摘要
半胱氨酸蛋白酶3(Caspase-3)作为细胞凋亡通路中重要的效应蛋白,在细胞凋亡过程中发挥着重要作用。为初步探究马氏珠母贝Caspase-3(PmCaspase-3)的生物学功能,本研究利用cDNA末端快速扩增(RACE)技术克隆获得PmCaspase-3基因cDNA的全长序列并对其序列特征进行分析;同时利用实时荧光定量PCR(RT-qPCR)方法分析了PmCaspase-3基因mRNA在马氏珠母贝不同组织和不同发育时期的表达差异。结果显示,PmCaspase-3基因cDNA全长为2233 bp,其中5'端非编码区长度为80 bp,3'端非编码长度为31 bp,开放阅读框长度为2088 bp,共编码695个氨基酸;生物信息学分析显示,PmCaspase-3含有Caspase家族特有的CASc结构域和半胱氨酸蛋白酶家族p20、p10活性位点以及多种磷酸化位点,经进化分析以及多序列比对可知与其他物种Caspase-3蛋白同源性较高。RT-qPCR结果表明,PmCaspase-3在肝胰腺中的表达量最高,在闭壳肌中的表达量最低;在发育过程中D型幼虫期和眼点期的表达量较高。
As an important effector protein in apoptosis pathway,Caspase-3 plays an important role in the process of apoptosis.In order to explore the biological function of Caspase-3 from Pinctada fucata martensii(PmCaspase-3),the full-length cDNA sequence of PmCaspase-3 was cloned by rapid amplification of cDNA ends(RACE)technology,and then its characteristics were analyzed.In the meantime,expression levels of PmCaspase-3 in different tissues and different development stages of P.f.martensii were tested by RT-qPCR.Results showed that the full length of PmCaspase-3 cDNA was 2233 bp,consisting of a 5'-terminal untranslated region(UTR)of 80 bp,a 3'-terminal UTR of 31 bp,and an open reading frame(ORF)of 2088 bp encoding a polypeptide of 695 amino acids.Bioinformatics analysis showed that PmCaspase-3 contained the CASc domain unique to the Caspase family and the active sites of p20 and p10 of the cysteine protease family,as well as a variety of phosphorylation sites.The construction of the phylogenetic tree of its amino acid sequences and the multi-sequence alignment showed that PmCaspase-3 of Pinctada fucata martensii had high homologywith PmCaspase-3 of other species.RT-qPCR results showed that the expression level of PmCaspase-3 was the highest in the hepatopancreas and the lowest in adductor muscle.During development,PmCaspase-3was highly expressed inD-shaped larvae and Eye-spotted larvae.
作者
房晓宸
卢金昭
梁海鹰
何军军
申铖皓
Fang Xiaochen;Lu Jinzhao;Liang Haiying;He Junjun;Shen Chenghao(Shenzhen Research Institute,Guangdong Ocean University,Shenzhen,518108;Fisheries College,Guangdong Ocean University,Zhanjiang,524088;Guangdong Provincial Engineering Research Center for Aquatic Animal Health Assessment,Shenzhen,518108;Guangdong Province,Pearl Breeding and Processing Engineering Technology Research Center,Zhanjiang,524025)
出处
《基因组学与应用生物学》
CAS
CSCD
北大核心
2021年第3期970-979,共10页
Genomics and Applied Biology
基金
国家自然科学基金(31472306)
广东省海港建设与渔业产业发展专项(A201608B15)
深圳市科技计划(JCYJ20180507183240459)共同资助。
