The cDNA cloning and mRNA expression of heat shock protein 70 gene in the haemocytes of bay scallop (Argopecten irradians, Lamarck 1819) responding to bacteria challenge and naphthalin stress.
Heat shock protein 70 (HSP70) is an important member of the heat shock protein superfamily, and it plays a key role in the process of protecting cells, facilitating the folding of nascent peptides and responding to stress. The cDNA of bay scallop Argopecten irradians HSP70 (designated AIHSP70) was cloned by the techniques of homological cloning and rapid amplification of cDNA end (RACE). The full length of AIHSP70 cDNA was 2651bp in length, having a 5' untranslated region (UTR) of 96bp, a 3' UTR of 575bp, and an open reading frame (ORF) of 1980bp encoding a polypeptide of 659 amino acids with an estimated molecular mass of 71.80kDa and an estimated isoelectric point of 5.26. BLAST analysis revealed that the AIHSP70 gene shared high identity with other known HSP70 genes. Three classical HSP signature motifs were detected in AIHSP70 by InterPro analysis. 3-D structural prediction of AIHSP70 showed that its N terminal ATPase activity domain and C terminal substrate-binding domain shared high similarity with that in human heat shock protein 70. The results indicated that the AIHSP70 was a member of the heat shock protein 70 family. A semi-quantitive RT-PCR method was used to analyse the expression of AIHSP70 gene after the treatment of naphthalin which is one kind of polycyclic aromatic hydrocarbon (PAH) and the challenge of bacteria. mRNA expression of AIHSP70 in scallop was up-regulated significantly after the stimulation of naphthalin and increased with increasing naphthalin concentration. A clearly time-dependent expression pattern of AIHSP70 was observed after the scallops were infected by Vibrio anguillarum, and the mRNA expression reached a maximum level at 8h and lasted to 16h, and then dropped progressively. The results indicated that AIHSP70 could play an important role in mediating the environmental stress and immune response in scallop.
Song
Linsheng
L
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China. lshsong@ms.qdio.ac.cn
Wu
Longtao
L
Ni
Duojiao
D
Chang
Yaqing
Y
Xu
Wei
W
Xing
Kezhi
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary / Q000737:chemistry; D015870:Gene Expression / Q000502:physiology*; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D006434:Hemocytes / Q000187:drug effects / Q000502:physiology; D008969:Molecular Sequence Data; D049897:Pectinidae / Q000235:genetics* / Q000382:microbiology / Q000502:physiology; D010802:Phylogeny; D011084:Polycyclic Aromatic Hydrocarbons / Q000633:toxicity*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014733:Vibrio / Q000502:physiology*
D018076:DNA, Complementary; D018840:HSP70 Heat-Shock Proteins; D011084:Polycyclic Aromatic Hydrocarbons; D012333:RNA, Messenger