Activity of the vascular cambium gives rise to secondary xylem for wood formation in trees. The transcription factor WUSCHEL-related HOMEOBOX4 (WOX4) is a central regulator downstream of the hormone and peptide signaling pathways that maintain cambial activity. However, the genetic regulatory network underlying WOX4-mediated wood formation at the post-transcriptional level remains to be elucidated. In this study, we identified the ubiquitin receptor PagDA1 in hybrid poplar (Populus alba × Populus glandulosa clone 84K) as a negative regulator of wood formation, which restricts cambial activity during secondary growth. Overexpression of PagDA1 in poplar resulted in a relatively reduced xylem due to decreased cambial cell division. By contrast, mutation of PagDA1 by CRISPR/Cas9 resulted in an increased cambial cell activity and promoted xylem formation. Genetic analysis demonstrated that PagDA1 functions antagonistically in a common pathway as PagWOX4 to regulate cambial activity. We propose that PagDA1 physically associates with PagWOX4 and modulates the degradation of PagWOX4 by the 26S proteasome. Moreover, genetic analysis revealed that PagDA1 exerts its negative effect on cambial development by modulating the stability of PagWOX4 in a ubiquitin-dependent manner mediated by the E3 ubiquitin ligase PagDA2. In sum, we have identified a cambial regulatory protein complex, PagDA1-PagWOX4, as a potential target for wood biomass improvement.

中文翻译：

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泛素化 DA1 通过调节杨树中 WOX4 的稳定性负向调节血管形成层活性。

维管形成层的活动导致树木形成次生木质部。转录因子 WUSCHEL 相关的 HOMEOBOX4 (WOX4) 是维持形成层活性的激素和肽信号通路下游的中央调节因子。然而，WOX4 介导的木材形成在转录后水平的遗传调控网络仍有待阐明。在这项研究中，我们鉴定了杂交杨树 (Populus alba × Populus glandulosa clone 84K) 中的泛素受体 PagDA1 作为木材形成的负调节因子，它限制了次生生长过程中的形成层活动。由于形成层细胞分裂减少，PagDA1 在杨树中的过度表达导致木质部相对减少。相比之下，CRISPR/Cas9 对 PagDA1 的突变导致形成层细胞活性增加并促进木质部形成。遗传分析表明，PagDA1 在与 PagWOX4 相同的通路中起拮抗作用，以调节形成层活动。我们提出 PagDA1 与 PagWOX4 物理结合并调节 26S 蛋白酶体对 PagWOX4 的降解。此外，遗传分析表明，PagDA1 通过以 E3 泛素连接酶 PagDA2 介导的泛素依赖性方式调节 PagWOX4 的稳定性，从而对形成层发育产生负面影响。总之，我们已经确定了一种形成层调节蛋白复合物 PagDA1-PagWOX4，作为木材生物量改进的潜在目标。我们提出 PagDA1 与 PagWOX4 物理结合并调节 26S 蛋白酶体对 PagWOX4 的降解。此外，遗传分析表明，PagDA1 通过以 E3 泛素连接酶 PagDA2 介导的泛素依赖性方式调节 PagWOX4 的稳定性，从而对形成层发育产生负面影响。总之，我们已经确定了一种形成层调节蛋白复合物 PagDA1-PagWOX4，作为木材生物量改进的潜在目标。我们提出 PagDA1 与 PagWOX4 物理结合并调节 26S 蛋白酶体对 PagWOX4 的降解。此外，遗传分析表明，PagDA1 通过以 E3 泛素连接酶 PagDA2 介导的泛素依赖性方式调节 PagWOX4 的稳定性，从而对形成层发育产生负面影响。总之，我们已经确定了一种形成层调节蛋白复合物 PagDA1-PagWOX4，作为木材生物量改进的潜在目标。