Construction of a screening system for key intracellular survival proteins of macrophages of Staphylococcus aureus
10.16016/j.2097-0927.202311091
- VernacularTitle:金黄色葡萄球菌巨噬细胞胞内生存关键分泌蛋白筛选体系的构建
- Author:
Yaojia SHI
1
;
Tian TIAN
;
Tingrong XIONG
;
Yu WANG
;
Xiaokai ZHANG
;
Quanming ZOU
Author Information
1. 475004 河南开封,河南大学药学院
- Keywords:
Staphylococcus aureus;
macrophages;
intracellular infection;
secretory protein
- From:
Journal of Army Medical University
2024;46(8):815-821
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a high-throughput screening system to obtain key Staphylococcus aureus (S.aureus)secretory proteins which required for S.aureus survival in macrophages.Methods Based on our validated eukaryotic expression vector library of S.aureus secretory proteins,DNA transfection was used to obtain an RAW264.7 macrophage array expressing S.aureus secretory proteins.After the RAW264.7 cells were infected with S.aureus,the extracellular bacteria were removed to observe the intracellular surviving situation of S.aureus.Finally,the screening results were validated by the overexpression and knockout S.aureus of corresponding secretory proteins.Results The optimal transfection dose (1.0 μg/well)of plasmids for RAW264.7,multiplicity of infection (MOI,1 .0 ),and infection time (4 h after removing extracellular bacteria of S.aureus ) were established respectively.To validate the screening results,the corresponding overexpression and knockout strains were constructed.And hypothetical protein and Serine protease E were found to promote the survival of intracellular S.aureus.Conclusion We successfully construct a screening system for key secreted secretory proteins which required for S.aureus surviving in macrophages,which may advance the study of the intracellular surviving mechanism of S.aureus.
