title: Cloning, expression and characterization of Ostrinia furnacalis serpin1, a regulator of the prophenoloxidase activation system.
abstract: Serine protease inhibitors of the serpin superfamily are regulators of proteases involved in a variety of physiological processes including immune responses. In this study, we have isolated a full-length serpin cDNA from Ostrinia furnacalis. The 1188 bp open reading frame encodes a 395-residue protein with a theoretical molecular mass of 43.3 kDa and an isoelectric point of 4.92. Ofserpin1 contains a putative signal peptide followed by a conserved domain including a reactive center loop (RCL) with a hinge region (E(344) to S(353)) and a predicted P1-P1' cleavage site (Leu(360)-Ser(361)). Ofserpin1 mRNA and protein were detected in all the tested tissues, particularly in hemocytes and integument. The recombinant protein inhibited chymotrypsin and trypsin in a dose-dependent manner, and were significantly cleaved by the enzyme trypsin and chymotrypsin. Ofserpin1 impeded the prophenoloxidase activation cascade by 45.6% at 16.5 μg, and affected activity of prophenoloxidase activating protease. Levels of Ofserpin1 transcripts in the integument were higher than those in hemocytes, fat body and midgut. After an immune challenge with Staphylococcus aureus and Escherichia coli, the relative mRNA levels of Ofserpin1 decreased in 2-10h post-infection (hpi) in integument and hemocytes compared to the untreated control. Our results suggested that Ofserpin1 has serine protease inhibitory activity and is likely involved in the regulation of prophenoloxidase activation system in O. furnacalis.
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Bing
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Taoyan
authors.initials: T
authors.lastname: Wu
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Tang
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Shiyang
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Qiuwen
authors.initials: Q
authors.lastname: Xu
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Ya
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China.
authors.forename: Yingjuan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Plant Protection, College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, Jiangsu, PR China. Electronic address: fengcj@yzu.edu.cn.
authors.forename: Congjing
authors.initials: C
authors.lastname: Feng
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004156:Catechol Oxidase / Q000235:genetics* / Q000378:metabolism; D002918:Chymotrypsin / Q000737:chemistry; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004792:Enzyme Precursors / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000254:growth & development; D015870:Gene Expression; D006434:Hemocytes / Q000201:enzymology* / Q000382:microbiology; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D034582:Integumentary System / Q000382:microbiology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009036:Moths / Q000201:enzymology* / Q000235:genetics / Q000382:microbiology; D016366:Open Reading Frames; D009928:Organ Specificity; D021382:Protein Sorting Signals; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D015843:Serpins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013211:Staphylococcus aureus / Q000254:growth & development; D014357:Trypsin / Q000737:chemistry
chemical_list: D018076:DNA, Complementary; D004792:Enzyme Precursors; D019476:Insect Proteins; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011994:Recombinant Proteins; D015843:Serpins; C038451:pro-phenoloxidase; D004156:Catechol Oxidase; D002918:Chymotrypsin; D014357:Trypsin
keywords: Bacterial challenge; Innate immunity; Ostrinia furnacalis; Prophenoloxidase activation; Serpin1