State Key Laboratory of Genetic Engineering, Ministry of Education Key Laboratory of Biodiversity Sciences and Ecological Engineering and Institute of Biodiversity Sciences, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai, 200438, China.
Department of Biology, The Huck Institutes of the Life Sciences, The Pennsylvania State University, University Park, PA, 16802, USA.
J Genet Genomics. 2020 Aug;47(8):477-492. doi: 10.1016/j.jgg.2020.09.001. Epub 2020 Sep 8.
The Arabidopsis bHLH010/089/091 (basic helix-loop-helix) genes are functionally redundant and are required for both anther development and normal expression of DYT1-activated anther-related genes. These three genes are conserved in Brassicaceae, suggesting that each of them is under selection pressure; however, little is known about the possible functional differences among these bHLH genes and between the bHLH and DYT1 genes. Here, we compared novel anther transcriptomic data sets from bHLH010/089/091 single and double mutants, with an anther transcriptomic data set from the wild type (WT) and a previously obtained anther transcriptomic data set from the bhlh010 bhlh089 bhlh091 triple mutant. The results revealed molecular phenotypes that support the functional redundancy and divergence of bHLH010, bHLH089, and bHLH091, as well as the functional overlap and difference between them and DYT1. DNA-binding analyses revealed that DYT1 and bHLH089 specifically recognize the TCATGTGC box to activate the expression of target genes, including ATA20, EXL4, and MEE48. In addition, among genes whose expression was affected in the bhlh010 bhlh089 double and bhlh010 bhlh089 bhlh091 triple mutants, genes that are involved in the stress response and cell signaling were enriched, which included 256 genes whose expression was preferentially induced by heat during early flower development. Moreover, the bhlh double mutants exhibited defective pollen development when the plants were grown under elevated temperature, suggesting that bHLH genes are important for anther gene expression under such conditions. These results are consistent with the observation that the heat-induced expression of several genes is less in the bhlh mutants than that in the WT. Therefore, our results provide important insights into the molecular mechanism underlying the activation of direct targets by DYT1-bHLH089 heterodimers and demonstrate the protective roles of bHLH010/089/091 in maintaining fertility upon heat stress.
拟南芥 bHLH010/089/091（碱性螺旋-环-螺旋）基因在功能上是冗余的，对于花药发育和 DYT1 激活的花药相关基因的正常表达都是必需的。这三个基因在十字花科中是保守的，这表明它们各自都受到选择压力的影响；然而，关于这些 bHLH 基因之间以及 bHLH 和 DYT1 基因之间可能存在的功能差异，我们知之甚少。在这里，我们比较了 bHLH010/089/091 单突变体和双突变体的新型花药转录组数据集，与野生型（WT）的花药转录组数据集以及之前获得的 bhlh010 bhlh089 bhlh091 三突变体的花药转录组数据集。结果揭示了支持 bHLH010、bHLH089 和 bHLH091 功能冗余和分化的分子表型，以及它们与 DYT1 之间的功能重叠和差异。DNA 结合分析表明，DYT1 和 bHLH089 特异性识别 TCATGTGC 框以激活靶基因的表达，包括 ATA20、EXL4 和 MEE48。此外，在 bhlh010 bhlh089 双突变体和 bhlh010 bhlh089 bhlh091 三突变体中，那些表达受到影响的基因，包括在早期花发育过程中受到热优先诱导的 256 个基因，参与了应激反应和细胞信号转导。此外，当在高温下生长时，bHLH 双突变体表现出花粉发育缺陷，这表明 bHLH 基因在这种条件下对于花药基因表达很重要。这些结果与观察到的 bhlh 突变体中几个基因的热诱导表达低于 WT 的结果一致。因此，我们的结果为 DYT1-bHLH089 异二聚体激活直接靶基因的分子机制提供了重要的见解，并证明了 bHLH010/089/091 在高温胁迫下维持育性的保护作用。
