Overexpression of Rubisco subunits with RAF1 increases Rubisco content in maize

Abstract

Rubisco catalyzes a rate-limiting step in photosynthesis and has long been a target for improvement due to its slow turnover rate. An alternative to modifying catalytic properties of Rubisco is to increase its abundance within C 4 plant chloroplasts, which might increase activity and confer a higher carbon assimilation rate. Here, we overexpress the Rubisco large (LS) and small (SS) subunits with the Rubisco assembly chaperone RAF1. While overexpression of LS and/or SS had no discernable impact on Rubisco content, addition of RAF1 overexpression resulted in a >30% increase in Rubisco content. Gas exchange showed a 15% increase in CO 2 assimilation (A SAT ) in UBI-LSSS-RAF1 transgenic plants, which correlated with increased fresh weight and in vitro V cmax calculations. The divergence of Rubisco content and assimilation could be accounted for by the Rubisco activation state, which decreased up to 23%, suggesting that Rubisco activase may be limiting V cmax , and impinging on the realization of photosynthetic potential from increased Rubisco content.

Increased Rubisco content in maize mitigates chilling stress and speeds recovery

“…At light‐saturated photosynthetic capacity ( A SAT ), RAF1‐LSSS exhibited a significant, 12% increase in CO 2 assimilation under control conditions, and was 17% higher than the control after 2 weeks at 14 °C (Figure d‐e). This is in line with our previous observation that CO 2 assimilation increased in RAF1‐LSSS by 15% under control conditions, relative to WT plants (Salesse‐Smith et al , ). Compared to control conditions, however, photosynthetic rates decreased by approximately 50% in response to chilling in all genotypes (Figure e), consistent with published results (Long and Spence, ).…”

“…It should be considered, however, that the slope may additionally reflect carbonic anhydrase activities, as well as CO 2 diffusion between cells (mesophyll conductance and BS leakiness). Under control conditions, PEPC carboxylation efficiency was similar between all genotypes (Figure a and c), in keeping with our prior observation that all genotypes had similar PEPC activity (Salesse‐Smith et al , ). During chilling stress, carboxylation efficiency decreased significantly in all genotypes, except for RAF1‐LSSS where only a minimal decrease was observed (Figure b‐c).…”

“…Three previously reported transgenic lines were used in the experiments here: UBI‐RAF1, UBI‐LSSS and UBI‐RAF1‐LSSS (Salesse‐Smith et al , ; Wostrikoff et al , ). These lines express RAF1 and/or the Rubisco large and small subunits (LSSS), driven by the highly expressed maize ubiquitin promoter.…”

“…Thus, increasing Rubisco content in maize could theoretically provide a means to enhance chilling tolerance. Our previous work documented a >30% increase in maize Rubisco content conditioned by transgenic overexpression of the Rubisco large (LS) and small subunits (SS) with Rubisco Assembly Factor 1 (RAF1; Salesse‐Smith et al , ). The current study examines the properties of these transgenic lines during chilling and recovery to better understand Rubisco’s limitation on C 4 photosynthesis during chilling stress, and to ascertain whether increased Rubisco content confers a metabolic advantage under such conditions.…”

Photosynthesis engineered to increase rice yield

“…Furthermore, it is anticipated that increased Rubisco production could only be achieved at the expense of nitrogen use efficiency -with consequences for sustainability 11 . Recent analyses of chloroplast volume in C4 crops and a demonstrated transgenic increase in Rubisco content in maize have disproved the space limitation hypothesis for C4 crops 12,13 and, now, Yoon et al 3 disprove the hypothesis for C3 rice. Under well-fertilized conditions, the engineered plants showed almost double the flag leaf Rubisco content relative to the untransformed control plants.…”