Effect of the WRKY76 transcription factor on starch biosynthesis and plant growth
Estabrooks, Hannah Margaret
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Starch is an important contributor to plant growth as excess photosynthate is stored in leaves as starch during the day to be mobilized at night and re-synthesized in sink tissues. Starch is also the principal constituent of cereal seeds and its variation greatly influence crop yields. The starch pathway is complex and its regulation is not fully understood. Transcription Factors (TFs) are known to act as master regulators of whole biosynthetic pathways and the work presented here was aimed at gaining a better understanding of starch production in leaves by identifying a TF which specifically regulates the leaf starch biosynthetic pathway. Leaf starch levels are regulated in part by ADP-glucose pyrophosphorylase (AGPase), the rate limiting step of starch biosynthesis. Transgenic rice event (NR16+) with increased leaf AGPase activity (due to overexpression of the AGPase large and small subunit transgenes Sh2r6hs and Bt2) was subjected to RNA-sequencing. The results indicated that the leaf specific AGP transgene, which increased leaf starch, also had upregulation of the WRKY76 TF. Another regulatory protein, F-Box, was chosen as a candidate due to the F-box family's involvement in plant development. The current study examines the potential of these gene products for increasing starch biosynthesis in leaves via leaf specific overexpression. Results indicate that overexpression of WRKY76 increases leaf starch an average of 39% at both the one month and anthesis growth stages in comparison to the Varietal Control Nipponbare. WRKY76 transgenic lines have enhanced phenotype with an improved harvest index due to biomass and immature panicles trending down by 4% and 21% respectively, while seed weight trended 12% higher. Events overexpressing WRKY76 were also found to upregulate important starch biosynthetic and carbon metabolism genes including AGPL1, AGPS2, SSIIIb, GBSII, Rbcs, PRK, and GS2 as well as leading to a general upregulation of leaf tissue carbohydrates. Events 1, 2, and 12 additionally had on average 13% increased photosynthetic rate at the one month growth stage. The findings of this study support WRKY76 as a dynamic regulator of the starch biosynthetic pathway conferring more efficient carbon assimilation leading to an increased harvest index.