转录因子TDF1对拟南芥分泌型绒毡层发育与功能的转录调控

During plant male gametophytes process, tapetum plays critical roles in pollen and pollen wall development. TDF1 encodes an R2R3 MYB transcription factor in Arabidopsis. The mutation of TDF1 shows the swollen and vacuolated tapetal cells with excessive division, and the failure differentiation of secretory tapetum finally leads to microspores degradation. However, the molecular mechanism of transcriptome regulated by TDF1 and its target genes and the biological functions of target genes activated by TDF1 for secretory tapetum development remain unknown. We have found that TDF1 can directly bind the promoters of CDI8, CEXP5 and AMS, respectively. In this project, 1) These candidate target genes of TDF1 will be further confirmed by bio-chemistry approaches. 2) Other proteins interacting with TDF1 will be screened by yeast two-hybrid assay and confirmed by tobacco Co-IP assay. 3) Expression patterns and subcellular locations of CDI8, CEXP5 and AMS will be observed; functions of CDI8 and CEXP5 for anther development will be analyzed by genetic and cytological approaches. 4) The recovery of tapetum and pollen wall development in rescue lines with tdf1 background will be observed. Based on these results, we will reveal the molecular mechanism regulated by TDF1 and its target genes for anther development. Furthermore, biological functions of CDI8, CEXP5 and AMS will be illustrated for secretory tapetum differentiation and pollen wall formation.

植物雄性生殖发育中，绒毡层对花粉发育起到重要作用。TDF1编码拟南芥MYB转录因子，该基因突变导致绒毡层细胞分裂增多、细胞肿胀且空泡化严重；最终分泌型绒毡层分化受阻、小孢子完全降解。然而，TDF1与其下游基因组成的转录调控在花药发育中的分子机制；以及TDF1下游基因对于分泌型绒毡层发育的生物学功能仍未知。申请人已初步确定TDF1能够结合在CDI8、CEXP5和AMS基因的启动子上。本项目中，1）明确CDI8、CEXP5和AMS基因受TDF1直接激活调控。2）筛选与TDF1相互作用的蛋白。3）对CDI8、CEXP5和AMS基因进行表达及功能分析。4）Rescue分析CDI8、CEXP5和AMS基因对于tdf1中绒毡层及花粉壁发育的补偿表型。由此，揭示TDF1与其靶基因组成的转录调控网络在花药发育中的分子机制与作用模式；探究TDF1下游基因对于分泌型绒毡层细胞分化及花粉壁形成的具体生物学功能。

植物雄性生殖发育中，绒毡层对花粉发育起到重要作用。TDF1编码拟南芥MYB转录因子，该基因突变导致绒毡层细胞分裂增多、细胞肿胀且空泡化严重；最终分泌型绒毡层分化受阻、小孢子完全降解。然而，TDF1直接调控哪些下游基因，在花药发育中其转录调控的具体分子机制；以及TDF1下游基因对于分泌型绒毡层发育的生物学功能均未知。本项目的研究结果显示TDF1直接激活调控AMS、和CDI8基因的表达，同时TDF1与AMS相互作用，在体内形成复合体并以feed-forward loop的形式共同激活CEXP5、MS188、TEK基因的表达；此外，CDI8和CEXP5基因在绒毡层转入分泌型的第6~7期中有高量表达，提示其参与分泌型绒毡层发育的可能性。这些研究结果揭示了TDF1与其靶基因组成的转录调控网络在花药发育中的具体分子机制与作用模式；并有助于进一步探究TDF1下游基因对于分泌型绒毡层发育的生物学功能。