Signal peptide of eosinophil cationic protein is toxic to cells lacking signal peptide peptidase.
Eosinophil cationic protein (ECP) is a toxin secreted by activated human eosinophils. The properties of mature ECP have been well studied but those of the signal peptide of ECP (ECPsp) are not clear. In this study, several chimeric proteins containing N-terminal fusion of ECPsp were generated, and introduced into Escherichia coli, Pichia pastoris, and human epidermoid carcinoma cell line A431 to study the function of ECPsp. We found that expression of ECPsp chimeric proteins inhibited the growth of E. coli and P. pastoris but not A431 cells. Primary sequence analysis and in vitro transcription/translation of ECPsp have revealed that it is a potential substrate for human signal peptide peptidase (hSPP), an intramembrane protease located in endoplasmic reticulum. In addition, knockdown of the hSPP mRNA expression in ECPsp-eGFP/A431 cells caused the growth inhibitory effect, whereas complementally expression of hSPP in P. pastoris system rescued the cell growth. Taken together, we have demonstrated that ECPsp is a toxic signal peptide, and expression of hSPP protects the cells from growth inhibition.
Wu
Chia-Mao
CM
Department of Life Science, Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan 30013, Republic of China.
Chang
Margaret Dah-Tsyr
MD
D016282:Aspartic Acid Endopeptidases / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D001798:Blood Proteins / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D047091:Eosinophil Granule Proteins; D004926:Escherichia coli; D017930:Genes, Reporter; D006801:Humans; D010843:Pichia; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012260:Ribonucleases / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity
D001798:Blood Proteins; D047091:Eosinophil Granule Proteins; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D012260:Ribonucleases; D016282:Aspartic Acid Endopeptidases; C460085:signal peptide peptidase