紫苏WRI1转录因子的鉴定、表达分析及低温胁迫响应

Identification,Expression Analysis and Low Temperature Stress Response of WRI1 Transcription Factor from Perilla frutescens

紫苏[Perilla frutescens(L.)Britt.]是一种新型油料经济作物。转录因子WRI1(WRINKLED1)参与调控植物油脂合成积累。为探究紫苏PfWRI1在油脂合成积累及胁迫响应过程中的分子调控机制,通过分子克隆技术分离得到PfWRI1的完整编码区,利用生物信息学分析工具和半定量PCR(sqRT-PCR)及实时荧光定量PCR(qRTPCR)技术对序列特征及表达水平等进行分析。结果显示:PfWRI1转录因子属于AP2(APETALA2)超家族,包含2个AP2保守结构域;亚细胞定位预测分析显示,PfWRI1转录因子定位于细胞核;无规则卷曲和α-螺旋是PfWRI1蛋白二级结构中的主要结构元件;系统进化分析表明,PfWRI1与芝麻和油梨WRI1的亲缘关系最近;互作蛋白预测表明,PfWRI1蛋白可能与LEC1(LEAFYCOTYLEDON1)、FATA(fatty acyl-acyl carrier protien thioesterase)、FATB和PDAT(phospholipids:diacylglycerol acyltransferase)等存在互作关系;PfWRI1在紫苏的不同组织中均有表达,随种子成熟其表达量呈现先升高后降低的模式,且在发育中后期(开花后30 d)种子的表达量最高,表明PfWRI1可能在紫苏种子油脂合成中具有调控作用;紫苏幼苗经过低温胁迫后,PfWRI1的表达量在胁迫12 h后显著上调,说明PfWRI1可能参与紫苏幼苗冷胁迫响应。这些结果为进一步研究PfWRI1在紫苏油脂合成积累和胁迫响应中的分子机制及功能奠定了基础,为紫苏和其他油料作物的遗传改良和新品种选育提供了理论依据。

Perilla[Perilla frutescens(L.)Britt.]is a new type of edible-medicinal oil crop.The transcription factor WRI1(WRINKLED1)is master regulator in transcriptional control of plant oil biosynthesis and accumulation.To explore the molecular mechanism of PfWRI1 transcription factor in regulating oil synthesis accumulation and stress response in perilla,PfWRI1transcription factor was cloned from perilla by molecular cloning technique.The sequence characteristics and expression levels were analyzed using bioinformatics analysis tools and semi-quantitative reverse transcription polymerase chain reaction(sqRTPCR)and quantitative real-time reverse transcription polymerase chain reaction(qRT-PCR)techniques.The results showed that PfWRI1 transcription factor is a member of the AP2(APETALA2)transcription factor family,which contains two AP2 conserved domains and is localized at the nucleus.Random coil andα-helix were the main structural elements in the secondary structure of PfWRI1 protein.Phylogenetic analysis showed that the closest relatives to PfWRI1 are Sesamum indicum and Persea americana WRI1.The prediction of interacting proteins indicated that PfWRI1 protein might interact with LEC1(LEAFYCOTYLEDON1),FATA(fatty acyl-acyl carrier protien thioesterase),FATB and PDAT(phospholipids:diacylglycerol acyltransferase).PfWRI1 expressed in all perilla organs/tissues tested,whereas their expression levels were different in these tissues.The expression level of PfWRI1 increased in the early developmental stage and then decreased,and which reached the peak l in seed at 30 days after flowering(DAF)with the seed maturity.It was suggested that PfWRI1 may play an important regulatory role in perilla seed oil synthesis.The expression of PfWRI1 transcription factor peaked after 12 hours under low temperature stress of perilla seedlings,indicating that PfWRI1 may be involved in the response to cold stress.This study lays a foundation for further research on the molecular mechanism and function of PfWRI1 in oil synthesis and accumulation and stress response of perilla,and provides a theoretical basis for genetic improvement and new variety breeding of perilla and other oil crops.