Seed coat lignification level is crucial in Capsicum spp seed longevity.

Capsicum (pepper) is known for its poor seed germination, particularly seed longevity is usually much shorter than other Solanaceae. However, the molecular mechanisms involved are mostly unknown in these species. The present study examines the differences in seed longevity among Capsicum species and varietal types. Feral or less domesticated species, such as Capsicum chinense and particularly Capsicum frutescens, showed higher germination rates than the more domesticated Capsicum annuum after accelerated seed aging treatments. In addition, variability was detected in the expression of genes involved in the response to seed deterioration. The differences observed in ASPG1 expression led us to study the seed protein profile in dry and germinating seeds. Seed storage protein mobilization during germination was faster in seed aging-resistant genotypes. Similarly, the transcriptional change observed for the orthologous gene of the trans-species regulator AtHB25 prompted us to study the structure and molecular components of the seed coat in peppers. All the Capsicum pepper accessions analyzed presented very lignified testa and we observed a positive correlation between the amount of lignin and seed viability. Our results provide essential information to explain the poor germination observed in pepper seeds and provide an experimental framework for future improvements in this important character.

Apoplastic lipid barriers regulated by conserved homeobox transcription factors extend seed longevity in multiple plant species.

Cutin and suberin are lipid polyesters deposited in specific apoplastic compartments. Their fundamental roles in plant biology include controlling the movement of gases, water and solutes, and conferring pathogen resistance. Both cutin and suberin have been shown to be present in the Arabidopsis seed coat where they regulate seed dormancy and longevity. In this study, we use accelerated and natural ageing seed assays, glutathione redox potential measures, optical and transmission electron microscopy and gas chromatography-mass spectrometry to demonstrate that increasing the accumulation of lipid polyesters in the seed coat is the mechanism by which the AtHB25 transcription factor regulates seed permeability and longevity. Chromatin immunoprecipitation during seed maturation revealed that the lipid polyester biosynthetic gene long-chain acyl-CoA synthetase 2 (LACS2) is a direct AtHB25 binding target. Gene transfer of this transcription factor to wheat and tomato demonstrated the importance of apoplastic lipid polyesters for the maintenance of seed viability. Our work establishes AtHB25 as a trans-species regulator of seed longevity and has identified the deposition of apoplastic lipid barriers as a key parameter to improve seed longevity in multiple plant species.

Identification of novel seed longevity genes related to oxidative stress and seed coat by genome-wide association studies and reverse genetics.

Seed longevity is a polygenic trait of relevance for agriculture and for understanding the effect of environment on the ageing of biological systems. In order to identify novel longevity genes, we have phenotyped the natural variation of 270 ecotypes of the model plant, Arabidopsis thaliana, for natural ageing and for three accelerated ageing methods. Genome-wide analysis, using publicly available single-nucleotide polymorphisms (SNPs) data sets, identified multiple genomic regions associated with variation in seed longevity. Reverse genetics of 20 candidate genes in Columbia ecotype resulted in seven genes positive for seed longevity (PSAD1, SSLEA, SSTPR, DHAR1, CYP86A8, MYB47 and SPCH) and five negative ones (RBOHD, RBOHE, RBOHF, KNAT7 and SEP3). In this uniform genetic background, natural and accelerated ageing methods provided similar results for seed-longevity in knock-out mutants. The NADPH oxidases (RBOHs), the dehydroascorbate reductase (DHAR1) and the photosystem I subunit (PSAD1) highlight the important role of oxidative stress on seed ageing. The cytochrome P-450 hydroxylase, CYP86A8, and the transcription factors, MYB47, KNAT7 and SEP3, support the protecting role of the seed coat during seed ageing.

PRX2 and PRX25, peroxidases regulated by COG1, are involved in seed longevity in Arabidopsis.

Permeability is a crucial trait that affects seed longevity and is regulated by different polymers including proanthocyanidins, suberin, cutin and lignin located in the seed coat. By testing mutants in suberin transport and biosynthesis, we demonstrate the importance of this biopolymer to cope with seed deterioration. Transcriptomic analysis of cog1-2D, a gain-of-function mutant with increased seed longevity, revealed the upregulation of several peroxidase genes. Reverse genetics analysing seed longevity uncovered redundancy within the seed coat peroxidase gene family; however, after controlled deterioration treatment, seeds from the prx2 prx25 double and prx2 prx25 prx71 triple mutant plants presented lower germination than wild-type plants. Transmission electron microscopy analysis of the seed coat of these mutants showed a thinner palisade layer, but no changes were observed in proanthocyanidin accumulation or in the cuticle layer. Spectrophotometric quantification of acetyl bromide-soluble lignin components indicated changes in the amount of total polyphenolics derived from suberin and/or lignin in the mutant seeds. Finally, the increased seed coat permeability to tetrazolium salts observed in the prx2 prx25 and prx2 prx25 prx71 mutant lines suggested that the lower permeability of the seed coats caused by altered polyphenolics is likely to be the main reason explaining their reduced seed longevity.

Arabidopsis COGWHEEL1 links light perception and gibberellins with seed tolerance to deterioration.

Light is a major regulator of plant growth and development by antagonizing gibberellins (GA), and we provide evidence for a role of light perception and GA in seed coat formation and seed tolerance to deterioration. We have identified two activation-tagging mutants of Arabidopsis thaliana, cog1-2D and cdf4-1D, with improved seed tolerance to deterioration linked to increased expression of COG1/DOF1.5 and CDF4/DOF2.3, respectively. These encode two homologous DOF transcription factors, with COG1 most highly expressed in seeds. Improved tolerance to seed deterioration was reproduced in transgenic plants overexpressing these genes, and loss of function from RNA interference resulted in opposite phenotypes. Overexpressions of COG1 and CDF4 have been described to attenuate various light responses mediated by phytochromes. Accordingly, we found that phyA and phyB mutants exhibit increased seed tolerance to deterioration. The phenotype of tolerance to deterioration conferred by gain of function of COG1 and by loss of function of phytochromes is of maternal origin, is also observed under natural aging conditions and correlates with a seed coat with increased suberin and reduced permeability. In developing siliques of the cog1-2D mutant the expression of the GA biosynthetic gene GA3OX3 and levels of GA1 are higher than in the wild type. These results explain the antagonism between phytochromes and COG1 in terms of the inhibition and the activation, respectively, of GA action.