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Identification of Mycoplasma suis MSG1 interaction proteins on porcine erythrocytes
Yaning Zhang1, Yao Zou, Peipei Ma
1Key Laboratory of Bacteriology, Ministry of Agriculture, College of Veterinary Medicine, Nanjing Agricultural University, 210095, Nanjing, China.
Abstract:
Adhesion protein MSG1 mediating adherence to porcine erythrocytes in Mycoplasma suis (M. suis) invasion has been identified previously. In order to determine the host membrane proteins that interact with MSG1, recombinant His-tagged MSG1 (rMSG1) was used to screen for interacting proteins in the protein extracts of porcine erythrocyte membrane. Potential rMSG1-interacting proteins were initially identified as band 3 and β-actin with molecular weight of 46 and 45 kDa, respectively. Immune fluorescence results showed that rMSG1 can specifically bind with the β-actin of HeLa, BHK-21, and HEK-293A cells, respectively. RNA interference assays further demonstrated that the interaction between β-actin and rMSG1 on HeLa cells was specific and dose dependent. Confocal microscopy showed that both rMSG1 and M. suis can partially co-localize with β-actin on the surface of porcine erythrocytes. Pull-down assays showed that rMSG1 can directly interact with β-actin. Our study is the first to report the interaction of MSG1 with β-actin, which will be of help to understand the pathogenesis of M. suis and develop a cultivation system.
Insights
Mycoplasma suis adhesion protein MSG1 directly interacts with host β-actin. This finding is crucial for understanding M. suis pathogenesis and developing new cultivation methods.
Area of Science:
- Microbiology
- Cell Biology
- Protein-Protein Interactions
Background:
- Mycoplasma suis (M. suis) causes disease in pigs, with its adhesion protein MSG1 previously identified as key for erythrocyte adherence.
- Understanding host-pathogen interactions is vital for controlling M. suis infections.
Purpose of the Study:
- To identify host membrane proteins interacting with M. suis MSG1.
- To elucidate the specific interaction between MSG1 and host cell components.
Main Methods:
- Recombinant His-tagged MSG1 (rMSG1) used for screening porcine erythrocyte membrane proteins.
- Immune fluorescence, RNA interference, confocal microscopy, and pull-down assays employed to validate interactions.
- Analysis of interactions with β-actin in various cell lines (HeLa, BHK-21, HEK-293A) and porcine erythrocytes.
Main Results:
- MSG1 was found to interact with porcine erythrocyte membrane proteins, including band 3 and β-actin.
- rMSG1 specifically binds to β-actin in multiple cell types.
- The interaction between MSG1 and β-actin is specific, dose-dependent, and direct.
- MSG1 and M. suis partially co-localize with β-actin on porcine erythrocytes.
Conclusions:
- This study establishes a direct interaction between M. suis MSG1 and host β-actin.
- This interaction provides novel insights into M. suis pathogenesis.
- The findings may aid in developing improved M. suis cultivation systems.
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