荒漠昆虫小胸鳖甲水通道蛋白基因MpAQP1克隆及低温对其表达的影响

摘要

水通道蛋白(aquaporin, AQP)参与细胞的水分平衡与运输,与生物的抗逆性相关,但AQP在昆虫耐寒性中的作用仅有少量报道.为了检测昆虫AQP基因的序列特征及其表达是否与低温相关,本研究从荒漠甲虫小胸鳖甲Microdera punctipennis 中克隆获得 AQP 的开放阅读框(ORF)序列,命名为 MpAQP1(GenBank 登录号:KX129951).生物信息学分析表明,MpAQP1的ORF长度为813 bp,编码270个氨基酸,分子量为28.59 kDa,理论等电点为7.73.MpAQP1属于萤火虫内嵌蛋白(PRIP)家族,具有6个跨膜结构域和5个螺旋环,含有高度保守的天冬酰胺-脯氨酸-丙氨酸基序(NPA基序).实时荧光定量PCR检测MpAQP1在低温下的表达谱发现,4℃低温处理1 h, MpAQP1开始上调表达,5 h达到最高峰,为对照组的34.39倍.研究表明MpAQP1为冷响应基因,水通道蛋白可能参与小胸鳖甲低温下的渗透调节而发挥耐寒性功能.%Aquaporin(AQP)involves in water balance and transport in cells, thus it is related to organisms'stress resistance.However,studies on the function of AQP in insect cold resistance are rare.To detect whether the expression of AQP gene is induced by low temperature, we firstly cloned the Open Reading Frame(ORF)sequence of the aquaporin gene from the desert beetle Microdera punctipennis,and named it as MpAQP1(GenBank accession number: KX129951).Bioinformatics analysis showed that MpAQP1 ORF is 813 bp encoding 270 amino acids with a molecular weight of 28.59 kDa, the theoretical isoelectric point is 7.73.MpAQP1 is a member of Pyrocoelia rufa integral protein(PRIP)family.MpAQP1 exhibited a typical structure with six transmembrane domains, five helix loops and two highly conserved asparagine-proline-alanine motifs(NPA motifs).Real time quantitative PCR results showed that the expression of MpAQP1 was up-regulated after the insect was treated at 4℃ for 1 h, and reached peak at 5 h,which was 34.39 times of the control group.Our study indicated that MpAQP1 was a cold responsive gene,which may play role in improving cold tolerance of M.punctipennis by involving in osmoregulation.