Nishimura Noriyuki, Tsuchiya Wataru, Suzuki Nahomi, Hirayama Takashi, Yamazaki Toshimasa
Division of Plant Molecular Regulation Research, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, 2-1-2 Kannondai, Tsukuba, 305-8602, Ibaraki, Japan.
Radiation Breeding Division, Institute of Crop Science, National Agriculture and Food Research Organization, 2425 Kamimurata, Hitachiohmiya, 319-2293, Ibaraki, Japan.
Plant J. 2025 Apr;122(2):e70180. doi: 10.1111/tpj.70180.
Abscisic acid (ABA) is an important phytohormone regulating seed dormancy and germination. DELAY OF GERMINATION 1 (DOG1) is a pivotal regulator of seed dormancy and regulates the ABA response by binding with ABA HYPERSENSITIVE GERMINATION 1 (AHG1) and heme. However, to date, the molecular function and regulatory mechanisms of DOG1 remain unclear, including the relationship between DOG1 and the ABA response. Here, we investigate the mechanism of DOG1 in the ABA response using RNA sequencing, genetics, and biochemistry experiments. Our data suggest that DOG1 and AHG1 regulate the expression of many common genes, including seed maturation and ABA response. Moreover, DOG1 acts upstream of ABA INSENSITIVE 5 (ABI5) and regulates ABI5 target genes including LATE EMBRYOGENESIS ABUNDANT 1 (EM1) and EM6. We therefore performed a genetic screen to isolate mutants that suppress the ABA hypersensitive phenotype of YFP-DOG1-overexpressing transgenic plants. Ten mutant alleles caused mutations in the DOG1 transgene region, including three premature stop codon mutations and seven single amino acid substitutions. One of these mutants, P178L, which contains an amino acid substitution, abolished the interaction with AHG1 and promoted the dimerization of DOG1. Furthermore, we identify a heme-binding residue, Cys96, that plays an important role in ABA response. Overall, these data suggest that DOG1 and AHG1 regulate ABA response via ABI5 and that the association between heme and AHG1 is critical for the function of DOG1 during the regulation of seed germination.
脱落酸（ABA）是一种调控种子休眠和萌发的重要植物激素。萌发延迟1（DOG1）是种子休眠的关键调节因子，通过与ABA超敏萌发1（AHG1）和血红素结合来调节ABA反应。然而，迄今为止，DOG1的分子功能和调控机制仍不清楚，包括DOG1与ABA反应之间的关系。在此，我们利用RNA测序、遗传学和生物化学实验研究DOG1在ABA反应中的机制。我们的数据表明，DOG1和AHG1调节许多共同基因的表达，包括种子成熟和ABA反应。此外，DOG1在ABA不敏感5（ABI5）的上游起作用，并调节ABI5的靶基因，包括晚期胚胎发生丰富蛋白1（EM1）和EM6。因此，我们进行了遗传筛选，以分离出能抑制YFP - DOG1过表达转基因植物ABA超敏表型的突变体。十个突变等位基因导致DOG1转基因区域发生突变，包括三个提前终止密码子突变和七个单氨基酸替换。其中一个突变体P178L含有一个氨基酸替换，消除了与AHG1的相互作用并促进了DOG1的二聚化。此外，我们鉴定出一个血红素结合残基Cys96，它在ABA反应中起重要作用。总体而言，这些数据表明DOG1和AHG1通过ABI5调节ABA反应，并且血红素与AHG1之间的关联对于DOG1在种子萌发调控过程中的功能至关重要。
