RUBISCO - THE CONSEQUENCES OF ALTERING ITS EXPRESSION AND ACTIVATION IN TRANSGENIC PLANTS

Transgenic tobacco (Nicotiana tabacum W38) hemizygous for a single antisense gene directed against Rubisco's small subunit had 35% of the Rubisco content of control leaves (15% when homozygous). CO2 assimilation (at 1000 mu mol quanta m(-2) s(-1) and 350 mu bar CO2) by the hemizygous leaves was reduced to 40% of that of the controls without material effect on stomatal conductance, chlorophyll content or other photosynthetic components. Leaf soluble protein was reduced commensurately with the reduction in Rubisco. CO2 assimilation rate in the hemizygous leaves remained limited by Rubisco activity at all, even very high, CO2 concentrations. This led to a simple, hyperbolic response of photosynthesis to intraplastid CO2 concentration from which the in vivo catalytic properties of Rubisco were inferred and compared with those of isolated Rubisco in vitro. Using a similar approach, the content of Rubisco activase was suppressed by incorporating a partial cDNA for activase into the tobacco genome in the antisense orientation with respect to a cauliflower mosaic virus 35S promoter. The progeny of a primary transformant with two anti-activase inserts had from <1% to 20% of the activase content of control plants. Quite severe suppression of activase, to less than 5% of the amount present in control leaves, was required before effects on photosynthesis and growth became apparent, indicating that one activase tetramer must be able to service, continuously, as many as 200 Rubisco octamers. Plants with lower activase contents could not grow unless the atmosphere was enriched with CO2. Their Rubisco was less carbamylated and they had lower CO2 assimilation rates than the controls. The rate of release of 2'-carboxyarabinitol-1-phosphate from Rubisco after illumination of the anti-activase leaves was also impaired. Older anti-activase plants accumulated increasing amounts of Rubisco in their younger leaves, but were unable to carbamylate it. The photosynthetic rate per carbamylated Rubisco active site in the strongly suppressed anti-activase leaves was only approximately 25% of that seen in control leaves, suggesting that activase may not only promote carbamylation of uncarbamylated Rubisco sites, but also accelerate turnover at carbamylated sites.