Cyrus Tang Hematology Center, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Prevention, Suzhou Medical College, Soochow University, Suzhou, China.
Cyrus Tang Hematology Center, Collaborative Innovation Center of Hematology, State Key Laboratory of Radiation Medicine and Prevention, Suzhou Medical College, Soochow University, Suzhou, China; NHC Key Laboratory of Thrombosis and Hemostasis, Jiangsu Institute of Hematology, The First Affiliated Hospital of Soochow University, Suzhou, China.
J Biol Chem. 2022 Dec;298(12):102643. doi: 10.1016/j.jbc.2022.102643. Epub 2022 Oct 26.
Transmembrane protease serine 2 (TMPRSS2) is a membrane-bound protease expressed in many human epithelial tissues, including the airway and lung. TMPRSS2-mediated cleavage of viral spike protein is a key mechanism in severe acute respiratory syndrome coronavirus 2 activation and host cell entry. To date, the cellular mechanisms that regulate TMPRSS2 activity and cell surface expression are not fully characterized. In this study, we examined two major post-translational events, zymogen activation and N-glycosylation, in human TMPRSS2. In experiments with human embryonic kidney 293, bronchial epithelial 16HBE, and lung alveolar epithelial A549 cells, we found that TMPRSS2 was activated via intracellular autocatalysis and that this process was blocked in the presence of hepatocyte growth factor activator inhibitors 1 and 2. By glycosidase digestion and site-directed mutagenesis, we showed that human TMPRSS2 was N-glycosylated. N-glycosylation at an evolutionarily conserved site in the scavenger receptor cysteine-rich domain was required for calnexin-assisted protein folding in the endoplasmic reticulum and subsequent intracellular trafficking, zymogen activation, and cell surface expression. Moreover, we showed that TMPRSS2 cleaved severe acute respiratory syndrome coronavirus 2 spike protein intracellularly in human embryonic kidney 293 cells. These results provide new insights into the cellular mechanism in regulating TMPRSS2 biosynthesis and function. Our findings should help to understand the role of TMPRSS2 in major respiratory viral diseases.
跨膜丝氨酸蛋白酶 2（TMPRSS2）是一种在许多人类上皮组织中表达的膜结合蛋白酶，包括气道和肺部。TMPRSS2 介导的病毒刺突蛋白裂解是严重急性呼吸综合征冠状病毒 2 激活和宿主细胞进入的关键机制。迄今为止，调节 TMPRSS2 活性和细胞表面表达的细胞机制尚未完全阐明。在这项研究中，我们研究了人类 TMPRSS2 的两种主要翻译后事件，酶原激活和 N-糖基化。在人胚肾 293、支气管上皮 16HBE 和肺肺泡上皮 A549 细胞的实验中，我们发现 TMPRSS2 通过细胞内自催化激活，而在存在肝细胞生长因子激活物抑制剂 1 和 2 的情况下，这一过程被阻断。通过糖苷酶消化和定点突变，我们表明人类 TMPRSS2 被 N-糖基化。在进化上保守的清道夫受体富含半胱氨酸结构域中的 N-糖基化对于内质网中钙连接蛋白辅助的蛋白质折叠以及随后的细胞内运输、酶原激活和细胞表面表达是必需的。此外，我们表明 TMPRSS2 在人胚肾 293 细胞中切割严重急性呼吸综合征冠状病毒 2 刺突蛋白。这些结果为调节 TMPRSS2 生物合成和功能的细胞机制提供了新的见解。我们的发现应该有助于理解 TMPRSS2 在主要呼吸道病毒疾病中的作用。
