Characterization of molecular functions of polyamines in fruit development and ripening in tomato (Solanum lycopersicum)
Date of Award
Doctor of Philosophy (PhD)
Horticulture and Landscape Architecture
Avtar K Handa
Committee Member 1
Michael V Mickelbart
Committee Member 2
Kashchandra G Raghothama
Committee Member 3
Putrescine (PUT), spermidine (SPD) and spermine (SPM) are three major polyamines (PAs) present in all living organisms. These biogenic amines have been implicated in diverse plant growth and development processes, including seed germination, tissue lignification, organogenesis, flowering, pollination, embryogenesis, fruit development, ripening, abscission, senescence, and stress responses. To elucidate molecular roles of PAs in fruit development and ripening, I characterized transgenic tomato plants ectopically expressing yeast spermidine synthase (ySpdSyn) or S-adenosylmethionine decarboxylase (ySAMdc) under constitutive CaMV 35S and/or fruit-specific SlE8 promoters. The ySpdSyn-expression enhanced PUT, SPD and SPM level in floral buds and fertilized developing ovaries by 2- to 3-fold compared to WT tissues with majority being sequestered as bound forms. Higher PA levels altered fruit shape of transgenic tomatoes to more obovoid than WT by regulating expression of fruit shape genes (SUN1 and OVATE), and cell division and expansion genes (CDKB2, CYCB2, KRP1 and CCS52B). Characterization of PA homeostasis during fruit growth and ripening revealed a strong correlation of conjugated PAs with transcripts abundance of PA biosynthesis (ODC, ADC, SAMdc3) and catabolizing genes (CuAO-like, PAO4-like) and the bound PAs to transcript levels of ySpdSyn and SAMdc2 suggesting a significant metabolic inter-conversion among the various forms of PAs. Co-expression of ySpdSyn and ySAMdc transgenes showed that SAMdc is the rate limiting step in biosynthesis of higher PAs with potential to alter PA homeostasis in fruit tissues. Characterization of ySpdSyn and ySAMdc transgenic and WT fruits showed that expression of transgenes was associated with higher firmness of ripened fruits both on-planta and after harvest up to 17 days after ripe stage. Free SPD/SPM levels were positively correlated with fruit firmness, accumulation of total solids and delay in fruit shriveling and inversely correlated with fresh fruit weight, juice pH and seed number in tomato fruits. Free PUT levels exhibited trends opposite to that seen with SPD/SPM confirming hypothesis that PUT and SPD/SPM ratios play significant roles in the outcome of biological functions of PAs. Evaluation of ySpdSyn lines under field conditions showed 50% increase in fruit yield per plant due to continued fruit set until late in the season and up to 60% increase in fruit fresh and dry weight much beyond the fruit breaker stage. The metabolomic changes in transgenic fruits were determined using the nuclear magnetic resonance spectroscopy (1H NMR) and compared to WT fruit metabolic profile during on-planta fruit ripening and post-ripening stages. Free SPD levels were positively correlated with Ile, Val, Glu, Gln, Trp, malate, citrate and trigonelline. The levels of Ala, Glu, Asp and UDP-NAcGLU were negatively correlated with free SPD levels but positively correlated with free PUT indicating differential function of these two PAs. Levels of fructose and AMP were also negatively correlated with free SPD. Conjugated and bound PAs exhibited a limited correlation with metabolome profiles. The node-edge network analyses among PAs, metabolites and their associated pathways showed that PAs upregulate many anabolic pathways, but negatively affect glycolysis, starch and sugar metabolism, and zeatin biosynthesis. Taken together these results indicate that SPD is associated with enhancing many metabolic pathways and delaying senescence-related processes leading to improved postharvest fruit quality. I have collated transcriptome of transgenic plants and mutants with altered PA levels. Its analyses revealed complex and differential relationships among PUT, SPD and SPM in regard to regulation of plant hormone biosynthesis and signaling. In summary, the use of transgenic plants with modified PA levels provide an insight into molecular functions of PAs in altering fruit architecture, improving fruit quality attributes, increasing fruit production and delaying ripening-related changes in tomatoes. Limited transcriptome profile suggest a complex crosstalk between PAs and plant growth hormones during fruit ripening. Metabolome profiles of transgenic fruits showed a significant impact of PAs on fruit quality improvement by restoring metabolic pathways during fruit ripening.
Anwar, Raheel, "Characterization of molecular functions of polyamines in fruit development and ripening in tomato (Solanum lycopersicum)" (2014). Open Access Dissertations. 1058.