Stages Of Leaf Development Research Articles

Early Initiation of Bundle Sheath Cells During Leaf Development as Visualised by SCARECROW Expression in Dicotyledonous C4 Plants.

The C4 type of dicotyledonous plants exhibit a higher density of reticulate veins than the C3 type, with a nearly 1:1 ratio of mesophyll cells (MCs) to bundle sheath cells (BSCs). To understand how this C4-type cell pattern is formed, we identified two SCARECROW (SCR) genes in C4 Flaveria bidentis, FbSCR1 and FbSCR2, that fully or partially complement the endodermal cell layer-defective phenotype of Arabidopsis scr mutant. We then created FbSCRs promoter β-glucuronidase reporter (GUS) lines of F. bidentis, which showed GUS expression in BSCs and their progenitor cells. The GUS expression pattern in F. bidentis transformants and comparison with the closely related C3-type Flaveria pringlei revealed that higher-order veins were initiated in the early leaf developmental stage. Treatment with an auxin polarity transport inhibitor decreased the MC area and led to vein formation without free ends, resulting in the formation of BSCs in positions adjacent to other BSCs. However, BSC differentiation was not affected, as evidenced by BSC specific FbSCR1 expression and RuBisCO accumulation. These results indicate that polar auxin transport is important for MC proliferation and/or differentiation, which leads to the formation of a C4-type cell pattern in which MCs and BSCs are equally adjacent.

Sequential Defense Strategies: From Ant Recruitment to Leaf Toughness.

Plants express many types of defenses in response to herbivory damage. These defenses can be displayed as a sequence or they can overlap, increasing efficiency in protection. However, leaf defense shifts during leaf development, including extrafloral nectaries (EFNs), are neglected in natural tropical systems. To address this gap, our study evaluates the shifts in defense strategies of Eriotheca gracilipes, focusing on extrafloral nectaries and leaf toughness across leaf development stages. We also simulate herbivory by cutting the leaves to address the role of visiting ants against herbivores. We observed that E. gracilipes exhibits a defense turnover, shifting from indirect defenses (e.g., EFNs) in young leaves to physical defenses in adult leaves. Simulated herbivory led to heightened ant visitation, which correlated with decreased herbivory rates, indicating that ant recruitment acts as an effective deterrent. We observed a peak of EFN activity in young leaves, increased foliar toughness in adult leaves, and reduced herbivory on ant-patrolled young leaves. Additionally, E. gracilipes demonstrated tolerance to up to 10% foliar loss with no significant impact on leaf asymmetry, although 50% foliar loss increased asymmetry in newly flushed leaves. These results highlight E. gracilipes' adaptive flexibility by attracting protective ants when vulnerable and enhancing structural defenses as leaves develops, E. gracilipes minimizes herbivory impact. This study provides valuable insight into the adaptive roles of EFNs and tolerance in E. gracilipes, contributing to a broader understanding of plant defense strategies.

Effect of Nitrogen Addition on the Intra-Annual Leaf and Stem Traits and Their Relationships in Two Dominant Species in a Subtropical Forest

Exploring the relationship between leaf and xylem growth and nitrogen (N) deposition on tree growth helps us better understand how N deposition affects tree growth, but relevant studies are still limited. We tested if leaf and xylem growth and their relationship were affected by N deposition across multiple stages of leaf development. We studied two dominant tree species (Castanopsis chinensis and Schima superba) in a subtropical forest in South China, monitoring their leaf and xylem growth traits under canopy N addition of 0 (CK) and 50 kg N ha−1 year−1 (CAN) and compared these traits and their relationships across different treatments, periods, and species. We found that CAN significantly increased C. chinensis’s leaf carbon (C) content, while the same treatment reduced leaf C content in S. superba. Specific leaf area (SLA), specific leaf weight (SLW), leaf N content, and C:N ratio showed more variation between species and within species across developmental stages than in response to different N addition treatments. Nitrogen addition also significantly increased the xylem width in both species, but the weekly xylem increment before July was notably higher in S. superba than in C. chinensis. Moreover, the leaf C content during early leaf development (ELD) following N addition was significantly and negatively correlated with the start date of xylogenesis in C. chinensis but had no significant effect on the onset of xylogenesis in S. superba. Additionally, regardless of species, SLA in ELD was negatively correlated with xylem width at the end of the early growing season (EW) under CK. However, this relationship became insignificant under CAN, suggesting that N addition alleviated the trade-off between SLA in ELD and EW. These results demonstrated that the relationship between leaf and xylem growth is influenced by both the leaf developmental stage and N availability. Traditional analyses of the leaf–xylem relationship, based solely on mature leaf traits, might oversimplify the effects of N deposition on tree growth.

Dynamic Chromatin Accessibility and Gene Expression Regulation During Maize Leaf Development.

Chromatin accessibility is closely associated with transcriptional regulation during maize (Zea mays) leaf development. However, its precise role in controlling gene expression at different developmental stages remains poorly understood. This study aimed to investigate the dynamics of chromatin accessibility and its influence on genome-wide gene expression during the BBCH_11, BBCH_13, and BBCH_17 stages of maize leaf development. Maize leaves were collected at the BBCH_11, BBCH_13, and BBCH_17 developmental stages, and chromatin accessibility was assessed using ATAC-seq. RNA-seq was performed to profile gene expression. Integrated analysis of ATAC-seq and RNA-seq data was conducted to elucidate the relationship between chromatin accessibility and transcriptional regulation. A total of 46,808, 38,242, and 41,084 accessible chromatin regions (ACRs) were identified at the BBCH_11, BBCH_13, and BBCH_17 stages, respectively, with 23.4%, 12.2%, and 21.9% of these regions located near transcription start sites (TSSs). Integrated analyses revealed that both the number and intensity of ACRs significantly influence gene expression levels. Motif analysis identified key transcription factors associated with leaf development and potential transcriptional repressors among genes, showing divergent regulation patterns in ATAC-seq and RNA-seq datasets. These findings demonstrate that chromatin accessibility plays a crucial role in regulating the spatial and temporal expression of key genes during maize leaf development by modulating transcription factor binding. This study provides novel insights into the regulatory mechanisms underlying maize leaf development, contributing to a deeper understanding of chromatin-mediated gene expression.

Ecophysiological and Molecular Analysis of Contrasting Genotypes for Leaf Senescence in Sunflower (Helianthus annuus L.) Under Differential Doses of N in Soil.

Leaf senescence in plants is the last stage of leaf development and is characterized by a decline in photosynthetic activity, an active degeneration of cellular structures, and the recycling of accumulated nutrients to areas of active growth, such as buds, young leaves, flowers, fruits, and seeds. This process holds economic significance as it can impact yield, influencing the plant's ability to maintain an active photosynthetic system during prolonged periods, especially during the grain filling stage, which affects plant weight and oil content. It can be associated with different stresses or environmental conditions, manifesting itself widely in the context of climate change and limiting yield, especially in crops of agronomic relevance. In this work, we study the stability of two widely described sunflower (Helianthus annuus L.) genotypes belonging to the INTA Breeding Program against differential N conditions, to verify their yield stability in control conditions and under N supply. Two inbred lines were utilized, namely R453 (early senescence) and B481-6 (late senescence), with contrasting nitrogen availability in the soil but sharing the same ontogeny cycle length. It was observed that, starting from R5.5, the B481-6 genotype not only delayed senescence but also exhibited a positive response to increased nitrogen availability in the soil. This response included an increase in intercepted radiation, resulting in a statistically significant enhancement in grain yield. Conversely, the R453 genotype did not show significant differences under varying nitrogen availability and exhibited a tendency to decrease grain yield when nitrogen availability was increased. The response to nitrogen can vary depending on the specific genotype.

Genetic Foundation of Leaf Senescence: Insights from Natural and Cultivated Plant Diversity.

Leaf senescence, the final stage of leaf development, is crucial for plant fitness as it enhances nutrient reutilization, supporting reproductive success and overall plant adaptation. Understanding its molecular and genetic regulation is essential to improve crop resilience and productivity, particularly in the face of global climate change. This review explores the significant contributions of natural genetic diversity to our understanding of leaf senescence, focusing on insights from model plants and major crops. We discuss the physiological and adaptive significance of senescence in plant development, environmental adaptation, and agricultural productivity. The review emphasizes the importance of natural genetic variation, including studies on natural accessions, landraces, cultivars, and artificial recombinant lines to unravel the genetic basis of senescence. Various approaches, from quantitative trait loci mapping to genome-wide association analysis and in planta functional analysis, have advanced our knowledge of senescence regulation. Current studies focusing on key regulatory genes and pathways underlying natural senescence, identified from natural or recombinant accession and cultivar populations, are highlighted. We also address the adaptive implications of abiotic and biotic stress factors triggering senescence and the genetic mechanisms underlying these responses. Finally, we discuss the challenges in translating these genetic insights into crop improvement. We propose future research directions, such as expanding studies on under-researched crops, investigating multiple stress combinations, and utilizing advanced technologies, including multiomics and gene editing, to harness natural genetic diversity for crop resilience.

Exploring silica accumulation in bamboo leaves: A study on phytolith morphology and epidermal patterning in the tropical giant bamboo Dendrocalamus copelandii.

Bamboo is a grass in the Poaceae family with various applications. Bamboo leaves can accumulate high silica. However, silica deposition in bamboo has received limited study. Therefore, this research investigated silica accumulation in Dendrocalamus copelandii leaves. The study includes the localisation of silica through phytolith morphology, examination of the distribution patterns of phytoliths in epidermal tissues, analysis of silica accumulation within specialised silica cells (short cells), and analysis of silicon concentration across various leaf developmental stages. We employed imaging techniques, including Differential interference contrast and Scanning electron microscope incorporated with an energy-dispersive X-ray spectrometer, to investigate silica accumulation in bamboo leaves. We also analysed silicon concentration using Inductively coupled plasma-optical emission spectroscopy. Leaves of D. copelandii exhibited 11 phytolith morphotypes, such as BILOBATE, POLYLOBATE, SADDLE, ACUTE, ACUTE BULBOSUS, MICROHAIR, STOMATA, BULLIFORM FLABELLATE, ELONGATE SINUATE, ELONGATE ENTIRE and TRACHEARY. Most of these phytoliths were found in short cells (BILOBATE, POLYLOBATE and SADDLE) of epidermal tissues. The short cells were arranged transversely along the leaf length. BILOBATE was found in both the abaxial and adaxial epidermis, while SADDLE was found only in the abaxial epidermis. Silica accumulation in the short cells of unexpanded leaves begins at the leaf apex, spreads to the middle and base positions, and accumulates first in the abaxial before the adaxial epidermis. Moreover, bamboo leaves accumulate more silicon concentration as they age. Phytolith morphotypes and silica accumulation in epidermal short cells are key factors in understanding silica deposition. Leaf age and climate significantly impact silicon concentration in bamboo leaves. Our findings are informative for archaeological studies and for plant taxonomical classification. The results are also applicable for biotechnological applications.

Mapping of quantitative trait loci (QTL) in Brassica napus L. for tolerance to water stress.

Brassica napus L. plants are sensitive to water stress conditions throughout their life cycle from seed germination to seed setting. This study aims at identifying quantitative trait loci (QTL) linked to B. napus tolerance to water stress mimicked by applications of 10% polyethylene glycol-6000 (PEG-6000). Two doubled haploid populations, each consisting of 150 genotypes, were used for this research. Plants at the two true leaf stage of development were grown in the absence (control) or presence (stress) of PEG-6000 under controlled environmental conditions for 48h, and the drought stress index was calculated for each genotype. All genotypes, along with their parents, were genotyped using the Brassica Infinium 90K SNP BeadChip Array. Inclusive composite interval mapping was used to identify QTL. Six QTL and 12 putative QTL associated with water stress tolerance were identified across six chromosomes (A2, A3, A4, A9, C3, and C7). Collectively, 2154 candidate genes for water stress tolerance were identified for all the identified QTL. Among them, 213 genes were identified as being directly associated with water stress (imposed by PEG-6000) tolerance based on nine functional annotations. These results can be incorporated into future breeding initiatives to select plant material with the ability to cope effectively with water stress.

Integrated Transcriptomic and Metabolomic Analysis Reveals Possible Molecular Mechanisms of Leaf Growth and Development in Disanthus cercidifolius var. longipes.

Background:Disanthus cercidifolius var. longipes is an ancient relic plant unique to China. However, the typical shade-loving plant is largely exposed to the sun, which poses a major challenge to its conservation. Methods: This study explored dynamic changes in primary and secondary metabolites in D. cercidifolius leaves at different stages of development, combining metabolomics and transcriptome analysis to discuss the differentially accumulated metabolites (DAMs) and differentially expressed genes (DEGs). Results: The DAMs and DEGs were enriched in pathways related to photosynthesis, carbon (C) metabolism, anthocyanin synthesis, plant hormone signal transduction, and flavonoid synthesis. At the initial stage of leaf development, many primary metabolites were synthesized in the leaves. Before leaf maturity, many primary metabolites were converted into secondary metabolites. Combined transcriptome and metabolome analysis showed that the metabolites and genes related to anthocyanin synthesis and flavonoid metabolism were upregulated. In contrast, the genes related to C metabolism and C fixation were downregulated. After leaf maturity, photosynthetic capacity increased, total flavonoid content peaked (implying the strongest photoprotection capacity), and the transformation of anthocyanins and flavonoids was weakened. Conclusions: Light intensity indirectly affects the accumulation of the primary and secondary metabolism of D. cercidifolius. With the enhancement of photoprotection, the photosynthetic energy capacity decreases. It is, therefore, inferable that D. cercidifolius has shading properties and achieves a stable nutrient supply during growth and development through these strategies. Thus, D. cercidifolius protection requires a shaded environment.

Light Energy Use Efficiency in Photosystem II of Tomato Is Related to Leaf Age and Light Intensity

The fundamental key to increase photosynthetic efficiency of crop plants lies in optimizing the light energy use efficiency. In our study, we used tomato to evaluate the allocation of absorbed light energy in young and mature leaves, and to estimate if the extent of photoinhibition and photoprotection can be affected by the leaf age. A reduced efficiency of the oxygen-evolving complex, in young leaves compared to mature ones, resulted in a donor-side photoinhibition, as judged from the significantly lower Fv/Fm ratio, in young leaves. The detected increased 1O2 production in young leaves was probably due to a donor-side photoinhibition. The effective quantum yield of photosystem II (PSII) photochemistry (ΦPSII), at low light intensity (LLI, 426 μmol photons m−2 s−1), was significantly lower in young compared to mature leaves. Moreover, the non-significant increase in non-photochemical energy loss in PSII (ΦNPQ) could not counteract the decreased ΦPSII, and as a result the non-regulated energy loss in PSII (ΦNO) increased in young leaves, compared to mature ones. The significantly lower ΦPSII in young leaves can be attributed to the increased reactive oxygen species (ROS) creation that diminished the efficiency of the open PSII reaction centers (Fv’/Fm’), but without having any impact on the fraction of the open reaction centers. The reduced excess excitation energy, in mature leaves compared to young ones, at LLI, also revealed an enhanced PSII efficiency of mature leaves. However, there was almost no difference in the light energy use efficiency between young and mature leaves at the high light intensity (HLI, 1000 μmol photons m−2 s−1). The ability of mature tomato leaves to constrain photoinhibition is possible related to an enhanced photosynthetic function and a better growth rate. We concluded that the light energy use efficiency in tomato leaves is influenced by both the leaf age and the light intensity. Furthermore, the degrees of photoinhibition and photoprotection are related to the leaf developmental stage.

Impact of field drying at different initial leaf ages on growth and yield formation of direct seeding rice in Chengdu plain

AbstractTo determine the optimal drying period for direct seeding rice, we conducted experiments using two rice cultivars: hybrid indica (Shuyou 217) and conventional indica (Huang Huazhan). The experiments explored the influence of varying initial leaf development stages on tillering dynamics, leaf area, dry matter production, yield, and yield components in direct seeding rice. The results indicated a significant correlation between the initial leaf development stage and the yield of direct seeding rice. As the initial leaf development stage progressed, the number of effective panicles increased, while key traits affecting panicle yield, such as grain/spikelet? Number per panicle, seed setting rate, and 1000-grain weight, decreased. Additionally, varietal differences in tillering reduction were observed across different leaf development stages. Shuyou 217 exhibited a decrease in heading rate with increasing leaf development stage, whereas Huang Huazhan showed no significant difference. With delayed seeding planting age, both cultivars demonstrated an increasing trend in the leaf area index, initially leading to a rise in effective leaf area proportion, followed by a decline. At the full heading and maturity stages, the dry weight of the stem sheath and panicles per plant decreased as the leaf development stage was delayed. However, the output and transport rates of stem sheath assimilated first increased and then decreased. From a practical perspective, field drying management should be aligned with varietal characteristics to reduce ineffective tillering, ensure sufficient dry matter accumulation during the full heading stage, and optimize post-heading dry matter export and transport. This strategy promotes the coordination between the “source” and “sink” dynamics in rice, ultimately maximizing yield in direct-seeded rice systems.

Use of infrared and mass spectrometry for the discrimination of metabolites from different stages of Schinus terebinthifolia Raddi leaves

The Brazilian Peppertree (Schinus terebinthifolia Raddi) is a medicinal plant with several biological activities, the most important of which are anti-inflammatory, antimicrobial and antiproliferative. Therefore, scientific research and investigation of its bioactive compounds are required. Leaves at three development stages were collected from three different genotypes of S. terebinthifolia. Methanol extracts were prepared and analyzed by infrared and mass spectrometry. The data obtained were treated by principal component analysis (PCA). The PCA model applied to the mass spectrometry data proved to be efficient to discriminate the three stages of development, in addition to indicating a differentiated tendency in the production of secondary metabolites by the genotypes. Two principal components were needed to describe the variance of the infrared spectrometry data (81.96%), while for mass spectrometry data three principal components were needed (55.73%). Ten substances were the most influential in the separation of the groups (young, intermediate, and old leaves or differentiation between genotypes A, B, and C), with emphasis on methyl digallate, quinic acid hexoside, digallic acid, and methyl gallate. Infrared spectroscopy data allowed us to differentiate young leaves from intermediate ones and the spectral bands that most influenced this differentiation (related to the groups O–H, C–O, CC, C–H and CO) correlated with the substances indicated by mass spectrometry. These results showed a different distribution of compounds among the leaf development stages, providing valuable information for those aiming to isolate specific substances of biological interest.

Genome-wide identification and expression analysis of two-component system genes in switchgrass (Panicum virgatum L.)

The two-component system (TCS) consists of histidine kinase (HK), histidine phosphate transfer protein (HP), and response regulatory factor (RR). It is one of the most crucial components of signal transduction in plants, playing a significant role in regulating plant growth, development, and responses to various abiotic stresses. Although TCS genes have been extensively identified in a variety of plants, the genome-wide recognition and examination of TCS in switchgrass remain unreported. Accordingly, this study identified a total of 87 TCS members in the genome of switchgrass, comprising 20 HK(L)s, 10 HPs, and 57 RRs. Detailed analyses were also conducted on their gene structures, conserved domains, and phylogenetic relationships. Moreover, this study analysed the gene expression profiles across diverse organs and investigated their response patterns to adverse environmental stresses. Results revealed that 87 TCS genes were distributed across 18 chromosomes, with uneven distribution. Expansion of these genes in switchgrass was achieved through both fragment and tandem duplication. PvTCS members are relatively conservative in the evolutionary process, but the gene structure varies significantly. Various cis-acting elements, varying in types and amounts, are present in the promoter region of PvTCSs, all related to plant growth, development, and abiotic stress, due to the TCS gene structure. Protein-protein interaction and microRNA prediction suggest complex interactions and transcriptional regulation among TCS members. Additionally, most TCS members are expressed in roots and stems, with some genes showing organ-specific expression at different stages of leaf and inflorescence development. Under conditions of abiotic stress such as drought, low temperature, high temperature, and salt stress, as well as exogenous abscisic acid (ABA), the expression of most TCS genes is either stimulated or inhibited. Our systematic analysis could offer insight into the characterization of the TCS genes, and further the growth of functional studies in switchgrass.

Roles of Germin-like Protein Family in Response to Seed Germination and Shoot Branching in Brassica napus.

Germin-like proteins (GLPs) play important roles in the regulation of various plant development processes, such as seed vigor, root and leaf development and disease resistance, while the roles of GLPs on agronomic traits are rarely studied in Brassica napus. Here, we identified GLPs family genes in rapeseed and analyzed their potential functions. There are 77 GLPs family genes (BnGLPs) in the Zhongshuang11 rapeseed reference genome, divided into a, b, c, d, e, f six subfamilies. Tissue expression profile analysis of BnGLPs revealed the following: e subfamily genes were highly expressed in early stages of silique, cotyledon, vegetative rosette and leaf development; f subfamily genes were highly expressed in seed development; genes of a subfamily were mainly expressed in the root; and genes of b, c, d subfamily exhibited low-level or no expression in above mentioned tissues. RT-qPCR analysis confirmed that the transcripts of two f subfamily members decreased dramatically during seed germination, suggesting that f subfamily proteins may play vital roles in the early stage of seed germination. Transcriptome analysis of axillary buds in sequential developing stages revealed that the transcripts of eight e subfamily genes showed a rapid increase at the beginning of shoot branching, implying that the e subfamily members played vital roles in branch development. These results demonstrate that rapeseed BnGLPs likely play essential roles in seedling development, root development and plant architecture, indicating that harnessing certain BnGLPs may contribute to the improvement of rapeseed yield.

Evaluating the Patterns of Maize Development in the Hetao Irrigation Region Using the Sentinel-1 GRD SAR Bipolar Descriptor.

Assessing maize yield is critical, as it is directly influenced by the crop's growth conditions. Therefore, real-time monitoring of maize growth is necessary. Regular monitoring of maize growth indicators is essential for optimizing irrigation management and evaluating agricultural yield. However, quantifying the physical aspects of regional crop development using time-series data is a challenging task. This research was conducted at the Dengkou Experimental Station in the Hetao irrigation area, Northwest China, to develop a monitoring tool for regional maize growth parameters. The tool aimed to establish a correlation between satellite-based physical data and actual crop growth on the ground. This study utilized dual-polarization Sentinel-1A GRD SAR data, accessible via the Google Earth Engine (GEE) cloud platform. Three polarization descriptors were introduced: θc (pseudo-scattering type parameter), Hc (pseudo-scattering entropy parameter), and mc (co-polar purity parameter). Using an unsupervised clustering framework, the maize-growing area was classified into several scattering mechanism groups, and the growth characteristics of the maize crop were analyzed. The results showed that throughout the maize development cycle, the parameters θc, Hc, and mc varied within the ranges of 26.82° to 42.13°, 0.48 to 0.89, and 0.32 to 0.85, respectively. During the leaf development stage, approximately 80% of the maize sampling points were concentrated in the low-to-moderate entropy scattering zone. As the plants reached the big trumpet stage, the entire cluster shifted to the high-entropy vegetation scattering zone. Finally, at maturity, over 60% of the sampling points were located in the high-entropy distribution scattering zone. This study presents an advanced analytical tool for crop management and yield estimation by utilizing precise and high-resolution spatial and temporal data on crop growth dynamics. The tool enhances the accuracy of crop growth management across different spatial and temporal conditions.

Transcriptome dynamics in developing leaves from C3 and C4 Flaveria species.

C4 species have evolved more than 60 times independently from C3 ancestors. This multiple and parallel evolution of the complex C4 trait suggests common underlying evolutionary mechanisms, which could be identified by comparative analysis of closely related C3 and C4 species. Efficient C4 function depends on a distinctive leaf anatomy that is characterised by enlarged, chloroplast-rich bundle sheath cells and narrow vein spacing. To elucidate the molecular mechanisms that generate the Kranz anatomy, we analysed a developmental series of leaves from the C4 plant Flaveria bidentis and the closely related C3 species Flaveria robusta by comparing anatomies and transcriptomes. Vascular density measurements of all nine leaf developmental stages identified three leaf anatomical zones whose proportions vary with respect to the developmental stage. We then deconvoluted the transcriptome datasets using non-negative matrix factorisation, which identified four distinct transcriptome patterns in the growing leaves of both species. By integrating the leaf anatomy and transcriptome data, we were able to correlate the different transcriptional profiles with different developmental zones in the leaves. These comparisons revealed an important role for auxin metabolism, in particular auxin homeostasis (conjugation and deconjugation), in establishing the high vein density typical of C4 species.

Developmental-specific regulation promotes the free amino acids accumulation in chlorotic tea plants (Camellia sinensis)

Chlorophyll-deficient tea plant exhibits a significantly higher accumulation of free amino acids (FAAs) than normal tea plants. This study focused on the impact of leaf color and the developmental stage on FAAs in six tea germplasms while maintaining all other conditions. The total FAAs content initially increased as the leaf matured during the one-bud-two-leaves (1B2L) and one-bud-three-leaves (1B3L) stages in green germplasms, then decreased or stabilized in the one-bud-four-leaves (1B4L) stage. In contrast, chlorotic germplasms showed continuous FAAs' content increase from 1B2L to 1B4L, thus being significantly positively correlated with total chlorophyll content. Interestingly, ethylamine content decreased with leaf maturation in both chlorotic and green germplasms, thus showing a significant negative correlation with L-theanine content only in chlorotic germplasms. Comparative RNA-seq analysis linked FAAs accumulation in chlorotic germplasm's 1B3L to photosynthesis inhibition and in 1B4L to nitrogen assimilation promotion. Feeding experiments revealed higher L-theanine synthesis and degradation abilities in chlorotic shoots versus green shoots, with synthesis efficiency exceeding degradation efficiency. Overall, this study uncovers a developmental-specific FAAs accumulation pattern in chlorotic germplasms and offers novel insights into the precise regulation by leaf color and developmental stage.

Genome-Wide Identification and Analysis of the WRKY Transcription Factor Family Associated with Leaf Senescence in Alfalfa

Leaves are the most significant parts of forage crops such as alfalfa. Senescence is the terminal stage of leaf development and is controlled by an integrated myriad of endogenous signals and environmental stimuli. WRKY transcription factors (TFs) play essential roles in regulating leaf senescence; however, only a few studies on the analysis and identification of the WRKY TF family in Medicago Sativa have been reported. In this study, we identified 198 WRKY family members from the alfalfa (M. sativa L.) cultivar ’XinjiangDaye’ using phylogenetic analysis and categorized them into three subfamilies, Groups I, II, and III, based on their structural characteristics. Group II members were further divided into five subclasses. In addition, several hormone- and stress-related cis-acting elements were identified in the promoter regions of MsWRKYs. Furthermore, 14 aging-related MsWRKYs genes from a previous transcriptome in our laboratory were selected for RT-qPCR validation of their expression patterns, and subsequently cloned for overexpression examination. Finally, MsWRKY5, MsWRKY66, MsWRKY92, and MsWRKY141 were confirmed to cause leaf yellowing in Nicotiana benthaminana using a transient expression system. Our findings lay a groundwork for further studies on the mechanism of M. sativa leaf aging and for the creation of new germplasm resources.

Assembly and maintenance of phyllosphere microbial diversity during rubber tree leaf senescence

Phyllosphere microorganisms execute important ecological functions including supporting host plant growth, enhancing host resistance to abiotic stresses, and promoting plant diversity. How leaf developmental stages affect plant-microbiome interactions and phyllosphere microbial community assembly and diversity is poorly understood. In this study, we utilized amplicon sequencing of 16S rRNA and ITS genes to investigate the composition and diversity of microbial communities across different leaf developmental stages of rubber trees. Our findings reveal that endophytic microbial communities, particularly bacterial communities, are more influenced by leaf senescence than by epiphytic communities. The high abundance of metabolism genes in the endosphere of yellow leaves contributes to the degradation and nutrient relocation processes. Nutrient loss leads to a higher abundance of α-Proteobacteria (r-selected microorganisms) in the yellow leaf endosphere, thereby promoting stochastic community assembly. As leaves age, the proportion of microorganisms entering the inner layer of leaves increases, consequently enhancing the diversity of microorganisms in the inner layer of leaves. These results offer insights into the mechanisms governing community assembly and diversity of leaf bacteria and fungi, thereby advancing our understanding of the evolving functions of microbial communities during leaf senescence in general, and for an important tropical crop species in particular.

Identification of transcription factors associated with leaf senescence in tobacco

Leaf senescence represents the final stage of leaf development, involving transcription factors (TFs)-mediated genetic reprogramming events. The timing of crop leaf senescence has a major influence on the yield and quality of crop in agricultural production. As important regulator of plant growth, the significance of TFs in the regulation of leaf senescence have been highlighted in various plant species by recent advances in genetics. However, studies on underlying molecular mechanisms are still not adequately explained. In this study, for analyzing the regulation of TFs on senescence of tobacco leaves, we combined gene differential expression analysis with weighted gene co-expression network analysis (WGCNA) to analyze the time-series gene expression profiles in senescing tobacco leaf. Among 3517 TF genes expressed in tobacco leaves, we identified 21, 35, and 183 TFs that were associated with early, middle, and late stages of tobacco leaf senescence, respectively. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation results reveal that these senescence response TFs are correlated with several biological pathways such as plant hormone signal transduction, ubiquitin mediated proteolysis and MAPK signaling pathway, indicating the roles of TFs in regulating leaf senescence. Our results provide implications for future studies of the potential regulatory mechanisms of TFs involved in senescence of tobacco leaves.