Brief Summary of Minutes of Annual Meeting

Accomplishments

CA (Khanday): Research at UC Davis focuses on understanding plant reproductive development and seed biology. The research enhances our understanding of seed development and germination to improve seed quality and vigor. This year, progress was made in identifying genetic and molecular mechanisms that regulate various seed quality traits. Key findings from this year include:

• A comprehensive developmental, physiological, and transcriptomic analysis of tomato seed development revealed key transition points associated with the acquisition of seed quality traits such as germinability, desiccation tolerance, reduced primary dormancy, and rapid germination. Several gene modules and candidate genes regulating these transitions during seed development were identified.
• To investigate the role of ABA biosynthesis in regulating seed dormancy and germination, expression of 9-cis-epoxycarotenoid dioxygenase (NCED) genes was examined. NCED enzymes regulate a key rate-limiting step in ABA biosynthesis pathway. Of the 11 NCED genes encoded in the tomato genome, five (SlNCED1, SlNCED2, SlNCED4, SlNCED5A, and SlNCED5B) were found to be expressed in seeds, each showing a distinct expression patterns during seed development.
• Loss-of-function mutants created in some of these seed-expressed NCED genes, demonstrated significantly enhanced vigor and stress-tolerance. These seeds show higher germination percentages, more rapid and synchronous germination compared to wild-type controls. They also show higher thermotolerance for germination than controls.

TX (Leskovar): Research at Texas A&M University, Texas A&M AgriLife Research

Objective 2: Capitalize on new technologies to assess and manipulate traits to enhance seedling quality

This year we focused on gaining more understanding on how selective strategies regulate or induce key morpho-physiological traits involved in seedling and grafted transplant adaptation. We also evaluated grafting as a tool to mitigate abiotic stress when exposed to different management strategies, under protected environments and organic systems.  We conducted four studies: 1) Pant biostimulants to enhance seedling and plant growth in seedless watermelon, 2) Role of microbial consortia and rootstocks for organic pepper production, 3) Heat stress responses of tomato genotypes across vegetative and reproductive stages, and 4) Grafted tomato rootstock screening under heat stress.

1) Plant biostimulants to enhance seedling and plant growth in seedless watermelon

The seedless watermelon study was conducted to evaluate microbial and seaweed extract biostimulants for their ability to enhance seed germination, seedling vigor, and canopy growth. A follow up trial evaluated seedlings treated with the same biostimulants at seeding, transplanting, and a day before moisture stress was initiated. Overall seaweed extracts significantly enhanced germination and emergence while no effect was observed for mycorrhizae treated seeds. Transplants treated with seaweed also had higher canopy and CO₂ assimilation under water deficit conditions.  These studies showed potential of microbial and seaweed biostimulant in seed germination, and enhanced seedling tolerance and growth in watermelon under moisture deficit stress.

2) Role of microbial consortia and rootstocks for organic pepper production

The grafted pepper study conducted in a high tunnel environment evaluated the performance of two Phytophthora-resistant rootstocks, CM-334 and YC-207, grafted with a common scion (Mama Mia Giallo), and the application of a mycorrhizal and bacterial mix consortium under two organic fertilization rates (optimal and reduced rate). The hypothesis explored whether the synergy of microbial inoculation and grafting could compensate for the reduced growth of the low rate by improving nutrient use efficiency.  Overall chlorophyll content was higher (~10 %) in inoculated plants compared to non-inoculated plants under low fertilization. Leaf net photosynthesis was similar at both fertilization rates, while inoculated plants had an increase in plant water use efficiency (~12 %). Microbial inoculation and low fertilization induced early flowering compared to non-inoculated plants grown at optimal fertilization rate. Under low fertilization, inoculated YC-grafted plants exhibited higher electron transport rate, stomatal conductance, and transpiration rate than other inoculated plants.  Our follow up studies will evaluate the correlation of these physiological traits with fruit yield, quality, and soil health to provide further understanding applicable to rootstock selection and microbial inoculation to optimize pepper production, particularly in resource-limited conditions.

3) Heat stress responses of tomato genotypes across vegetative and reproductive stages

We evaluated heat stress responses of ten tomato genotypes across the vegetative and reproductive stages over two seasons. Across all cultivars, ‘Big Beef’ showed larger declines in shoot and root biomass (74.1% and 70.4%), canopy leaf area (59.4%), stigma receptivity (82.1%), and pollen viability (60.3%) compared to the average genotype, but its pollen and ovule numbers (14.6% and 3.1%) were less reduced than the average. In contrast, ‘Cherokee Purple’ demonstrated similar thermotolerance levels during the vegetative and flowering stages to those of genotypes with tolerant yield responses. From that study we concluded that heat-tolerant or heat-sensitive responses in the vegetative or flowering stages did not always correlate with fruit yield reductions in tomato under heat stress.

4) Grafted tomato rootstock screening under heat stress  

This study assessed the productivity of novel rootstock-grafted tomato plants against the industry standard (Maxifort rootstock). A total of 27 scion/rootstock combinations, using two processing tomato industry scions (H1996 and H1015), and including non-grafted and self-grafted controls were compared under field conditions during summer 2025.  Based on yield performance and canopy growth, a set of four rootstocks (diploids) were selected for further studies.  Based on yield performance, best rootstock genotypes in combination were: X20-07 (1^st), X21-02 (3^rd), X20-06 (4^th), X21-09 (10^th). Genotypes that exhibited higher shoot biomass than non-grafted, self-grafted and Maxifort were also X20-07 and X21-09.  

FL ( Pérez): Coastal dune restoration programs are essential for protecting natural areas and human created infrastructure. Such programs rely on a mix of engineering and nature-based solutions to re-establish or stabilize dune systems. Dune stabilizing plants such as Uniola paniculata (sea oats) represent an essential component of nature-based solutions. However, the supply and quality of seeds from genetically appropriate source material can be problematic. Lack of information regarding factors associated with sea oats seed quality are lacking. This knowledge gap is magnified by the highly fragmented yet wide (> 1300 km) spatial distribution of sea oats along the US Atlantic and Gulf coasts.

Likewise, knowledge of environmental and management factors that influence wiregrass (Aristida beyrichiana) seed quality is lacking. Wiregrass is a foundational species due to its ability to carry regenerative fires and occurs in the critically endangered longleaf pine ecosystem throughout the southeastern US. Seed production is stimulated almost exclusively by growing season burns. Wiregrass is a key species used in long leaf pine ecosystem restoration to re-establish the vegetation-fire feedback system. But wiregrass seed quality is highly variable. The gap in wiregrass seed biology knowledge therefore limits restoration activities.

 Objective 1.1 Understand how developmental and environmental mechanisms affect seed quality – We worked with five state agencies to collects seeds of sea oats from 18 populations over a two-year period. We then investigated biochemical, physiological, and physical seed traits associated with seed quality and how these traits varied over a large latitudinal gradient.

We discovered that higher seed antioxidant concentrations were common in parts of the distribution with extreme environmental conditions. This correlates with sea oats populations growing at latitudinal extremes that experienced relatively warmer or colder temperatures. However, germination capacity was not coupled with baseline antioxidant concentrations. Higher antioxidant concentrations evident after imbibition were associated with poor seed germination in warmer conditions.

We also found that seed traits such as surface area and surface-to-volume ratio were positively correlated with aging stress tolerance. Similarly, the initial respiratory capacity and baseline antioxidant concentrations were positively correlated with aging stress survival. Alternatively, a spike in antioxidant concentrations at 36 hours after imbibition was negatively correlated with aging stress survival. We also ascertained that panicle density had the largest effects on seed production throughout the sea oats distribution.

Our results indicate that climate conditions and environmental components associated with temperature and precipitation may largely influence the initial pool of antioxidants in sea oats seeds. For example, a high concentration of antioxidants following seed imbibition could be indicative of seed damage or poor viability that may be influenced by environmental stress. Moreover, stable levels of antioxidants through the early imbibition process were typical of seeds with extended aging stress survival while unstable levels corresponded to reduced aging stress survival. Seed traits including surface-to-volume ratio and increased metabolism time were better positive predictors of extended aging stress survival.

We studied wiregrass seed quality in terms of seed fill and pathogen presence across 19 wiregrass populations throughout Florida. Seeds were collected from these sites over multiple seasons. We also studied factors that influence seedling emergence.

We found that wiregrass seeds while desiccation tolerant also expressed a short lifespan when confronted with ageing stress. We discovered that the type of habitat (e.g., xeric vs. mesic) and burn timing displayed the largest effects on the production of filled versus empty seeds. However, the type of habitat and rainfall amount during the seed developmental period were the strongest predictors of filled and infected seeds. Additionally, we uncovered that wiregrass seedling production was most strongly influenced by seasonal temperatures. Wiregrass seeds tend to avoid transitioning to seedlings during winter conditions. But there may be strong interannual differences in this response. The type of habitat where wiregrass occurred had a small effect on the seed to seedling transition.

KY (Finneseth): Joined group February 10, 2025. Established field plots of 60 native perennial species at the University of Kentucky Horticulture Research Farm (Lexington, KY) to explore seed production protocols and commercial feasibility. Preliminary data collected include mortality ratings, in-season performance ratings, and flowering interval. Seed harvest is underway and laboratory quality studies will be conducted over the winter.

KY (Kawashima): This year, we investigated how plants utilize actin filaments (F-actin) in the coenocytic endosperm to regulate final seed size, using Arabidopsis thaliana as a model. Our lab demonstrated that F-actin forms aster-shaped structures around each nucleus in the coenocytic endosperm, and that enhancing F-actin function by overexpressing actin genes increases seed size. Conversely, reducing F-actin function by expressing a dominant-negative form of actin results in smaller seeds. Through transcriptomic analyses, we identified a unique actin gene specifically expressed in the coenocytic endosperm. Interestingly, the mutant of this gene produces enlarged seeds, while its overexpression in the endosperm leads to smaller seeds. Biochemical assays further revealed that this actin isoform possesses distinct properties compared to conventional, ubiquitously expressed actins: faster polymerization and reduced bundling. We hypothesize that this endosperm-specific actin regulates nuclear movement and determines the number of nuclei retained in the endosperm, thereby controlling endosperm size and ultimately final seed size.

KY (Downie): The Natural Protection and repair mechanism in orthodox seeds encompasses a host of molecules and pathways put in place to permit survival of maturation desiccation and repair of essential entities following the reintroduction of water. In the first category are protective molecules including the LATE EMBRYOGENESIS ABUNDANT PROTEINs (LEAPs), intrinsically disordered proteins found in most organisms that can withstand desiccation at some stage of their life-cycle) that are hypothesized to provide protection to stressed cells. So too are non-reducing sugars (NRS; trehalose, sucrose, raffinose) which are thought to provide protection to membranes and proteins during desiccation tolerance as well as to mitigate the negative influence of reactive oxygen species. Following imbibition, one protein repair mechanism is the activity of PROTEIN ISOASPARTYL METHYLTRANSFERASE (PIMT) that acts to change isoAspartic acid (isoAsp, an abnormal amino acid formed non-enzymatically in proteins from either Asparagine [Asn] or Aspartic acid [Asp]) back to Asp. Together, these molecules are components of the natural protection and repair (NPR) mechanism in orthodox seeds, the focus of our studies.

We found that the pathway providing precursors to raffinose production also contributes to the vigor of Zea mays (maize) seeds during germination at sub-optimal temperatures (Li et al. 2025). Additionally, we ascertained that raffinose or one of its degradation products, galactose, contributes to flooding tolerance in maize (Yan et al. 2024). This is apparently through the stimulation of adventitious root formation close to the surface where oxygen availability in flooded soils is greater.

LEAPs have been thought to act as simple shield molecules, physically separating two otherwise reactive molecules from each other as water is removed from the system. The evidence for this is convincing but it does not illustrate the entire mode of action of the LEAPs. They, or at least some of them, have preferred client proteins to which they physically bind, including the formation of homodimers that can influence subcellular localization, or rapidly render the LEAPs innocuous upon the return of homeostasis to the cell. We have documented the preferred client proteins of one such LEAP in the dehydrin family (Unêda-Trevisoli, Dirk et al. 2024). Ms. Deepshikha Sai Bassetti, Ms. Ashley Morrison, and Mr. James Campbell, all University of Kentucky Agriculture and Medical Biotechnology students at the time, were co-authors on this publication (Unêda-Trevisoli, Dirk, et al. 2024.) We have published a review of one mechanism of action of some LEAPs in forming biomolecular condensates (Elder et al. 2025).

All proteins containing Asn or Asp (if not at the end of the peptide or immediately followed by proline) are susceptible to isoAsp formation and susceptibility (likelihood of forming isoAsp) is protein dependent. Upon dehydration the non-enzymatic, water independent, conversion of Asn and Asp to the intermediate succinimide is un-repairable. Succinimide in the stored proteome builds up over time. Upon imbibition the entire population of succinimide, unstable and highly reactive in water, converts to isoAsp (75%) or Asp (25%). The formation of isoAsp adds an additional carbon in the peptide chain of the protein misaligning important regulatory and catalytic domains. Fortunately, the protein repair PIMT recognizes isoAsp and, using S-Adenosyl methionine (SAM), methylates the alpha carbon, destabilizing it, which results in the conversion of isoAsp back to succinimide. The entire process repeats until the isoAsp is stoiciometrically converted back to Asp. Plants have two PIMTs. Previously, we have examined the prefered client protein repertoire of ZmPIMT2, unveiling a propensity to repair mitochondrial proteins without which the seed is of poor vigor. We have now unveiled the client proteins of ZmPIMT1 which includes many proteins involved in translation. Gratifyingly, client proteins identified using protein pull-down assays with ZmPIMT1 included a DEAD-box RNA HELICASE ortholog previously identified as a PIMT1 target in Arabidopsis! Additionally, many POLY-A BINDING PROTEINs (PABPs) were ZmPIMT1 targets. In addition to enhancing seed longevity, these PABPs are instrumental for alleviating seed dormancy in Arabidopsis (Zhang et al. 2025, in press). This article will be introduced in an In Brief summary.

SD (GU): 1. Completed a multi-year project to identify adaptive mechanisms of seed pigment traits. Seeds are coated with red or dark pigments presumably to promote plant adaptation. Allelic variants of the red (Rc/rc) and purple (Pb/pb) pericarp color genes were assembled into the same genetic background of rice. Genotypic differences in the developmental and histological patterns of pigment traits, seed flavonoid chemicals, seed dormancy, and germination velocity were detected. The biochemical and physiological data were used to model genetic component effects and genotype-by-environmental effects of the two genes.

OR (Sabry): Oregon State University (OSU) conducts studies in both basic and applied research, focusing on the application of new technologies to assess and manipulate traits to enhance seed quality. In 2025, our project addressed the following issues:

1. Development of digital PCR test to detect and quantify annual contamination in perennial ryegrass samples. This project is the fruit of collaboration with scientists from the USDA-ARS FRRL in Logan, UT and Oregon state University. The same research group developed KASP and qPCR tests to distinguish between annual and perennial ryegrass.
2. Determine the reliability of fluorescence and growout tests in detecting annual ryegrass in perennial samples.
3. Developing a short grow-out method to differentiate between annual and perennial ryegrass by using morphological traits of seedlings.
4. Elias collaborated with international researchers in two projects: 1) Enhancing salt tolerance in rye (Secale cereale) through ZnO and SiO2 nanoparticles and mycorrhizal fungi: Insights into antioxidant activation and physiological responses; and 2) Develop a novel approach in using insect-based spinach-food waste for gene targeting to cancer tissues.
5. Elias is also working on the 5^th ed. of the “Principles of Seed Science and Technology”.

The seed research at Oregon State University focuses on solving actual problems that are facing growers in Oregon and the US, answering their buzzling questions, increasing crop productivity and quality, and contributing to farmers welfare. For example, project 1 (above) aimed to solve the problem of contamination of annual ryegrass in perennial seed lots which has been one of the major problems in grass seed industry, especially in Oregon, the largest producer of ryegrass in the USA, and the 2^nd in the world. Project 2 (above) will help determine the reliability of the fluorescence test as a fast, economic test to detect annual ryegrass in perennial types. Project 3 (above) will provide US grass seed growers with an effective tool to differentiate between annual and perennial ryegrass when the fluorescence test over estimates the contamination of perennial ryegrass samples with annual types. Project 4 (above) aimed to explore methods for enhancing salt tolerance in rye (Secale cereale).

Collaborated with scientists from the USDA-ARS and from other universities help expand the usefulness of our studies beyond only one state (Oregon) of even one country (the US). Sabry is also a Co-PI in a project that has been submitted to NIFA on turfgrass genomics, with Texas A&M.

MI (Isaacs): Started a 3-year project to collaboratively build seed systems with midwestern Native American communities. Developed and distributed surveys to partner tribal communities to learn about their seed keeping practices, reasons for growing, and challenges with access to appropriate seed. Designed and facilitated 3 Native Seed Workshops with Winnebago, Santee, Ponca, and Omaha Nations: Creating A Shared Vision for Seed Saving, Seed Production, And Access to Native Foods (Total 60 participants). These workshops were in collaboration with Center for Rural Affairs and two branches of Nebraska Indian College, and Ponca Tribe of Nebraska, and Iowa State University. We determined future goals related to improving seed production and storage of native seed and identified seed training needs. At MSU, 30 students completed Isaacs’ course in Spring 2025, Grain Grading and Seed Quality. They learned the principles of seed quality and important factors for testing and regulating seed.