Department of Biological Sciences, KAIST, Daejeon 305-701, Korea.
Plant Cell. 2011 Apr;23(4):1404-15. doi: 10.1105/tpc.110.080721. Epub 2011 Apr 5.
A previous study showed that SOMNUS (SOM), which encodes a C3H-type zinc finger protein, is a key negative regulator of seed germination that acts downstream of PHYTOCHROME INTERACTING FACTOR3-LIKE5 (PIL5). However, it was not determined if PIL5 is the sole regulator of SOM expression. Public microarray data suggest that the expression of SOM mRNA is regulated also by ABSCISIC ACID INSENSITIVE3 (ABI3), another key regulator of seed germination. By analyzing abi3 mutants and ABI3 overexpression lines, we show here that ABI3 activates the expression of SOM mRNA collaboratively with PIL5 in imbibed seeds. Chromatin immunoprecipitation analysis coupled with electrophoretic mobility shift assay indicate that ABI3 activates the expression of SOM mRNA by directly binding to two RY motifs present in the SOM promoter in vivo, which is further supported by the greatly decreased expression of a reporter gene driven by a SOM promoter bearing mutated RY motifs. At the protein level, the ABI3 protein interacts with the PIL5 protein. The ABI3-PIL5 interaction, however, does not affect targeting of ABI3 and PIL5 to SOM promoters. Taken together, our results indicate that ABI3 and PIL5 collaboratively activate the expression of SOM mRNA by directly binding to and interacting with each other at the SOM promoter.
先前的研究表明，编码 C3H 型锌指蛋白的 SOM 是种子萌发的关键负调控因子，其作用于 PHYTOCHROME INTERACTING FACTOR3-LIKE5（PIL5）下游。然而，尚未确定 PIL5 是否是 SOM 表达的唯一调节剂。公共微阵列数据表明，SOM mRNA 的表达也受到另一个种子萌发关键调节剂——ABA 不敏感 3（ABI3）的调控。通过分析 abi3 突变体和 ABI3 过表达系，我们在这里表明，ABI3 在吸胀种子中与 PIL5 共同激活 SOM mRNA 的表达。染色质免疫沉淀分析结合电泳迁移率变动分析表明，ABI3 通过直接结合 SOM 启动子中存在的两个 RY 基序来激活 SOM mRNA 的表达，这进一步得到了由携带突变 RY 基序的 SOM 启动子驱动的报告基因表达大大降低的支持。在蛋白质水平上，ABI3 蛋白与 PIL5 蛋白相互作用。然而，ABI3-PIL5 相互作用并不影响 ABI3 和 PIL5 靶向 SOM 启动子。综上所述，我们的结果表明，ABI3 和 PIL5 通过直接结合和相互作用于 SOM 启动子，共同激活 SOM mRNA 的表达。
