Jiarui Pan

SUMMARY Drought stress caused by global warming has resulted in significant tree mortality, driving the evolution of water conservation strategies in trees. Although phytohormones have been implicated in morphological adaptations to water deficits, the molecular mechanisms underlying these processes in woody plants remain unclear. Here, we report that overexpression of PtoMYB142 in Populus tomentosa results in a dwarfism phenotype with reduced leaf cell size, vessel lumen area, and vessel density in the stem xylem, leading to significantly enhanced drought resistance. We found that PtoMYB142 modulates gibberellin catabolism in response to drought stress by binding directly to the promoter of PtoGA2ox4 , a GA 2 ‐oxidase gene induced under drought stress. Conversely, knockout of PtoMYB142 by the CRISPR/Cas9 system reduced drought resistance. Our results show that the reduced leaf size and vessel area, as well as the increased vessel density, improve leaf relative water content and stem water potential under drought stress. Furthermore, exogenous GA 3 application rescued GA‐deficient phenotypes in PtoMYB142 ‐overexpressing plants and reversed their drought resistance. By suppressing the expression of PtoGA2ox4 , the manifestation of GA‐deficient characteristics, as well as the conferred resistance to drought in PtoMYB142 ‐overexpressing poplars, was impeded. Our study provides insights into the molecular mechanisms underlying tree drought resistance, potentially offering novel transgenic strategies to enhance tree resistance to drought.

MicroRNA319 (miR319) and its targets TEOSINTE-BRANCHED1/CYCLOIDEA/PCF (TCP) transcription factors are well-characterized regulators of leaf and flower development, yet their role in root development remains elusive. Here, we demonstrated that overexpression of miR319a led to a decrease in the number and density of lateral roots in poplar, while repressing miR319a by short tandem target mimics (STTM) promoted lateral root (LR) development. The auxin signaling repressors IAA3.1 and IAA3.2 were upregulated in miR319a-OE plants but downregulated in miR319a-STTM plants. After exogenous applications of naphthaleneacetic acid (NAA), which exhibited the characteristics and physiological functions of the endogenous auxin indole-3-acetic acid, the number and density of LR in WT increased by 30% and 44%, respectively. In miR319a-OE plants, the LR number increased by 23% and 48%, and the LR density increased by 10% and 26%. NAA treatment can partially compensate for the phenotype of inhibited LR development caused by the overexpression of miR319a. After N-1-naphthylphthalamic acid (NPA) treatment, which is a key inhibitor of the directional (polar) transport of the auxin hormone in plants, the LR number in WT decreased by 70%. In the overexpression plants, the number of lateral roots decreased by 85–87%, and in the STTM plants, the number of lateral roots decreased by about 83%. It was proved that NPA treatment could reverse the phenotype of increased LR number in miR319a-STTM plants. Expression analysis revealed that miR319a significantly inhibited the expression of the key auxin-regulated genes IAA3.1 and IAA3.2, suggesting that auxin signaling might mediate its effects on lateral root formation. Additionally, we compared the fluorescence signal in the reporter line with GFP expression driven by the auxin-responsive DR5 promoter within the genetic backgrounds of WT, miR319a-OE, and miR319a-STTM plants, which revealed that auxin signaling was stronger in the epidermal cells and elongation zone cells in the LR of miR319a-OE plants, whereas in LR of WT and miR319a-STTM plants, auxin signaling was more pronounced in the root tip meristematic cells. Furthermore, transactivation assays and expression analysis indicated that IAA3.2 was a downstream target of TCP19. Chromatin immunoprecipitation coupled with quantitative PCR (ChIP-qPCR) confirmed that TCP19 directly bound to the promoter region of IAA3.2. These findings establish that miR319a targeted and cleaved TCP19, and TCP19 further directly and negatively regulates the expression of IAA3.2, thereby controlling LR development in Populus tomentosa (P. tomentosa). The formation of LR can expand the plant root system, which is of great significance for the vegetative propagation of plants and the in-vitro regeneration of explants. Moreover, the formation of LR is an important strategy for plants to cope with environmental stresses. This study provides a theoretical basis for breeding poplars more suitable for vegetative propagation.

Wood, as a natural and renewable resource, plays a crucial role in industrial production and daily life. Lignin, as one of the three major components of the plant cell secondary wall, plays a key role in conferring mechanical strength and enhancing stress resistance. The caffeic acid-O-methyltransferase (COMT) family of oxygen-methyltransferases is a core regulatory node in the downstream pathway of lignin biosynthesis. Here, our report shows that caffeic acid-O-methyltransferase 2 (COMT2) exhibits high conservation across several species. Tissue expression analysis reveals that COMT2 is specifically highly expressed in the secondary xylem of Populus tomentosa stems. We demonstrated that the specific overexpression of COMT2 in fiber cells of Populus tomentosa led to a significant increase in plant height, stem diameter, internode number, and stem dry weight. Furthermore, we found that the specific overexpression of COMT2 in fiber cells promotes xylem differentiation, lignin accumulation, and the thickening of the secondary cell wall (SCW) in fiber cells. Our results indicate that key downstream lignin biosynthesis enzyme genes are upregulated in transgenic plants. Additionally, mechanical properties of stem bending resistance, puncture resistance, and compressive strength in the transgenic lines are significantly improved. Moreover, we further created the DUFpro:COMT2 transgenic lines of Populus deltoides × Populus. euramericana cv ‘Nanlin895’ to verify the functional conservation of COMT2 in closely related poplar species. The DUFpro:COMT2 Populus deltoides × Populus. euramericana cv ‘Nanlin895’ transgenic lines exhibited phenotypes similar to those observed in the P. tomentosa transgenic plants, which showed enhanced growth, increased lignin accumulation, and greater wood strength. Overall, the specific overexpression of the caffeic acid O-methyltransferase gene COMT2 in poplar stem fiber cells has enhanced the wood biomass, wood properties, and mechanical strength of poplar stems.