Membrane-bound carboxypeptidase E facilitates the entry of eosinophil cationic protein into neuroendocrine cells

The Biochemical Journal
Chia-Mao WuMargaret Dah-Tsyr Chang

Abstract

ECP (eosinophil cationic protein) is a major component of eosinophil granule proteins, and is used as a clinical biomarker for asthma and allergic inflammatory disease. ECP has been implicated in damage to the cell membrane of many tissue types, but the mechanism is not well known. In the present study, mECP-eGFP-6H, a recombinant fusion protein containing mature ECP (mECP), enhanced green fluorescence protein (eGFP) and a His(6) tag (6H), has been expressed, purified and added to GH3 neuroendocrine cells to study the internalization ability of ECP. We found that mECP-eGFP-6H entered into GH3 neuroendocrine cells and inhibited the growth of the cells with an IC(50) of 0.8 microM. By yeast two-hybrid screening and immunoprecipitation, we have identified a specific protein-protein interaction between mECP and CPE (carboxypeptidase E), a well characterized metalloprotease. Further in vivo yeast two-hybrid screening has also revealed that residues 318-387 located in a region of unknown function in mature CPE are indispensable for association with mECP. In addition, the uptake of mECP-eGFP-6H is suppressed by dominant-negative expression of the recycling defect mutant pre-pro-HA-CPE(S471A,E472A) in GH3 cells, suggesting that the ent...Continue Reading

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Citations

Mar 10, 2012·Endocrine Reviews·Niamh X CawleyY Peng Loh
Aug 2, 2008·Molecular Endocrinology·Joshua J Park, Y Peng Loh
Aug 25, 2004·Biochemical and Biophysical Research Communications·Chia-Mao Wu, Margaret Dah-Tsyr Chang
Jan 27, 2009·Journal of Neurochemistry·Shigeru OisoKatsushi Yamada
Mar 23, 2013·Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine·Hui-Chen ChenChia-Hung Hsieh

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