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8 recordsLinked to original sources
PubMed2026

A single-nucleus transcriptome atlas of soybean anthers.

Anther development is crucial for plant sexual reproduction. However, a high-resolution, cell-type-specific transcriptomic atlas of this process is lacking for the legume crop soybean (Glycine max). Here, we construct a comprehensive transcriptional atlas of developing soybean anthers using single-nucleus RNA sequencing (snRNA-seq). We identify and characterize nine distinct cell types spanning both somatic and reproductive lineages. Our analysis reveals robust transcriptional continuity across anther developmental stages and dynamic reprogramming during key transitions. Notably, the shift from diploid meiocytes to haploid unicellular microspores is marked by the induction of previously inactive genes, despite an overall reduction in transcript abundance. Subsequently, within bicellular microspores, generative and vegetative cell lineages exhibit sharply divergent transcriptional programs: generative cells specialize in mRNA export and turnover, whereas vegetative cells up-regulate translational machinery. Evolutionary analysis further indicates that generative-cell-specific genes are subject to more relaxed purifying selection compared to those specific to vegetative cells. Functional validation using mutants generated by CRISPR/Cas9-mediated genome editing and EMS mutagenesis reveals the essential roles of OSD1A and PKSA in pollen development and fertility. This high-resolution atlas provides fundamental insights into the transcriptional regulation of soybean anther development and serves as a valuable resource for manipulating male fertility to advance hybrid breeding programs. The data are available at https://databases.genedenovo.com/pollen.

Glycine max↗
PubMed2025

Genetic effect of the Ph1 locus on transcriptome atlas of anther development-related genes, meiotic chromosome behavior and agronomic traits in bread wheat.

Proper spatiotemporal expression of meiosis-related genes (MRGs) and other male-microsporogenesis/microgametogenesis-related genes (MMRGs) is crucial for normal anther development, yet their expression patterns remain largely unknown in wheat. The Ph1 locus in wheat is known to contain the Ph1 gene that plays a dual role in promoting pairing between homologous chromosomes but repressing pairing between homoeologous chromosomes, but its genetic function is still unclear. Here, we investigated these issues by conducting a comprehensive transcriptome analysis during wheat anther development in Chinese Spring (CS) and its ph1b deletion mutant under greenhouse and field conditions. Our results revealed that MRGs and MMRGs are predominantly expressed during pre-meiosis stages, with MMRGs also being highly expressed in meiotic-II. Gene co-expression analysis showed that C2H2 and B3 transcriptional factors (TFs) are associated with MRGs, and MYB regulators interacted mainly with MMRGs during microgametogenesis. Deletion of genes within the Ph1 locus failed to induce compensatory transcriptional activation of their homoeologous counterparts, while genes outside the Ph1 locus showed environmental-specific responses, especially during meiotic-II and mature pollen stages. Notably, early disjunction of bivalent chromosomes is a primary factor leading to defective meiocytes during metaphase I. Furthermore, the ph1b deletion mutant exhibited a substantially delayed heading date, potentially contributing to environment-stable and environment-specific alterations in fertility and grain-related traits. Our study highlights the significant impact of the Ph1 locus on the transcriptome during anther development, and a previously unheeded effect on meiotic chromosome pairing and agronomic traits, suggesting potential for genetic manipulations within the Ph1 locus for wheat improvement.

Triticum↗
PubMed2026

PAT: An Image Analysis Tool for Automated Scoring of Pollen in Alexander-Stained Anthers.

Quantitative pollen viability analysis is a critical but labor-intensive step in plant reproductive biology. Existing deep-learning Segment Anything Models (SAM) fail to reliably segment viable pollen in Alexander-stained anthers. To address this, we fine-tuned an existing Cellpose-SAM model for pollen segmentation. We integrated it into PAT (Pollen Analysis Tool), a cross-platform desktop application. PAT features instance segmentation with interactive quality control, an in-app model retraining module, and publication-ready statistical outputs. We deployed PAT in an EMS suppressor screen of semi-sterile Arabidopsis smg7-6 mutants, enabling efficient candidate prioritization for whole-genome sequencing and mapping of the candidate mutation. This screen led to the identification of a point mutation in CAP-D2 (capd2-2), a Condensin I subunit, that rescues the smg7-6 meiotic phenotype. Notably, mutation in a Condensin II subunits (CAP-D3 and CAP-H2) does not confer rescue. Further characterization suggests the capd2-2 allele is hypomorphic, showing no defects in vegetative growth, chromocenter compaction, or transposable element silencing. Collectively, we demonstrate that accessible AI tools have the potential to bridge gaps in plant phenotyping and accelerate the pace of biological discovery.

Alexander staining↗
PubMed2026

Circadian- and light-regulated oscillatory expression of CSA in rice leaves is required for pollen fertility.

The oscillatory expression of CSA in rice leaves is regulated by the circadian clock and red/far-red light signals, mediated through DOF5 and PIL11, and is required for normal pollen fertility. Photoperiod-sensitive male-sterile lines represent a pivotal innovation in the development of hybrid rice. However, the underlying mechanisms governing photoperiod-sensitive male reproductive development remain poorly understood. Our previous studies demonstrated that the carbon starved anther (csa) mutant exhibits male sterility under short-day (SD) conditions but partial fertility under long-day (LD) conditions. In this study, we report that CSA expression follows an oscillatory rhythm in rice leaves under both SD and LD conditions, a pattern regulated by both circadian clock and light signals. Tissue-specific RNA interference knockdown of CSA in leaves was associated with reduced pollen viability, suggesting that CSA expression in leaves contributes to normal male fertility. Promoter truncation assay results indicate that distinct regions of the CSA promoter contribute differentially to the regulation of CSA expression in leaves versus anthers, and that both the CSA expression level in anthers and the rhythmic expression pattern of CSA in leaves are associated with the restoration of male fertility. Using dual-luciferase, yeast one-hybrid, and electrophoretic mobility shift assays, we identified two proteins, PIL11 and DOF5, which directly bind to specific motifs (an E-box and T/AAAAG motif) within the CSA promoter truncation, thereby regulating its transcription. These findings elucidate novel mechanisms linking light sensing to the expression of circadian-controlled genes, thus connecting photoperiod with male reproductive development in rice.

Oryza↗
PubMed2024

Premeiotic 24-nt phasiRNAs are present in the Zea genus and unique in biogenesis mechanism and molecular function.

Reproductive phasiRNAs (phased, small interfering RNAs) are broadly present in angiosperms and play crucial roles in sustaining male fertility. While the premeiotic 21-nt (nucleotides) phasiRNAs and meiotic 24-nt phasiRNA pathways have been extensively studied in maize (Zea mays) and rice (Oryza sativa), a third putative category of reproductive phasiRNAs-named premeiotic 24-nt phasiRNAs-have recently been reported in barley (Hordeum vulgare) and wheat (Triticum aestivum). To determine whether premeiotic 24-nt phasiRNAs are also present in maize and related species and begin to characterize their biogenesis and function, we performed a comparative transcriptome and degradome analysis of premeiotic and meiotic anthers from five maize inbred lines and three teosinte species/subspecies. Our data indicate that a substantial subset of the 24-nt phasiRNA loci in maize and teosinte are already highly expressed at the premeiotic phase. The premeiotic 24-nt phasiRNAs are similar to meiotic 24-nt phasiRNAs in genomic origin and dependence on DCL5 (Dicer-like 5) for biogenesis, however, premeiotic 24-nt phasiRNAs are unique in that they are likely i) not triggered by microRNAs, ii) not loaded by AGO18 proteins, and iii) not capable of mediating PHAS precursor cleavage. In addition, we also observed a group of premeiotic 24-nt phasiRNAs in rice using previously published data. Together, our results indicate that the premeiotic 24-nt phasiRNAs constitute a unique class of reproductive phasiRNAs and are present more broadly in the grass family (Poaceae) than previously known.

Zea mays↗
PubMed2024

Premeiotic 24-nt phasiRNAs are present in the Zea genus and unique in biogenesis mechanism and molecular function.

Reproductive phasiRNAs are broadly present in angiosperms and play crucial roles in sustaining male fertility. While the premeiotic 21-nt phasiRNAs and meiotic 24-nt phasiRNA pathways have been extensively studied in maize (Zea mays) and rice (Oryza sativa), a third putative category of reproductive phasiRNAs-named premeiotic 24-nt phasiRNAs-have recently been reported in barley (Hordeum vulgare) and wheat (Triticum aestivum). To determine whether premeiotic 24-nt phasiRNAs are also present in maize and related species and begin to characterize their biogenesis and function, we performed a comparative transcriptome and degradome analysis of premeiotic and meiotic anthers from five maize inbred lines and three teosinte species/subspecies. Our data indicate that a substantial subset of the 24-nt phasiRNA loci in maize and teosinte are already highly expressed at premeiotic phase. The premeiotic 24-nt phasiRNAs are similar to meiotic 24-nt phasiRNAs in genomic origin and dependence on DCL5 for biogenesis, however, premeiotic 24-nt phasiRNAs are unique in that they are likely (i) not triggered by microRNAs, (ii) not loaded by AGO18 proteins, and (iii) not capable of mediating cis-cleavage. In addition, we also observed a group of premeiotic 24-nt phasiRNAs in rice using previously published data. Together, our results indicate that the premeiotic 24-nt phasiRNAs constitute a unique class of reproductive phasiRNAs and are present more broadly in the grass family (Poaceae) than previously known.

Maize↗
PubMed2026

Functional study of the AfRAP2 gene in Amorpha fruticosa L. tolerance to saline-alkali and drought stress.

BACKGROUND: Amorpha fruticosa L. is a leguminous shrub with high tolerance to drought, poor soil, and saline-alkali stress conditions. As a member of the family of transcription factors in higher plants, the ethylene response factor AP2/ERF plays a crucial role in both plant adaptation to abiotic stress and in growth and development. In this study, based on genes identified from the transcriptomic sequencing of Amorpha fruticosa L. under drought stress, the upregulated gene AfRAP2 was isolated from its seedlings, with the aim of elucidating its stress-response function using molecular biological techniques. RESULTS: In this study, the AfRAP2 gene was cloned from the leaves of Amorpha fruticosa L. using RT-PCR. Bioinformatics analysis revealed that AfRAP2 contains an AP2 domain and belongs to the DREB subfamily of the AP2/ERF transcription factor family, showing close phylogenetic relationships with LaEREBP from Lathyrus albus. Real-time quantitative PCR (RT-qPCR) results indicate that AfRAP2 is expressed in various tissues of Amorpha fruticosa L., with the highest expression in leaves and the lowest in stems, furthermore, its expression is significantly upregulated in roots and leaves upon induction by NaHCO3 and PEG6000. Subcellular localization experiments confirmed that the AfRAP2 protein is localized to the nucleus, and GUS histochemical staining assay revealed that its promoter drives GUS expression in anthers. Resistance analysis of overexpressing yeast strains showed that yeast transformed with the AfRAP2 gene exhibited significantly better growth under sorbitol, mannitol, and NaHCO3 stress conditions compared to the control, indicating that this gene enhances yeast tolerance to drought and saline-alkali stress. We screened transgenic tobacco and Populus davidiana × P. alba var. Pyramidalis. The results showed that under natural drought and saline-alkali stress treatments, the transgenic lines exhibited significantly improved growth and higher activities of the physiological indicators of catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD) compared with wild-type plants, indicating that the overexpression of the AfRAP2 gene plays a key role in the response to saline-alkali stress and drought stress. CONCLUSION: In summary, AfRAP2 contains an AP2 domain and belongs to the DREB subfamily of transcription factors, under abiotic stress induced by NaHCO₃ and mannitol, it can induce the expression of the AfRAP2 gene in tobacco and Populus davidiana×P. alba var. pyramidalis. AfRAP2 plays a vital role in the plant response to saline-alkali stress and drought stress and is a promising candidate gene for stress-tolerant breeding.

Plant Proteins↗
PubMed2026

Heat stress impact on rice reproductive processes: challenges and new approaches.

Heat stress represents one of the most severe abiotic constraints to rice (Oryza sativa L.) productivity and is expected to intensify under ongoing climate change, particularly affecting the reproductive phase and leading to substantial yield and grain quality losses. This review synthesizes current knowledge on the impacts of heat stress on rice reproduction, with a focus on both male and female reproductive structures and their interactions. Evidence from anatomical, physiological, transcriptomic, and metabolomic studies to describe how elevated temperatures disrupt key reproductive processes, including microsporogenesis, anther dehiscence, pollen viability, pollen-pistil interactions, fertilisation, and embryo sac development were integrated in this review. It further discusses the genotype-dependent differences in reproductive thermotolerance; and key genes, metabolites, and pathways associated with heat stress perception, signalling, and tolerance are highlighted. Finally, it is briefly discussed how recent advances in breeding strategies, functional genomics and genome-editing technologies, particularly CRISPR-based approaches, are providing new opportunities to enhance reproductive resilience to heat stress and how it is essential to close the existing molecular knowledge gaps in the development of heat-tolerant rice varieties capable of sustaining productivity in a warming climate.

Oryza sativa (L.)↗