Xiong Shuang-Xi, Lu Jie-Yang, Lou Yue, Teng Xiao-Dong, Gu Jing-Nan, Zhang Cheng, Shi Qiang-Sheng, Yang Zhong-Nan, Zhu Jun
College of Life and Environment Sciences, Shanghai Normal University, 100 Guilin Road, Shanghai, 200234, China.
Plant J. 2016 Dec;88(6):936-946. doi: 10.1111/tpj.13284. Epub 2016 Sep 17.
The sexine layer of pollen grain is mainly composed of sporopollenins. The sporophytic secretory tapetum is required for the biosynthesis of sporopollenin. Although several enzymes involved in sporopollenin biosynthesis have been reported, the regulatory mechanism of these enzymes in tapetal layer remains elusive. ABORTED MICROSPORES (AMS) and MALE STERILE 188/MYB103/MYB80 (MS188/MYB103/MYB80) are two tapetal cell-specific transcription factors required for pollen wall formation. AMS functions upstream of MS188. Here we report that AMS and MS188 target the CYP703A2 gene, which is involved in sporopollenin biosynthesis. We found that AMS and MS188 were localized in tapetum while CYP703A2 was localized in both tapetum and locule. Chromatin immunoprecipitation (ChIP) showed that MS188 directly bound to the promoter of CYP703A2 and luciferase-inducible assay showed that MS188 activated the expression of CYP703A2. Yeast two-hybrid and electrophoretic mobility shift assays (EMSAs) further demonstrated that MS188 complexed with AMS. The expression of CYP703A2 could be partially restored by the elevated levels of MS188 in the ams mutant. Therefore, our data reveal that MS188 coordinates with AMS to activate CYP703A2 in sporopollenin biosynthesis of plant tapetum.
花粉粒的外壁层主要由孢粉素组成。孢子体分泌型绒毡层是孢粉素生物合成所必需的。尽管已经报道了几种参与孢粉素生物合成的酶，但这些酶在绒毡层中的调控机制仍然不清楚。败育小孢子（AMS）和雄性不育188/MYB103/MYB80（MS188/MYB103/MYB80）是花粉壁形成所需的两个绒毡层细胞特异性转录因子。AMS在MS188的上游起作用。在这里我们报道AMS和MS188靶向参与孢粉素生物合成的CYP703A2基因。我们发现AMS和MS188定位于绒毡层，而CYP703A2定位于绒毡层和小室。染色质免疫沉淀（ChIP）表明MS188直接结合到CYP703A2的启动子上，荧光素酶诱导分析表明MS188激活CYP703A2的表达。酵母双杂交和电泳迁移率变动分析（EMSA）进一步证明MS188与AMS形成复合物。在ams突变体中，MS188水平的升高可以部分恢复CYP703A2的表达。因此，我们的数据揭示了在植物绒毡层的孢粉素生物合成中，MS188与AMS协同激活CYP703A2。
