Ghent University, Department of Plant Biotechnology and Bioinformatics, Technologiepark 71, B-9052 Ghent, Belgium.
VIB Center for Plant Systems Biology, Technologiepark 71, B-9052 Ghent, Belgium.
Sci Adv. 2024 Mar 15;10(11):eadj2570. doi: 10.1126/sciadv.adj2570. Epub 2024 Mar 13.
Ubiquitination plays a crucial role throughout plant growth and development. The E3 ligase DA2 has been reported to activate the peptidase DA1 by ubiquitination, hereby limiting cell proliferation. However, the molecular mechanisms that regulate DA2 remain elusive. Here, we demonstrate that DA2 has a very high turnover and auto-ubiquitinates with K48-linkage polyubiquitin chains, which is counteracted by two deubiquitinating enzymes, UBIQUITIN-SPECIFIC PROTEASE 12 (UBP12) and UBP13. Unexpectedly, we found that auto-ubiquitination of DA2 does not influence its stability but determines its E3 ligase activity. We also demonstrate that impairing the protease activity of DA1 abolishes the growth-reducing effect of DA2. Last, we show that synthetic, constitutively activated DA1-ubiquitin fusion proteins overrule this complex balance of ubiquitination and deubiquitination and strongly restrict growth and promote endoreduplication. Our findings highlight a nonproteolytic function of K48-linked polyubiquitination and reveal a mechanism by which DA2 auto-ubiquitination levels, in concert with UBP12 and UBP13, precisely monitor the activity of DA1 and fine-tune plant organ size.
泛素化在植物生长和发育过程中起着至关重要的作用。已有报道称 E3 连接酶 DA2 通过泛素化激活肽酶 DA1，从而限制细胞增殖。然而，调节 DA2 的分子机制仍不清楚。在这里，我们证明 DA2 的周转率非常高，并通过 K48 连接的多泛素链进行自身泛素化，这被两种去泛素化酶 UBIQUITIN-SPECIFIC PROTEASE 12 (UBP12) 和 UBP13 抵消。出乎意料的是，我们发现 DA2 的自身泛素化不会影响其稳定性，而是决定其 E3 连接酶活性。我们还证明，破坏 DA1 的蛋白酶活性会消除 DA2 的生长抑制作用。最后，我们表明，合成的、组成性激活的 DA1-泛素融合蛋白会破坏这种泛素化和去泛素化的复杂平衡，并强烈限制生长并促进内复制。我们的研究结果强调了 K48 连接的多泛素化的非蛋白水解功能，并揭示了 DA2 自身泛素化水平与 UBP12 和 UBP13 协同作用，精确监测 DA1 活性并精细调节植物器官大小的机制。
