Department of Plant Biology, University of Georgia, Athens, GA 30602, USA.
Biomedical Microscopy Core, University of Georgia, 164 Coverdell Center, Athens, GA 30602, USA.
Plant Cell Physiol. 2021 Mar 25;62(1):53-65. doi: 10.1093/pcp/pcaa140.
Secondary wall deposition in xylem vessels is activated by Vascular-Related NAC Domain proteins (VNDs) that belong to a group of secondary wall NAC (SWN) transcription factors. By contrast, Xylem NAC Domain1 (XND1) negatively regulates secondary wall deposition in xylem vessels when overexpressed. The mechanism by which XND1 exerts its functions remains elusive. We employed the promoter of the fiber-specific Secondary Wall-Associated NAC Domain1 (SND1) gene to ectopically express XND1 in fiber cells to investigate its mechanism of action on secondary wall deposition. Ectopic expression of XND1 in fiber cells severely diminished their secondary wall deposition and drastically reduced the expression of SWN-regulated downstream transcription factors and secondary wall biosynthetic genes but not that of the SWN genes themselves. Transactivation analyses revealed that XND1 specifically inhibited SWN-activated expression of these downstream genes but not their MYB46-activated expression. Both the NAC domain and the C-terminus of XND1 were required for its inhibitory function and its NAC domain interacted with the DNA-binding domains of SWNs. XND1 was shown to be localized in the cytoplasm and the nucleus and its co-expression with VND6 resulted in the cytoplasmic sequestration of VND6. Furthermore, the C-terminus of XND1 was indispensable for the XND1-mediated cytoplasmic retention of VND6 and its fusion to VND6 was able to direct VND6 to the cytoplasm and render it unable to activate the gene expression. Since the XND1 gene is specifically expressed in xylem cells, these results indicate that XND1 acts through inhibiting VND functions to negatively regulate secondary wall deposition in xylem vessels.
木质部导管次生壁的沉积是由血管相关 NAC 结构域蛋白（VNDs）激活的，VNDs 属于次生壁 NAC（SWN）转录因子的一个亚类。相比之下，过量表达时，木质部 NAC 结构域 1（XND1）负调控木质部导管次生壁的沉积。XND1 发挥功能的机制仍不清楚。我们利用纤维特异性次生壁相关 NAC 结构域 1（SND1）基因的启动子在纤维细胞中外源表达 XND1，以研究其对次生壁沉积的作用机制。纤维细胞中外源表达 XND1 严重减少了次生壁的沉积，并显著降低了 SWN 调控的下游转录因子和次生壁生物合成基因的表达，但 SWN 基因本身的表达不受影响。转录激活分析表明，XND1 特异性抑制 SWN 激活的这些下游基因的表达，而不是 MYB46 激活的表达。XND1 的 NAC 结构域和 C 端都需要其抑制功能，其 NAC 结构域与 SWN 的 DNA 结合结构域相互作用。XND1 被证明定位于细胞质和细胞核中，与 VND6 共表达导致 VND6 在细胞质中的隔离。此外，XND1 的 C 端对于 XND1 介导的 VND6 细胞质保留是必不可少的，其与 VND6 的融合能够将 VND6 导向细胞质并使其无法激活基因表达。由于 XND1 基因在木质部细胞中特异性表达，这些结果表明 XND1 通过抑制 VND 功能负调控木质部导管次生壁的沉积。
