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2014年7月,吴庆宇博士和何杨课题组J Biol Chem上发表文章

发布日期:2014/07/22

 

2014年7月11日,吴庆宇博士课题组在J Biol Chem(2014;289:19500-7)杂志上在线发表题为“N-Glycosylation Is Required for Matriptase-2 Autoactivation and Ectodomain Shedding.”的文章。

 

作者:

Jiang Jiang1,2, Jianfeng Yang1, Ping Feng2, Bin Zuo1, Ningzheng Dong3, Qingyu Wu4*,  Yang He3*

 

1Cyrus Tang Hematology Center and Ministry of Education Engineering Center of Hematological Disease, Soochow University, Suzhou 215123, China.

2Department of Clinic Laboratories, the Second Affiliated Hospital, Soochow University, Suzhou 215004, China.

3Cyrus Tang Hematology Center and Ministry of Education Engineering Center of Hematological Disease, Ministry of Health Key Laboratory of Thrombosis and Hemostasis, Jiangsu Institute of Hematology, the First Affiliated Hospital, and the Collaborative Innovation Center of Hematology, Soochow University, Suzhou 215006, China.

4the Cyrus Tang Hematology Center and MOE Engineering Center of Hematological Disease, Collaborative Innovation Center of Hematology, Soochow University, Suzhou 215123, China, Molecular Cardiology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195.

*Corresponding author

 

摘要:

Matriptase-2 is a hepatic membrane serine protease that regulates iron homeostasis. Defects in matriptase-2 cause iron deficiency anemia. In cells, matriptase-2 is synthesized as a zymogen. To date, how matriptase-2 expression and activation are regulated remains poorly understood. Here we expressed human matriptase-2 in HEK293 and hepatic BEL-7402,SMMC-7721, and QGY-7703 cells. By labeling cell surface proteins and Western analysis, we examined matriptase-2 cell surface expression, zymogen activation, and ectodomain shedding. Our results show that matriptase-2 was activated on the cell surface but not intracellularly. Activated matriptase-2 underwent ectodomain shedding, producing soluble fragments in the conditioned medium. By testing inactive mutants, R576A and S762A, we found that matriptase-2 activation and shedding were mediated by its own catalytic activity and that the one-chain form of matriptase-2 had little activity in ectodomain shedding. We made additional matriptase-2 mutants, N136Q, N184Q, N216Q, N338Q, N433Q, N453Q, and N518Q, in which each of the predicted N-glycosylation sites was mutated. All of these mutants were expressed on the cell surface. However, mutants N216Q, N453Q, and N518Q, but not the other mutants, had impaired zymogen activation and ectodomain shedding. Our results indicate that N-glycans at specific sites are critical for matriptase-2 activation. Together, these data provide new insights into the cell surface expression, zymogen activation, and ectodomain shedding of matriptase-2.

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