Graduate Research Assistant University of Illinois Urbana-Champaign Urbana, Illinois
Body of Abstract: The rubisco activase (RCA) protein removes sugar phosphates from Ribulose-1,5-bisphosphate carboxylase-oxygenase (RUBISCO), keeping the enzyme active. In maize, two Rca genes encode for the α and β isoforms. The β isoform is encoded by Rca1 (Zm00001eb164390) and is constitutively expressed in leaves, while the α isoform is encoded by Rca3 (Zm00001eb164380) and is up-regulated during heat stress. While studies conducted in important C3 crops have shown the importance of the α isoform in heat stress response, the role of Rca3 in C4 maize is not yet understood. We present the characterization of Rca3 in maize using Dissociation (Ds) and Mutator (Mu) transposable element insertions. Mutant alleles were screened and verified using Sanger Sequencing with UniformMu insertion (rca3-m1::Mu) in the third exon, and the Ds insertion (rca3-m2::Ds) in the second exon. Results from RT-PCR indicate that the insertions disrupted Rca3 expression at 42°C. Gas exchange measurements at 42°C were used to identify variability between the response of the wildtype and mutant lines when heat stress is applied for four hours. Results showed that rca3-m2::Ds mutant plants have significantly lower photosynthetic rates after being exposed to 42°C for four hours. The rca3-m1::Mu allele had a less severe phenotype than rca3-m2::Ds allele, which we attribute to the position of the insertion that creates a knockdown rather than a knockout of Rca3. To check the relative expression of RNA in the wildtype and mutant lines at normal and high temperatures, qPCR was performed. Our results show that Rca3 is not expressed in rca3-m2::Ds mutants, while Rca3 is detected in rca3-m1::Mu plants, but with a reduced expression, which explains the less severe phenotype observed. Our results show that Rca3 in maize is important in maintaining photosynthetic rates at high temperatures and can be studied to improve crop resilience to temperature fluctuations in the future.