OBJECTIVE: Rock outcrop vegetation is distributed worldwide and hosts a diverse and unique flora that evolved under harsh environmental conditions. Unfortunately, seed ecology in such ecosystems has received little attention, especially regarding seed traits, germination responses to abiotic factors and the potential role of phylogenetic relatedness on such features Here, we provide the first quantitative and phylogenetically-informed synthesis of the seed functional ecology of Brazilian rock outcrop vegetation, with a particular focus on quartzitic and ironstone campo rupestre.
METHODS: Using a database of functional trait data, we calculated the phylogenetic signal of seven seed traits for 371 taxa and tested whether they varied among growth forms, geographic distribution, and microhabitats. We also conducted meta-analyses that included 4,252 germination records for 102 taxa to assess the effects of light, temperature, and fire-related cues on the germination of campo rupestre species and explored how the aforementioned ecological groups and seed traits modulate germination responses.
RESULTS: All traits and germination responses showed a moderate-to-strong phylogenetic signal. Campo rupestre species responded positively to light and had maximum germination between 20-25 ºC. The effect of temperatures beyond this range was moderated by growth form, species geographic distribution, and microhabitat. Seeds exposed to heat shocks above 80 °C lost viability, but smoke accelerated germination. We found a moderating effect of seed mass for in responses to light and heat shocks, with larger, dormant seeds tolerating heat better but less sensitive to light. Species from xeric habitats evolved phenological strategies to synchronise germination during periods of increased soil water availability.
CONCLUSIONS: Phylogenetic relatedness plays a major role in shaping seed ecology of Brazilian rock outcrop vegetation. Nevertheless, seed traits and germination responses varied significantly between growth forms, species geographic distribution and microhabitats, providing support to the regeneration niche hypothesis and the role of functional traits in shaping germination in these ecosystems.

OBJECTIVE: Epimedium brevicornu Maxim. is a perennial persistent C3 plant of the genus Epimedium Linn. in the family Berberaceae that exhibits severe physiological and morphological seed dormancy.We placed mature E. brevicornu seeds under nine stratification treatment conditions and explored the mechanisms of influence by combining seed embryo growth status assessment with related metabolic pathways and gene co-expression analysis.
RESULTS: We identified 3.9 °C as the optimum cold-stratification temperature of E. brevicornu seeds via a chilling unit (CU) model. The best treatment was variable-temperature stratification (10/20 °C, 12/12 h) for 4 months followed by low-temperature stratification (4 °C) for 3 months (4-3). A total of 63801 differentially expressed genes were annotated to 2587 transcription factors (TFs) in 17 clusters in nine treatments (0-0, 0-3, 1-3, 2-3, 3-3, 4-3, 4-2, 4-1, 4-0). Genes specifically highly expressed in the dormancy release treatment group were significantly enriched in embryo development ending in seed dormancy and fatty acid degradation, indicating the importance of these two processes. Coexpression analysis implied that the TF GRF had the most reciprocal relationships with genes, and multiple interactions centred on zf-HD and YABBY as well as on MYB, GRF, and TCP were observed.
CONCLUSIONS: In this study, analyses of plant hormone signal pathways and fatty acid degradation pathways revealed changes in key genes during the dormancy release of E. brevicornu seeds, providing evidence for the filtering of E. brevicornu seed dormancy-related genes.

Preharvest sprouting (PHS) is a serious problem in rice production as it leads to reductions in grain yield and quality. However, the underlying mechanism of PHS in rice remains unclear. In this study, we identified and characterized a preharvest sprouting and seedling lethal (phssl) mutant. The heterozygous phssl/+ mutant exhibited normal plant development, but severe PHS in paddy fields. However, the homozygous phssl mutant was seedling lethal. Gene cloning and genetic analysis revealed that a point mutation in OsABA3 was responsible for the mutant phenotypes. OsABA3 encodes a molybdenum cofactor (Moco) sulfurase. The activities of the sulfureted Moco-dependent enzymes such as aldehyde oxidase (AO) and xanthine dehydrogenase (XDH) were barely detectable in the phssl mutant. As the final step of abscisic acid (ABA) de novo biosynthesis is catalyzed by AO, it indicated that ABA biosynthesis was interrupted in the phssl mutant. Exogenous application of ABA almost recovered seed dormancy of the phssl mutant. The knock-out (ko) mutants of OsABA3 generated by CRISPR-Cas9 assay, were also seedling lethal, and the heterozygous mutants were similar to the phssl/+ mutant showing reduced seed dormancy and severe PHS in paddy fields. In contrast, the OsABA3 overexpressing (OE) plants displayed a significant increase in seed dormancy and enhanced plant resistance to PHS. The AO and XDH activities were abolished in the ko mutants, whereas they were increased in the OE plants. Notably, the Moco-dependent enzymes including nitrate reductase (NR) and sulfite oxidase (SO) showed reduced activities in the OE plants. Moreover, the OE plants exhibited enhanced resistances to osmotic stress and bacterial blight, and flowered earlier without any reduction in grain yield. Taken together, this study uncovered the crucial functions of OsABA3 in Moco sulfuration, plant development, and stress resistance, and suggested that OsABA3 is a promising target gene for rice breeding.

Vivipary is a prominent feature of mangroves, allowing seeds to complete germination while attached to the mother plant, and equips propagules to endure and flourish in challenging coastal intertidal wetlands. However, vivipary-associated genetic mechanisms remain largely elusive. Genomes of two viviparous mangrove species and a non-viviparous inland relative were sequenced and assembled at the chromosome level. Comparative genomic analyses between viviparous and non-viviparous genomes revealed that DELAY OF GERMINATION 1 (DOG1) family genes (DFGs), the proteins from which are crucial for seed dormancy, germination, and reserve accumulation, are either lost or dysfunctional in the entire lineage of true viviparous mangroves but are present and functional in their inland, non-viviparous relatives. Transcriptome dynamics at key stages of vivipary further highlighted the roles of phytohormonal homeostasis, proteins stored in mature seeds, and proanthocyanidins in vivipary under conditions lacking DFGs. Population genomic analyses elucidate dynamics of syntenic regions surrounding the missing DFGs. Our findings demonstrated the genetic foundation of constitutive vivipary in Rhizophoraceae mangroves.

In many regions, the climate is changing faster during winter than during the other seasons, and a loss of snow cover combined with increased temperature variability can expose overwintering organisms to harmful conditions. Understanding how species respond to these changes during critical developmental times, such as seed germination, helps us assess the ecological implications of winter climate change. To address this concern, we measured the breaking of seed dormancy and cold tolerance of temperate grassland species in the lab and field. In the lab, we ran germination trials testing the tolerance of 17 species to an extreme cold event. In the field, we deployed seeds of two species within a snow manipulation experiment at three locations and measured germination success biweekly from seeds subjected to ambient and reduced snow cover from winter into spring. From lab trials, cold tolerance varied among species, with seed germination decreasing <10%-100% following extreme cold events. Cold tolerance was related to seed traits, specifically less round seeds, seeds that required cold stratification, and seeds that mature later in the season tended to be more impacted by extreme cold temperatures. This variation in seed cold tolerance may contribute to altered community composition with continued winter climate change. In the field, germination increased through late winter, coinciding with the accumulation of days where temperatures were favorable for cold stratification. Through spring, germination success decreased as warm temperatures accumulated. Collectively, species-specific seed cold tolerances and mortality rates may contribute to compositional changes in grasslands under continued winter climate change.

Different methodologies have been applied for the selection of preharvest sprouting resistance in cereal breeding programs. We describe here a series of methods used in practical wheat breeding programs in Japan, including phenotyping based on germination score after artificial rain treatments and genotyping using DNA markers. These methods can be modified and applied to breeding programs in which preharvest sprouting is a problem during cereal cultivation.

Genome-wide association study (GWAS) is widely used to characterize genes or quantitative trait loci (QTLs) associated with preharvest sprouting and seed dormancy. GWAS can identify both previously discovered and novel QTLs across diverse genetic panels. The high-throughput SNP arrays or next-generation sequencing technologies have facilitated the identification of numerous genetic markers, thereby significantly enhancing the resolution of GWAS. Although various methods have been developed, the fundamental principles underlying these techniques remain constant. Here, we provide a basic technological flow to perform seed dormancy assay, followed by GWAS using population structure control, and compared it with previous identified QTLs and genes.

Seed dormancy is an important agronomic trait in cereal crops. Throughout the domestication of cereals, seed dormancy has been reduced to obtain uniform germination. However, grain crops must retain moderate levels of seed dormancy to prevent problems such as preharvest sprouting in wheat (Triticum aestivum) and barley (Hordeum vulgare). To produce modern cultivars with the appropriate seed dormancy levels, it is important to identify the genes responsible for seed dormancy. With recent advances in sequencing technology, several causal genes for seed dormancy quantitative trait loci (QTLs) have been identified in barley and wheat. Here, we present a method to identify causal genes for seed dormancy QTLs in barley, a method that is also applicable to other cereals.

Seed dormancy is an important trait in cereal breeding, as it prevents preharvest sprouting (PHS). Although seed dormancy is a multifactorial trait, seed color has been demonstrated to be a major dormancy-related factor controlled by few genes. The R-1 gene is a seed color regulator that encodes a MYB-type transcription factor in wheat. A set of genetic markers designed against R-1 can provide a powerful tool for swift wheat breeding. Depth of seed dormancy varies not only among lines but also during seed development in each line. In this chapter, we describe how developmental seeds can be collected to perform germination tests, how seed color can be observed after NaOH staining, and how to genotype wheat R-1 genes using multiplex PCR.

Seed germination is a critical phase for the life cycle and propagation of higher plants. This study explores the role of SlWRKY37, a WRKY transcription factor in tomato, in modulating seed germination. We discovered that SlWRKY37 expression is markedly downregulated during tomato seed germination. Through CRISPR/Cas9-mediated editing, we demonstrate that SlWRKY37 knockout enhances germination, while its overexpression results in a delay compared to the wild type. Transcriptome analysis revealed 679 up-regulated and 627 down-regulated genes in Slwrky37-CRISPR deletion mutants relative to the wild type. Gene ontology (GO) enrichment analysis indicated these differentially expressed genes are linked to seed dormancy, abscisic acid homeostasis, and protein phosphorylation pathways. Bioinformatics and biochemical assays identified SlABI5-like7 and SlLEA2 as key transcriptional targets of SlWRKY37, integral to tomato seed dormancy regulation. Additionally, SlWRKY37 was found to be post-translationally phosphorylated at Ser65, a modification crucial for its transcriptional activation. Our findings elucidate the regulatory role of SlWRKY37 in seed dormancy, suggesting its potential as a target for gene editing to reduce seed dormancy in tomato breeding programs.