Tao Yao

In many plant species, the role of ethylene as a phytohormone signal has been well-studied, yet its direct metabolism in planta has received less attention. Consequently, this gap constrains our understanding of the full impact ethylene has on plant systems. During multiple evaluations of resilience in Populus spp., we detected an unidentified metabolite with plausible links to ethylene. In this study, we structurally characterize and identify this molecule as an ethylene glycol glucose conjugate (EGGC). Ethylene-perturbation experiments revealed EGGC abundance is positively correlated with endogenous ethylene production, supporting EGGC as an ethylene-derived metabolite. Stable-isotope labeled EGGC supplied to the media was detected in stem/leaf tissue, indicating long-distance translocation of the compound. These studies also revealed enzymatic conversion in stem/leaf tissue of EGGC to ethylene glycol (EG), with EG levels maintained and accompanied by shifts in downstream metabolites (i.e., glycolic acid and oxalic acid). These results suggest that EGGC participates in active metabolism rather than serves as a waste product. Collectively, the findings reported in this study demonstrate that EGGC is a biomarker of whole-plant ethylene production and position it as a tool for monitoring ethylene-associated stress responses.

Single-cell RNA sequencing (scRNA-seq) has transformed transcriptomic studies by enabling gene expression profiling at the resolution of individual cells within and across a broad range of tissue types, revealing cellular heterogeneity that is obscured in bulk tissue transcriptomes. Over the past decade, improvements in microfluidics and library preparation have drastically increased throughput, allowing tens of thousands of cells to be assayed in a single experiment. Although initially developed in animal systems, scRNA-seq has rapidly emerged as a powerful and widely adopted approach in plant biology. Beyond transcriptomics, the integration of single-cell data with chromatin accessibility, proteomics, metabolomics, and spatial omics is enabling a system-level understanding of plant gene regulation and cellular organization. Network-based analytical frameworks further support the reconstruction of gene regulatory networks and the interpretation of complex single-cell data. In this review, we summarize the current technological landscape of plant single-cell studies, discuss key experimental and analytical challenges, and review emerging strategies for validating single-cell discoveries. We also discuss future directions in applying single-cell technologies to woody perennials plants and bioenergy-relevant crops, emphasizing their potential to accelerate the discovery of cell type-specific regulatory mechanisms underlying growth, stress resilience, and biomass production.

Flooding poses a substantial challenge to plant survival and productivity, particularly in riparian genera like Populus. This study examines the physiological, morphological, metabolic and molecular responses of Populus deltoides Bartram ex Marshall ‘D-124’ and P. trichocarpa Torr. & Gray × P. deltoides hybrid clone ‘52-225’ under control and inundated conditions to identify differences in flooding tolerance. Under flooding conditions, physiological and cellular stress was more pronounced in P. deltoides ‘D-124’ than in the hybrid clone ‘52-225,’ as evidenced by lower transpiration (E), photosynthesis (A) and chlorophyll content. In contrast, ‘52-225’ showed reduced reactive oxygen species accumulation, suggesting better cellular function under stress. Morphologically, ’52-225’ produced more shoot-born roots, which likely enhance oxygen transport and metabolic activity during flooding. Metabolite profiling revealed both overlapping and distinct patterns of sugar and amino acid accumulation between genotypes. Gene expression analysis revealed that flooding-responsive genes, including ALCOHOL DEHYDROGENASE 1 and HYPOXIA RESPONSIVE ERF 2, were activated in both genotypes, with a more pronounced response noted in ‘52-225’. These findings extend our understanding of flooding tolerance mechanisms in Populus by connecting physiological traits, stress responses and genetic regulation. This research contributes to the development of more flooding-resilient poplar varieties, with potential applications in breeding and restoration programs for flooding-prone environments.

SUMMARY: Terpenes play crucial roles in plant growth, development, and stress responses. The biosynthesis of terpenes is influenced by abiotic stress factors, such as drought, temperature, or light. However, the molecular network underlying how terpenes are regulated in response to environmental stimuli remains largely unknown. Here, we identified the autophagy protein SgATG8a as a key mediator of GABA‐regulated terpene production and drought tolerance in Sindora glabra. SgATG8a, evolutionarily related to the animal GABA receptor‐associated protein (GABARAP) subfamily, localizes in both the nucleus and cytoplasm. Exogenous GABA treatment not only increased the expression level of terpene synthase genes (SgTPSs) but also led to enhanced accumulation of six main terpene components in Sindora glabra. In addition, GABA alleviated the photosynthesis damage and enhanced leaf biomass under drought conditions. Consistently, overexpression of SgATG8a in Arabidopsis increased terpene synthase gene (SgTPS) expression, leading to the enhanced production of four major terpenes and improved the tolerance of transgenic plants to drought stress by regulating reactive oxygen species (ROS) scavenging systems. Moreover, the transcription factors SgWRKY13 and SgERF4 were identified as interacting partners of SgATG8a, activating SgTPS3 expression. Lectin receptor‐like kinase (LecRK1) is involved in the GABA‐mediated pathway by interacting with the SgWRKY13/SgERF4‐SgATG8a proteins, and the LecRK1‐SgWRKY13/SgERF4 phosphorylation module fine‐tunes the transcription of the downstream SgTPS3 gene. Taken together, these findings reveal a novel role for GABA in regulating terpene biosynthesis and drought tolerance, providing insights into the molecular mechanism underlying GABA‐mediated terpene production.

Summary: RNA plays a central role in plants, governing various cellular and physiological processes. Monitoring its dynamic abundance provides a discerning understanding of molecular mechanisms underlying plant responses to internal (developmental) and external (environmental) stimuli, paving the way for advances in plant biotechnology to engineer crops with improved resilience, quality and productivity. In general, traditional methods for analysis of RNA abundance in plants require destructive, labour‐intensive and time‐consuming assays. To overcome these limitations, we developed a transformative innovation for in vivo RNA imaging in plants. Specifically, we established a synthetic split ribozyme system that converts various RNA signals to orthogonal protein outputs, enabling in vivo visualisation of various RNA signals in plants. We demonstrated the utility of this system in transient expression experiments (i.e., leaf infiltration in Nicotiana benthamiana) to detect RNAs derived from transgenes and tobacco rattle virus, respectively. Also, we successfully engineered a split ribozyme‐based biosensor in Arabidopsis thaliana for in vivo visualisation of endogenous gene expression at the cellular level, demonstrating the feasibility of multi‐scale (e.g., cellular and tissue level) RNA imaging in plants. Furthermore, we developed a platform for easy incorporation of different protein outputs, allowing for flexible choice of reporters to optimise the detection of target RNAs.

SUMMARY: In plants, pre‐mRNA alternative splicing has been demonstrated to be a crucial tier that regulates gene expression in response to salt stress. However, the underlying mechanisms remain elusive. Here, we studied the roles of DIGEORGE‐SYNDROME CRITICAL REGION 14‐like (AtDGCR14L) in regulating pre‐mRNA splicing and salt stress tolerance. We discovered that Arabidopsis AtDGCR14L is required for maintaining plant salt stress tolerance and the constitutively spliced and active isoforms of important stress‐ and/or abscisic acid (ABA)‐responsive genes. We also identified the interaction between AtDGCR14L and splicing factor U1‐70k, which needs a highly conserved three amino acid (TWG) motif in DGCR14. Different from wild‐type AtDGCR14L, the overexpression of the TWG‐substituted AtDGCR14L mutant did not change salt stress tolerance or pre‐mRNA splicing of stress/ABA‐responsive genes. Additionally, SWITCH3A (SWI3A) is a core subunit of the SWI/SUCROSE NONFERMENTING (SWI/SNF) chromatin‐remodeling complexes. We found that SWI3A, whose splicing depends on AtDGCR14L, actively enhances salt stress tolerance. These results revealed that AtDGCR14L may play an essential role in crosstalk between plant salt‐stress response and pre‐mRNA splicing mechanisms. We also unveiled the potential role of SWI3A in controlling salt stress tolerance. The TWG motif in the intrinsically disordered region of AtDGCR14L is highly conserved and crucial for DGCR14 functions.

SQUAMOSA Promoter-Binding Protein-Like (SPL) transcription factors play vital roles in plant development and stress responses. In this study, we report a comprehensive DNA Affinity Purification sequencing (DAP-seq) analysis for 14 of the 16 SPL transcription factors in Arabidopsis thaliana, providing valuable insights into their DNA-binding specificities. We performed Gene Ontology (GO) analysis of the target genes to reveal their convergent and diverse biological functions among SPL family proteins. Comparative analysis between the paralogs AtSPL9 and AtSPL15 revealed differences in their binding motifs, suggesting divergent regulatory functions. Additionally, we expanded our investigation to homologs of AtSPL9/15 in Zea mays (ZmSBP8/30) and Triticum aestivum (TaSPL7/13), identifying conserved and unique DNA-binding patterns across species. These findings provide key resources for understanding the molecular mechanisms of SPL transcription factors in regulating plant development and evolution across different species.

Horizontal gene transfer (HGT) is a fundamental evolutionary process that plays a key role in bacterial evolution. The likelihood of a successful transfer event is expected to depend on the precise balance of costs and benefits resulting from pathway acquisition. Most experimental analyses of HGT have focused on phenotypes that have large fitness benefits under appropriate selective conditions, such as antibiotic resistance. However, many examples of HGT involve phenotypes that are predicted to provide smaller benefits, such as the ability to catabolize additional carbon sources. We have experimentally reproduced one such HGT event in the laboratory, studying the effects of transferring a pathway for catabolism of the plant-derived aromatic compound salicyl alcohol into soil isolates from the Pseudomonas genus. We find that pathway acquisition enables rapid catabolism of salicyl alcohol with only minor disruptions to existing metabolic and regulatory networks of the new host. However, this new catabolic potential does not confer a measurable fitness advantage during competitive growth in the rhizosphere. We conclude that the phenotype of salicyl alcohol catabolism is readily transferred by HGT but is selectively neutral under environmentally-relevant conditions. We propose that this condition is common and that HGT of many pathways will be self-limiting, because the selective benefits are small and negative frequency-dependent.