[O41] Genetic Improvement of Crop Yield

[O41-02] A Challenge for the Improvement of Photosynthetic Capacity by the Introduction of C₄-Like Rubisco in Rice

^○Hiroshi Fukayama¹, Keita Shiomi¹, Yuri Taketani¹, Hiroki Yoshikawa², Daisuke Sasayama¹, Tomoko Hatanaka¹, Tetsushi Azuma¹, Takuya Yoshizawa², Shun-Ichi Tanaka², Hiroyoshi Matsumura² (1.Graduate School of Agricultural Science, Kobe University, Japan, 2.Department of Biotechnology, Ritsumeikan University, Japan)

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is the key enzyme of photosynthetic CO₂ fixation. Rubisco in C₄ plants generally shows higher catalytic rate (k_cat) than that in C₃ plants. In our previous study, the chimeric incorporation of sorghum Rubisco small subunit (RbcS) significantly increased the Rubisco k_cat in rice. In this study, we knocked out rice RbcS multigene family by CRISPR/Cas9 and completely replaced the rice RbcS with sorghum RbcS in rice Rubisco. Obtained hybrid-Rubisco (CSS-Rubisco) showed almost C₄-plant-like catalytic properties, i.e., higher k_cat and K_m for CO₂. Transgenic lines expressing the hybrid-Rubisco accumulated moderately reduced levels of Rubisco and showed slight but significantly higher photosynthetic capacity at high CO₂ condition than non-transgenic rice. Grown under ambient CO₂ condition (40 Pa), the total dry weight in CSS lines was significantly lower than that in WT, whereas it was recovered to the level of WT under elevated CO₂ condition (100 and 300 Pa). The crystal structure of CSS-Rubisco in the sulfate-bound forms revealed a substantial structural difference in the β-hairpin (βC-βD) of RbcS around Leu101 in sorghum RbcS, which is likely to impact the flexibility of the 60s loop of Rubisco catalytic site. These results suggest that Leu101 in sorghum RbcS can be an important determinant of the kinetic properties of Rubisco and a promising target of improving the photosynthetic capacity in C₃ plants.