蓖麻RcSEC14p基因的克隆及低温胁迫下的表达分析

Cloning of RcSEC14p Gene in Ricinus communis L. and Expression Analysis under Cold Stress

Sec14-like蛋白参与细胞的基本生物学过程,如磷脂代谢、膜运输、信号转导及逆境响应。本研究采用RT-PCR的方法在蓖麻叶片中成功克隆了一个磷脂酰肌醇转运蛋白基因RcSEC14p (Accession No.MK205181)。RcSEC14p基因全长包含一个1 752 bp的完整开放阅读框,推测编码583个氨基酸。生物信息分析发现RcSEC14p为不稳定的亲水性蛋白;蛋白质二级结构元件多为α螺旋和无规卷曲。三级结构分析表明RcSEC14p结构内部含有一个潜在的磷脂结合口袋,可能在脂质代谢和磷酸肌醇信号转导等方面发挥重要的作用。多重序列比对表明蓖麻RcSEC14p与麻风树同源蛋白的相似性高达77.03%。系统进化树分析表明RcSEC14p属于Sec-like蛋白家族的SFH亚家族成员,N端包含一个SEC结构域,C端包含一个Nodulin结构域。q RT-PCR分析发现RcSEC14p受冷胁迫下调表达。该基因的下调表达可能会降低蓖麻的抗冷性。该研究为进一步探讨RcSEC14p是否通过调节磷脂信号转导途径参与冷胁迫响应提供依据。

Sec14-like protein participated in the basic biological process of cells, such as phospholipid metabolism,membrane traffic, signal transduction and stress response. In this study, a phosphatidylinositol transfer protein gene RcSEC14 p(Accession No.MK205181) was successfully cloned from castor leaves by RT-PCR method. Full length of RcSEC14 p gene consists of a complete open reading frame(1 752 bp) encoded a putative polypeptide of 583 amino acids. Bioinformatic analysis showed that RcSEC14 p was an unstable hydrophilic protein. Most of the protein secondary structural elements were α-helices and random coil. Tertiary structure analysis revealed that there was a potential phospholipid binding pocket in the structure of RcSEC14 p that may play a critical role in lipid metabolism and phosphoinositide signaling transduction. Multiple sequence alignment displayed that the similarity between castor RcSEC14 p and Jatropha carcas L. homologous protein was as high as 77.03%. Phylogenetic tree analysis showed that RcSEC14 p belongs to the SFH subfamily of Sec-like protein family, whose N-terminal contained a SEC domain and C-terminal contained a Nodulin domain. qRT-PCR analysis showed that the expression of RcSEC14 p was down-regulated by cold stress. Down-regulation of the gene expression might reduce the cold tolerance of castor. This study laid the foundation to test whether RcSEC14 p is involved in cold stress response by modulating phospholipid signaling transduction which deserves further investigation.