Gene network analysis of poplar root transcriptome in response to drought stress identifies a PtaJAZ3PtaRAP2.6-centered hierarchical network

Using time-series transcriptomic data from poplar roots undergoing polyethylene glycol (PEG)-induced drought stress, we built a genetic network model of the involved putative molecular responses. We found that the network resembled a hierarchical structure. The highest hierarchical level in this structure is occupied by 9 genes, which we called superhubs because they were primarily connected to 18 hub genes, which are then connected to 2,934 terminal genes. We were only able to regenerate transgenic plants overexpressing two of the superhubs, suggesting that the majority of the superhubs might interfere with the regeneration process and did not allow recovery of transgenic plants. The two superhubs encode proteins with closest homology to JAZ3 and RAP2.6 genes of Arabidopsis and were consequently named PtaJAZ3 and PtaRAP2.6. PtaJAZ3 and PtaRAP2.6 overexpressing transgenic lines showed a significant increase in both root elongation and lateral root proliferation and these responses were specific for the drought stress conditions and were highly correlated with the levels of overexpression of the transgenes. Several lines of evidence suggest of regulatory interactions between the two superhubs. Both superhubs were significantly induced by methyl jasmonate (MeJA). Because jasmonate signaling involves ubiquitin-mediated proteasome degradation, treatment with proteasome inhibitor abolished the MeJA induction for both genes. PtaRAP2.6 was upregulated in PtaJAZ3 transgenics but PtaJAZ3 expression was not affected in the PtaRAP2.6 overexpressors. The discovery of the two genes and further future insights into the associated mechanisms can lead to improved understanding and novel approaches to regulate root architecture in relation to drought stress.本研究基于经聚乙二醇（polyethylene glycol, PEG）模拟干旱胁迫的杨树根部时序转录组数据，构建了参与该胁迫应答过程的推定分子应答遗传网络模型。研究发现该遗传网络呈现层级结构特征：该层级结构的最高层由9个基因构成，我们将其命名为超级枢纽基因（superhubs），因其主要与18个枢纽基因（hub genes）相连，而这些枢纽基因又进一步连接至2934个末端基因。本研究仅成功获得了2个超级枢纽基因过表达的转基因植株，这表明多数超级枢纽基因可能会干扰植株再生过程，导致无法获得对应的转基因植株。这两个超级枢纽基因编码的蛋白与拟南芥（Arabidopsis）的JAZ3和RAP2.6基因同源性最高，因此分别被命名为PtaJAZ3和PtaRAP2.6。过表达PtaJAZ3和PtaRAP2.6的转基因株系，其主根伸长与侧根增殖均显著提升，且该应答反应仅在干旱胁迫条件下出现，并与转基因的过表达水平呈高度正相关。多项实验证据表明这两个超级枢纽基因之间存在调控互作关系：两个超级枢纽基因均能被茉莉酸甲酯（methyl jasmonate, MeJA）显著诱导；由于茉莉酸信号通路依赖泛素介导的蛋白酶体降解途径，蛋白酶体抑制剂处理可完全阻断MeJA对这两个基因的诱导表达。在过表达PtaJAZ3的转基因植株中，PtaRAP2.6的表达量显著上调，但在过表达PtaRAP2.6的转基因株系中，PtaJAZ3的表达并未受到影响。本研究发现的这两个基因，以及后续对其相关分子机制的深入解析，将有助于我们更好地理解干旱胁迫下杨树根部构型的调控机制，并为开发新型根部构型调控策略提供理论依据。