探讨了细胞周期阻滞药物对SARS-CoV-2感染的影响。结果显示，palbociclib和其他细胞周期阻滞化合物可以使细胞周期阻滞在G0/G1期，并上调ACE2的mRNA和蛋白水平，而不改变其亚细胞定位。mRNA表达调控方面，palbociclib等细胞周期阻滞方法可以上调STAT3、c-Jun的表达从而激活ACE2启动子活性，上调其mRNA表达水平。蛋白表达调控方面，palbociclib通过下调S期激酶相关蛋白2（S-phase kinase-associated protein 2, SKP2）抑制ACE2的泛素-蛋白酶体和溶酶体途径的蛋白降解。此外，细胞周期阻滞诱导ACE2表达增加，促进了SARS-CoV-2感染。综上所述，本研究证明了ACE2的表达在增殖细胞中下调，细胞周期抑制剂可以通过升高mRNA水平和减少蛋白降解来上调ACE2的蛋白表达水平，促进SARS-CoV-2的感染，这提示细胞周期抑制剂可能不适合作为SARS-CoV-2感染的治疗药物。

As the receptor for SARS-CoV-2 entry, expression level of ACE2 is closely related to SARS-CoV-2 infection. Multiple studies have shown that ACE2 could regulate cell proliferation. However, little is known about how cell proliferation state affects the expression level of ACE2. CDK4/6 inhibitor palbociclib was reported to be one of the top-scored repurposed drugs to treat COVID-19. In this study, we aimed to investigate the effect of palbociclib and other cell cycle arresting compounds on the regulation of ACE2 expression and subsequent viral infection, providing a basis for the treatment of COVID-19. Four methods were used to achieve cell cycle arrest in multiple cell lines. At the transcriptional level, we used RT-qPCR to detect changes in ACE2 mRNA levels, and further explored the transcriptional regulation of STAT3 and c-Jun transcription factors predicted by bioinformatics analysis tools on ACE2 using RT-qPCR, western blot, and dual luciferase reporter gene assays. At the protein level, we conducted an in-depth study of the protein degradation pathway of ACE2 using western blot, immunofluorescence, flow cytometry, and bioinformatics analysis. Finally, we explored the effects of cell cycle arresting drugs on SARS-CoV-2 infection using SARS-CoV-2 pseudoviral infection model in both Huh-7 and Vero E6 cells. We demonstrated that palbociclib and other methods could arrest cells in G0/G1 phase and up-regulate ACE2 mRNA and protein levels without altering its subcellular localization. At the transcriptional level, palbociclib and other cell cycle arresting compounds could upregulate expression levels of STAT3 and c-Jun, followed by activation of ACE2 promoter and elevation of ACE2 mRNA levels. At the protein level, palbociclib inhibited ubiquitin-proteasome and lysosomal degradation of ACE2 through down-regulating SKP2. In addition, increased ACE2 expression induced by cell cycle arrest facilitated SARS-CoV-2 pseudoviral infection. In conclusion, we demonstrated that ACE2 expression was down-regulated in proliferating cells. Cell cycle inhibitors could increase ACE2 expression through promoting mRNA levels and inhibiting protein degradation of ACE2 and facilitate SARS-CoV-2 infection, which might not work as effective therapeutic agents to treat SARS-CoV-2 infection.