日本質量分析学会 第66回質量分析総合討論会

Program

Poster Presentations

Day 3, May 17(Thu.)  Poster

A method based on two-dimensional fluorescence difference gel electrophoresis to analyze an ectodomain shedding

(St. Marianna Univ. School of Medicine)
oKazuki Omoteyama, Toshiyuki Sato, Atsuhiro Tsuchiya, Masaaki Sato, Mitsumi Arito, Naoya Suematsu, Manae Kurokawa, Tomohiro Kato

[Objective] Cell surface proteins digested at their extracellular domains and released from the surface (ectodomain shedding) are possibly associated with pathophysiology of various diseases. There has been no method to comprehensively detect these shedded proteins. Previously, we reported a method for detection of the proteins. In the current study, we tried to analyze the proteins by 2D-DIGE. [Methods] We used a human cervical carcinoma cell line of HeLa which were previously reported to express A Disintegrin And Metalloprotease 17 (ADAM17). Cell surface proteins of living Hela cells were labeled by Cy5 dye and then the cells were cultured in a medium containing TAPI-2 (ADAM17 inhibitor). As an internal control, cell surface proteins of which were labeled with Cy3 dye were cultured. Proteins in the cultured media were isolated and then were analyzed by 2D-DIGE. [Results] Many protein spots were visualized by 2D-DIGE. Multiple protein spots decreased their intensity by the treatment with TAPI-2. [Conclusion] We found that multiple protein spots were affected by TAPI-2. The affected protein would be released from the cell surface by ADAM17. This method is thought useful to comprehensively analyze cell surface proteins affected by the ectodomain shedding.