Four Closely-related RING-type E3 Ligases, APD1-4, are Involved in Pollen Mitosis II Regulation in Arabidopsis
Four Closely-related RING-type E3 Ligases, APD1-4, are Involved in Pollen Mitosis II Regulation in Arabidopsis
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DOI：
10.1111/j.1744-7909.2012.01152.x
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发表时间：
2012-10-01
影响因子：
11.4
通讯作者：
Qu, Li-Jia
中科院分区：
文献类型：
--
作者：
Luo, Guo;Gu, Hongya;Qu, Li-Jia
蛋白质的泛素化是真核生物重要的调控机制之一。在高等植物中，蛋白质泛素化在许多生物学过程中起着至关重要的作用，包括激素信号传导、光形态建成和病原体防御。然而，蛋白质泛素化在生殖过程中的作用尚不清楚。在这项研究中，我们确定了四个植物特异性RING指基因命名为异常花粉发育1（APD 1）到APD 4，作为调节花粉有丝分裂II（PMII）在拟南芥（L.）。apd 1和apd 2双突变体在成熟花粉阶段表现出显著增加的双细胞样花粉的百分比。进一步下调的APD 3和APD 4转录在apd 1 apd 2的RNA干扰（RNAi）导致更严重的异常双细胞样花粉表型比apd 1 apd 2，表明细胞分裂是有缺陷的男性配子发生。这4个基因在雄配子体发育过程中均表现为多阶段、不同水平的表达。利用绿色荧光融合蛋白（GFP）GFP-APD 1和GFP-APD 2的共聚焦分析表明APD与细胞内膜相关联。此外，APD 2在体外具有E2依赖性E3连接酶活性，并鉴定了5个APD 2相互作用蛋白。我们的研究结果表明，这四个基因可能参与，冗余，在调节PMII过程中的男性配子发生。
Ubiquitination of proteins is one of the critical regulatory mechanisms in eukaryotes. In higher plants, protein ubiquitination plays an essential role in many biological processes, including hormone signaling, photomorphogenesis, and pathogen defense. However, the roles of protein ubiquitination in the reproductive process are not clear. In this study, we identified four plant-specific RING-finger genes designated Aberrant Pollen Development 1 (APD1) to APD4, as regulators of pollen mitosis II (PMII) in Arabidopsis thaliana (L.). The apd1 apd2 double mutant showed a significantly increased percentage of bicellular-like pollen at the mature pollen stage. Further downregulation of the APD3 and APD4 transcripts in apd1 apd2 by RNA interference (RNAi) resulted in more severe abnormal bicellular-like pollen phenotypes than in apd1 apd2, suggesting that cell division was defective in male gametogenesis. All of the four genes were expressed in multiple stages at different levels during male gametophyte development. Confocal analysis using green florescence fusion proteins (GFP) GFP-APD1 and GFP-APD2 showed that APDs are associated with intracellular membranes. Furthermore, APD2 had E2-dependent E3 ligase activity in vitro, and five APD2-interacting proteins were identified. Our results suggest that these four genes may be involved, redundantly, in regulating the PMII process during male gametogenesis.