cDNA cloning, characterization and mRNA expression of a pacifastin light chain gene from the Chinese mitten crab Eriocheir sinensis.
The pacifastin family, characterized by several conserved arrays of six cysteine residues, is a newly identified serine protease inhibitor (SPI) family discovered uniquely in arthropods and plays important roles in multiple biological processes. In the present study, the full-length cDNA of a pacifastin light chain (designated ESPLC) was cloned from the Chinese mitten crab Eriocheir sinensis by expressed sequence tags (ESTs) and PCR techniques. The 1036bp ESPLC cDNA contained an 831bp open reading frame (ORF) encoding a putative pacifastin-related peptide of 276 amino acids, a 5'-untranslated region (UTR) of 67bp, and a 3'-UTR of 138bp. Six putative conserved domains sharing a characteristic cysteine array (Cys-Xaa(9-12)-Cys-Asn-Xaa-Cys-Xaa-Cys-Xaa(2-3)-Gly-Xaa(3-4)-Cys-Thr-Xaa(3)-Cys) were identified in the deduced amino acid sequence of ESPLC. The conservation of these PLDs (pacifastin light chain domains) and the relative higher similarity of ESPLC to other pacifastin-related precursors suggested that ESPLC was a member of pacifastin family. The mRNA transcripts of ESPLC were found to be higher expressed in hepatopancreas, gill and haemolymph than in gonad, muscle and heart, with the highest expression level in hepatopancreas. The ESPLC mRNA expression in haemolymph of Chinese mitten crab was up-regulated at 2h and 12h after challenged with Listonella anguillarum. The tissue distribution and temporal characteristics of ESPLC mRNA expression, similar to that of prophenoloxidase gene in E. sinensis, suggested that ESPLC was potentially involved in the response against invading bacteria, with the possibility that it functioned in the prophenoloxidase system in E. sinensis.
Gai
Yunchao
Y
Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China.
Wang
Lingling
L
Song
Linsheng
L
Zhao
Jianmin
J
Qiu
Limei
L
Wang
Bo
B
Mu
Changkao
C
Zheng
Peilin
P
Zhang
Ying
Y
Li
Ling
L
Xing
Kezhi
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003386:Brachyura / Q000276:immunology / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D005786:Gene Expression Regulation / Q000502:physiology*; D016905:Gram-Negative Bacterial Infections / Q000276:immunology / Q000378:metabolism; D053526:Listonella; D008969:Molecular Sequence Data; D010802:Phylogeny; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
D018076:DNA, Complementary; D011506:Proteins; D012333:RNA, Messenger; C106709:pacifastin