Hsp60 in caudal fin regeneration from Paramisgurnus dabryanus: Molecular cloning and expression characterization

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• 作者：Li Li ; Shengna Zhai ; Lele Wang ; Songbo Si ; Hailan Wu ; Zhongjie Chang
• 关键词：hsp60 gene ; Fin regeneration ; Wounding stimulation ; Gene expression ; Paramisgurnus dabryanus
• 刊名：Fish & Shellfish Immunology
• 出版年：February, 2014
• 年：2014
• 卷：36
• 期：2
• 页码：401-408
• 全文大小：4493 K

文摘

Heat shock protein 60 (Hsp60) is a kind of highly conserved immunogenic molecule involved in a wide range of biochemical processes in response to external stressors. Its multifunction in regulating immune responses and modulating signal pathway interests us in investigating its role in fin regeneration that has become an excellent and interesting model for studying the molecular basis of morphogenesis. We firstly clarified basical process and crucial period of caudal fins regeneration in Paramisgurnus dabryanus by histological analysis. Then we cloned full-length cDNA of hsp60 from P. dabryanus (designated as PdHsp60) by RACE method. The cDNA contains a 124聽bp 5鈥睻TR, a 1731聽bp open reading frame (ORF) encoding 576 amino acids and a 510bp 3鈥睻TR (Accession no.: KF544774). The phylogenetic tree shows that the PdHsp60 fits within the hsp60 clade. And quantitative RT-PCR detected the PdHsp60 began to increase rapidly its expression at 1聽dpa and reached its peak at 2聽dpa. Next, spatial distribution analysis of PdHsp60 in fins showed that PdHsp60 located mainly in the deeper lay of regenerated epidermis when PdHsp60 expressed most. After the PdHsp60 had been cloned into the pET-32a vector, SDS-PAGE and Western blotting analysis confirmed that the PdHsp60 protein was efficiently expressed in Escherichia coli BL21. These findings have revealed that PdHsp60, a highly conserved gene related to the innate immune system and stress response during vertebrate evolution, is involved in response to wounding stimulation鈥攊n the formation of wound epidermis which occurs as the first phase of fin regeneration after fin amputation in caudal fin regeneration.