拟南芥ATLP-3基因的表达模式探究

Expression Pattern Analysis of ATLP-3 in Arabidopsis thaliana

拟南芥(Arabidopsis thaliana) ATLP-3 (AT1G75030)基因编码蛋白是类甜蛋白家族(Thaumatin-Like Protein Superfamily)成员之一。目前对于类甜蛋白功能的研究主要集中于抗病和环境胁迫响应两方面,而对于蛋白的基础功能解析和调控机理研究较少。本实验通过分析转录组数据发现ATLP-3基因在花器官发育过程中高量表达,通过构建ATLP-3启动子连接GUS的转基因植株,染色并进行石蜡切片分析发现该基因在花药发育的第5期到第10期绒毡层中特异表达,说明该蛋白可能参与调控绒毡层的功能。进一步分析发现花药发育过程中发挥调控功能的转录因子TDF1、AMS和MS188在ATLP-3启动子区均有结合位点,高通量测序结果显示ATLP-3表达量在以上转录因子的突变体中相比于野生型显著下调。进一步实验发现,将ProATLP-3:GUS引入AMS功能缺失的突变体背景中,GUS信号相比于野生型明显下降,说明ATLP-3的表达受到AMS的正向调控。本实验还获得了ATLP-3基因功能缺失突变体并分析其花粉表型。以上研究结果将为深入探究ATLP-3基因的功能提供依据,为更多类甜蛋白家族成员功能的解析提供参考。

The protein encoded by ATLP-3(AT1G75030)gene of Arabidopsis thaliana belongs to the Thau-matin-Like Protein Superfamily.Until now,most studies about ATLP proteins function are focused on disease re-sistance and environmental stress response,while the basic function analysis and regulatory mechanism of the pro-teins are rarely studied.In this study,we found that ATLP-3 was highly expressed during flower developmental stages by analyzing transcriptome data.Transgenic plants with ATLP-3 promoter connected to GUS were con-structed,stained and analyzed in paraffin sections.It was found that ATLP-3 gene was specifically expressed in the tapetum layer of stage 5~10 anthers,this suggests that ATLP may be involved in the regulation of tapetum func-tion.Further analysis showed that the ATLP-3 promoter region contains binding sites of the transcription factors TDF1,AMS and MS188,which play a key role during anthers development.And RNA-seq results showed that the expression of ATLP-3 was significantly down-regulated in the mutants of these transcription factors compared with the wild-type.Theremore Pro ATLP-3:GUS was introduced into ams background,and GUS staining showed that the signal of GUS was significantly reduced compared with that in the wild tpye,indicating that the expression of ATLP-3 was positively regulated by AMS.The null mutant of ATLP-3 was obtained and its pollen phenotype was analyzed.This study will lay a foundation for further exploring the function of ATLP-3,and also provide a refer-ence for the analysis of the functions of ATLP proteins.