ACE2蛋白Cys141、Cys344和Cys498位点的棕榈酰化修饰促进其在细胞外囊泡的定位

doi:10.16352/j.issn.1001-6325.2025.09.1151

基础医学与临床 ›› 2025, Vol. 45 ›› Issue (9): 1151-1157.doi: 10.16352/j.issn.1001-6325.2025.09.1151

ACE2蛋白Cys141、Cys344和Cys498位点的棕榈酰化修饰促进其在细胞外囊泡的定位

阳晶晶¹, 马艳¹, 王悦¹, 孙阳², 李盼^2*, 郭峰^1*

1.徐州医科大学基础医学院江苏省免疫与代谢重点实验室病原生物学与免疫学教研室,江苏徐州 221004;
2.徐州医科大学细胞治疗产业学院江苏省肿瘤生物治疗研究所,江苏徐州 221121

收稿日期:2025-06-03 修回日期:2025-07-09 出版日期:2025-09-05 发布日期:2025-08-27
通讯作者: ^*PanLi@xzhmu.edu.cn; Feng.Guo@xzhmu.edu.cn
基金资助:
国家自然科学基金(82370051,82373905);江苏省研究生科研与实践创新计划项目(KYCX23_2944);基础医学国家级实验教学示范中心学生科创项目(2024BMS29)

Palmitoylation of ACE2 at Cys141, Cys344, and Cys498 facilitates its extracellular vesicles localization

YANG Jingjing¹, MA Yan¹, WANG Yue¹, SUN Yang², LI Pan^2*, GUO Feng^1*

1. Jiangsu Province Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Basic Medical School, Xuzhou Medical University, Xuzhou 221004;
2. State Key Laboratory Jiangsu Center for the Collaboration and Innovation of Cancer Biotherapy, Cancer Institute, Xuzhou Medical University, Xuzhou 221121, China

Received:2025-06-03 Revised:2025-07-09 Online:2025-09-05 Published:2025-08-27
Contact: ^*PanLi@xzhmu.edu.cn; Feng.Guo@xzhmu.edu.cn

摘要/Abstract

摘要： 目的鉴定严重急性呼吸系统综合征冠状病毒(SARS-CoV)和SARS-CoV-2细胞受体——血管紧张素转化酶2(ACE2)的关键S-棕榈酰化修饰位点,并探讨该修饰对ACE2定位和功能的影响。方法通过多位点突变策略,将ACE2蛋白的8个半胱氨酸(Cys)残基全部突变为丝氨酸,获得无棕榈酰化能力的突变体(8CS)。随后在该背景下逐一回突单个半胱氨酸残基,通过点击化学结合生物正交标记方法,检测各突变体的棕榈酰化修饰水平,筛选可能发生棕榈酰化的关键位点。进一步结合免疫荧光染色及细胞外囊泡提取实验,评估各修饰位点对ACE2定位的影响。结果优化了基于点突变技术的多位点棕榈酰化筛选体系,鉴定出ACE2蛋白的3个关键棕榈酰化位点:Cys141、Cys344 和 Cys498。功能研究表明,这些位点的棕榈酰化修饰显著促进ACE2向细胞外囊泡的富集。结论 ACE2蛋白Cys141、Cys344 和 Cys498位点发生的S-棕榈酰化修饰是其靶向细胞外囊泡定位的重要因素,提示其可能在病毒受体呈递和ACE2相关信号通路中发挥重要作用。

关键词: S-棕榈酰化, 多点突变, 翻译后修饰, 细胞外囊泡, 血管紧张素转化酶2(ACE2)

Abstract: Objective To identify S-palmitoylation sites on angiotensin-converting enzyme 2(ACE2), the cellular receptor for severe acute respiratory syndrome corona virus(SARS-CoV) and SARS-CoV-2, and to investigate the functional relevance of these modifications in regulating ACE2 localization and activity. Methods An optimized multi-site mutagenesis strategy was performed by simultaneously substitution of all eight cysteine (Cys) residuesin ACE2 by serine to create a non-palmitoylatable mutant (8CS). Then individual cysteine residues were re-introduced one by one. Palmitoylation level of mutants was evaluated using a bioorthogonal click chemistry method to identify palmitoylation-competent residues. Immune-fluorescence staining and extra-cellular vesicle isolation assays were then used to evaluate the impact of specific palmitoylation sites on ACE2 sub-cellular localization. Results An efficient strategy for multi-site palmitoylation site mapping was optimized and successfully identified three critical palmitoylation sites on ACE2: Cys141, Cys344 and Cys498. Functional analyses showed that palmitoylation of these sites significantly promotes the enrichment of ACE2 in extra-cellular vesicles. Conclusions S-palmitoylation at Cys141, Cys344 and Cys498 is essential for the trafficking of ACE2 to extra-cellular vesicles, which suggests a potential regulatory mechanism of its impact on viral receptor presentation and ACE2-associated signaling pathways.

Key words: S-palmitoylation, multi-site mutagenesis, post-translational modification, extra cellular vesicle, angiotensin-converting enzyme 2(ACE2)

中图分类号:

阳晶晶, 马艳, 王悦, 孙阳, 李盼, 郭峰. ACE2蛋白Cys141、Cys344和Cys498位点的棕榈酰化修饰促进其在细胞外囊泡的定位[J]. 基础医学与临床, 2025, 45(9): 1151-1157.

YANG Jingjing, MA Yan, WANG Yue, SUN Yang, LI Pan, GUO Feng. Palmitoylation of ACE2 at Cys141, Cys344, and Cys498 facilitates its extracellular vesicles localization[J]. Basic & Clinical Medicine, 2025, 45(9): 1151-1157.

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ACE2蛋白Cys141、Cys344和Cys498位点的棕榈酰化修饰促进其在细胞外囊泡的定位

Palmitoylation of ACE2 at Cys141, Cys344, and Cys498 facilitates its extracellular vesicles localization

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