Erxu Pi

Since the first plant gene essential for arbuscular mycorrhizal (AM) symbiosis was identified, more than 170 such genes have been discovered. However, these findings remain scattered across species, and a systematic synthesis is urgently needed to guide future functional studies and breeding applications. This review provides a systematic classification and contextual overview of the currently reported plant genes essential for AM symbiosis, covering leguminous species (e.g., Medicago truncatula, Lotus japonicus) and non-leguminous species (e.g., Oryza sativa, Solanum lycopersicum). We classify these genes into functional modules corresponding to key stages of AM symbiosis: SL-mediated pre-symbiotic signaling, chitin signal perception, activation of the common symbiosis signaling pathway (CSSP), calcium (Ca²⁺) oscillation generation, arbuscule development and maintenance, and nutrient exchange. Beyond classification, we highlight conserved genetic modules across plant lineages and discuss their implications for engineering AM symbiosis in non-host or poorly symbiotic crops. This synthesis establishes a foundational genetic resource for molecular breeding aimed at enhancing nutrient-use efficiency and sustainable crop production.

The calcium/calmodulin-dependent protein kinase (CCaMK) DMI3 (Doesn't Make Infections 3) decodes rhizobia-induced Ca2+ signals through phosphorylation of its interacting partner, IPD3, thereby initiating root nodule symbiosis (RNS). However, the precise mechanism by which DMI3 activates this pathway remains unclear. Here, we show that phosphorylation of a triplet motif (S343S344T345) located at the C-terminus of the calmodulin-binding domain acts as a molecular switch. We identified T345 as a phosphorylation site that exhibits regulatory functions similar to those of S343 and S344. Phosphomimic mutations at any single residue of the 3 sites blocked DMI3-dependent nodulation, indicating that the kinase must initially remain nonphosphorylated at these positions. Conversely, simultaneous phosphorylation of at least 2 residues was required for symbiotic activity, revealing a transition from a phosphorylation-free state to a hyperphosphorylated status (simultaneous phosphorylation of at least 2 residues) at these sites. Phosphomimic mutations within the triplet enhanced DMI3 autophosphorylation (auto-P) but severely impaired its interaction with Ca2+/CaM and IPD3. In addition, the physical interaction between DMI3 and IPD3 suppressed IPD3-triggered MtNIN transcription and nodulation, indicating that DMI3 also acts as a negative regulator by sequestering activated IPD3. Thus, we propose a model in which repeated auto-P at the triplet contributes to RNS activation by dissociating the CaM-DMI3-IPD3 complex, thereby releasing phosphorylated and activated IPD3 to initiate the downstream transcriptional cascade. Our findings uncover a dual role for DMI3—as both a kinase and a scaffold—and clarify how auto-P within the triplet motif licenses IPD3 to trigger RNS.

Increasing biotic and abiotic stresses are seriously impeding the growth and yield of staple crops and threatening global food security. As one of the largest classes of regulators in vascular plants, WRKY transcription factors play critical roles governing flavonoid biosynthesis during stress responses. By binding major W-box cis-elements (TGACCA/T) in target promoters, WRKYs modulate diverse signaling pathways. In this review, we optimized existing WRKY phylogenetic trees by incorporating additional plant species with WRKY proteins implicated in stress tolerance and flavonoid regulation. Based on the improved frameworks and documented results, we aim to deduce unifying themes of distinct WRKY subfamilies governing specific stress responses and flavonoid metabolism. These analyses will generate experimentally testable hypotheses regarding the putative functions of uncharacterized WRKY homologs in tuning flavonoid accumulation to enhance stress resilience.

Ethylene Responsive Factor (ERF) subfamily comprise the largest number of proteins in the plant AP2/ERF superfamily, and have been most extensively studied on the biological functions. Members of this subfamily have been proven to regulate plant resistances to various abiotic stresses, such as drought, salinity, chilling and some other adversities. Under these stresses, ERFs are usually activated by mitogen-activated protein kinase induced phosphorylation or escape from ubiquitin-ligase enzymes, and then form complex with nucleic proteins before binding to cis-element in promoter regions of stress responsive genes. In this review, we will discuss the phylogenetic relationships among the ERF subfamily proteins, summarize molecular mechanism how the transcriptional activity of ERFs been regulated and how ERFs of different subgroup regulate the transcription of stress responsive genes, such as high-affinity K⁺ transporter gene PalHKT1;2, reactive oxygen species related genes LcLTP, LcPrx, and LcRP, flavonoids synthesis related genes FtF3H and LhMYBSPLATTER, etc. Though increasing researches demonstrate that ERFs are involved in various abiotic stresses, very few interact proteins and target genes of them have been comprehensively annotated. Hence, future research prospects are described on the mechanisms of how stress signals been transited to ERFs and how ERFs regulate the transcriptional expression of stress responsive genes.

Protein phosphorylation is one of the most important posttranslational modifications. The phosphorylation and dephosphorylation of proteins regulate almost every cellular process, and the understanding of their functions can provide insights into the regulation of living systems at the molecular level. In recent years, both the rapid developments of enrichment approaches for phosphoproteins and MS techniques have improved the research scope and depth of phosphoproteomics. Using NaCl-treated soybean roots as the experimental materials, this chapter introduces the protein extraction, digestion with filter-aided sample preparation (FASP), eight-plex iTRAQ labeling, TiO₂-based enrichment of phosphopeptides, LC-MS/MS analysis, as well as bioinformatic methods and protocols.

Basic helix-loop-helix proteins (bHLHs) comprise one of the largest families of transcription factors in plants. They have been shown to be involved in responses to various abiotic stresses, such as drought, salinity, chilling, heavy metal toxicity, iron deficiency, and osmotic damages. By specifically binding to cis-elements in the promoter region of stress related genes, bHLHs can regulate their transcriptional expression, thereby regulating the plant’s adaptive responses. This review focuses on the structural characteristics of bHLHs, the regulatory mechanism of how bHLHs are involved transcriptional activation, and the mechanism of how bHLHs regulate the transcription of target genes under various stresses. Finally, as increasing research demonstrates that flavonoids are usually induced under fluctuating environments, the latest research progress and future research prospects are described on the mechanisms of how flavonoid biosynthesis is regulated by bHLHs in the regulation of the plant’s responses to abiotic stresses.

The bZIP family is one of the most widely distributed and conserved transcription factor categories in eukaryotes. They are widely involved in processes of plant abiotic stress responses, such as improving the ability of plants to adapt to drought, salinity and cold. To comprehensively understand how plant bZIPs regulating these tolerances, this article will have an overview of their distributions on different species, characteristics of protein architecture, regulation mechanisms for target genes’ transcriptional expression, and relevant research progresses in recent years. Specially, we will discuss the possible roles of flavonoids, whose synthesis could be regulated by bZIPs, in abovementioned plant tolerant responses.

Background: Trees of the genusTaxusare highly valuable medicinal plants with multiple pharmacological effects on various cancer treatments. Paclitaxel fromTaxustrees is an efficient and widely used anticancer drug, however, the accumulation of taxoids and other active ingredients can vary greatly amongTaxusspecies. In our study, the metabolomes of threeTaxusspecies have been investigated.Results: A total of 2246 metabolites assigned to various primary and secondary metabolic pathways were identified using an untargeted approach. Analysis of differentially accumulated metabolites identified 358 T. media-, 220 T. cuspidata-, and 169 T. mairei-specific accumulated metabolites, respectively. By searching the metabolite pool, 7 MEP pathway precursors, 11 intermediates, side chain products and derivatives of paclitaxel, and paclitaxel itself were detected. Most precursors, initiated intermediates were highly accumulated inT. mairei, and most intermediate products approaching the end point of taxol biosynthesis pathway were primarily accumulated inT. cuspidataandT. media. Our data suggested that there were higher-efficiency pathways to paclitaxel inT. cuspidataandT. mediacompared with inT. mairei. As an important class of active ingredients inTaxustrees, a majority of flavonoids were predominantly accumulated inT. maireirather thanT. mediaandT. cuspidata. The variations in several selected taxoids and flavonoids were confirmed using a targeted approach.Conclusions: Systematic correlativity analysis identifies a number of metabolites associated with paclitaxel biosynthesis, suggesting a potential negative correlation between flavonoid metabolism and taxoid accumulation. Investigation of the variations in taxoids and other active ingredients will provide us with a deeper understanding of the interspecific differential accumulation of taxoids and an opportunity to accelerate the highest-yielding species breeding and resource utilization.