SyPro POPLAR: IMPROVING POPLAR BIOMASS PRODUCTION UNDER ABIOTIC STRESS CONDITIONS: AN INTEGRATED OMICS, BIOINFORMATICS, SYNTHETIC BIOLOGY AND GENETIC ENGINEERING APPROACH.

The objective is to produce a suite of promoters that are short, precise and strong for induction. These synthetic promoters will be used to drive the expression of known and novel stress-responsive genes in trasnsgenic poplar trees under controlled and field conditions. Our objectives are: (1) Study the function of selected stress-responsive genes; (2) Discover novel motifs and construct stress-responsive synthetic promoters; (3) Use these promoters to drive the expression of genes known to confer abiotic stress tolerance in a variety of crops and develop abiotic-stress tolerant poplar seedlings; (4) Evaluate stress tolerance and yield potential of the transgenic poplar trees in the field. 

Collaborators:

  • Amir Ahkami, Pacific Northwest National Laboratory.
  • C. Neal Stewart, University of Tennessee.
  • Stephen DiFazio, West Virginia University.

THE EFFECTS OF MODIFICATIONS OF SOURCE-SINK RELATIONSHIPS ON CROP STRESS TOLERANCE

Abiotic stress is the primary cause of crop plant yield losses worldwide. Improving yield production and stability under stressful environments is needed to fulfill the food demand of the ever-growing world population, and an estimated increase of 50% in grain yield of major crops swill be needed by 2050. Drought, the most prominent threat to agricultural production worldwide, accelerates leaf senescence, leading to a decrease in canopy size, loss in photosynthesis and reduced yields. We hypothesized that it may be possible to enhance drought tolerance by altering sink/source relationships in the plant by promoting the stress-induced synthesis of cytokinins. The regulated expression of IPT (isopentenyltransferase) under the control of PSARK significantly improved drought tolerance in both laboratory and field conditions. Transgenic plants produced higher yields than wild-type plants in the field and the seeds from PSARK::IPTplants were normal, indicating that the nutritional value of the transgenic seeds was not altered. We used a multidisciplinary approach that combined genomics, proteomics, metabolomics and enzyme function analysis to identify and characterize cellular/biochemical components that regulate Carbon and Nitrogen metabolism in plants grown under water deficient conditions. Stress-induced cytokinin production had a positive effect on nitrate uptake as well as on the expression of genes associated with primary N assimilation and N re-assimilation, enhanced higher protein synthesis and the strengthening of the transgenic plants sink capacity. A System Biology approach was applied to identify genes and gene networks mediating the stress-response of crops to abiotic stress. A number of genes have been identified, and their expression has been modified in a number of crop species. The role of these genes and their effects on survival to stress is under study.

Collaborators:

  • Nir Sade, Tel Aviv University.

CELLULAR AND MOLECULAR DETERMINANTS FOR CLIMACTERIC AND NON-CLIMACTERIC BEHAVIOR IN PLUM FRUITS

Japanese plums represent the most abundant and variable group among tree species and include most of the fresh-market plums commercialized worldwide. We characterized and compared two Japanese plum cultivars, “Santa Rosa” (SR) and its bud-sport mutant “Sweet Miriam” (SM). These cultivars share the same genetic background but display contrasting ripening behaviors (SR, climacteric and SM, non-climacteric). Both cultivars differ in their sugar metabolism conferring the SM fruits with unusual quality properties (lower glucose and fructose, higher sorbitol and galactose-metabolism related sugars, etc). The main objective of this research is to characterize the differences in the complex sugar metabolic pathways between the cultivars and their possible crosstalk with ethylene biosynthesis-related enzymes, underlying their climacteric and non-climacteric fruit ripening behaviors. Fruits from each each cultivar were harvested at an early (S2: pit hardening) and late (S4: fully-ripe) stages of fruit development and assessed using a Systems Biology approach. Transcriptomics, proteomics and metabolomics methodologies, together with targeted gene expression and enzymatic activity assays were analyzed to reveal complex sugar metabolic interrelations and identify differences between the cultivars that could be associated to the observed changes in sugar homeostasis as well as ethylene biosynthesis and ethylene signaling. This experimental system provides a unique tool to study metabolic pathways underlying climacteric and non-climacteric fruit ripening behaviors and offers several practical applications. Understanding mechanisms that allow fruits to ‘switch’ to a sorbitol-based metabolism would have a great industry impact, since sorbitol is an alternative and healthier natural sweetener to sucrose. In addition, it could also allow the identification of candidate genes for breeding programs focused on fruit quality improvement.

Collaborators:

  • Macarena Farcuh, University of Maryland.

ENGINEERING NITROGEN FIXATION IN CEREAL CROPS

The interactions between plant roots and the microbe-rich soil environment are critical for plant fitness. It is estimated that plants exude up to 20% of their fixed carbon and many of these compounds (organic acids, amino acids, phenolics, secondary metabolites, etc.) help shaping microbial composition in the rhizosphere and rhizoplane, the region of soil containing the plant roots and the external root surface, respectively. Recently, we developed a novel approach in which rice plants were genetically modified to increase the production of compounds that stimulate the formation of biofilms in diazotrophic bacteria in the soil and promote the bacterial colonization of plant tissues, improving Biological Nitrogen Fixation (BNF) in a cereal crop (Yan et al., 2022). We performed a chemical screening to identify compounds that induce biofilm formation in diazotrophic bacteria and demonstrated that apigenin and other flavones induced bacterial biofilm synthesis, protecting the bacterial nitrogenase from damage by oxygen, and enhancing nitrogen-fixation activity. We genetically modified a flavone biosynthetic pathway in rice plants and generated enhanced flavone exudation into the rhizosphere, inducing BNF. As a consequence, modified rice plants produced more grain yield at limiting N-fertilizer.

We have identified several biosynthetic pathways in rice, wheat and other cereal crop plants that produce a number of metabolites that stimulate biofilm formation in soil diazotrophic bacteria and we are using CRISPR-based gene editing to modify the biosynthetic pathways in the plants, increasing compounds’ exudation into the rhizosphere with the concomitant increase in BNF and grain yields at reduced N-inorganic fertilizer concentrations.

Collaborators: Bayer Inc.