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

Transgenic tobacco (Nicotiana tabacum W38) hemizygous for a single ant isense gene directed against Rubisco's small subunit had 35% of the Ru bisco content of control leaves (15% when homozygous). CO2 assimilatio n (at 1000 mu mol quanta m(-2) s(-1) and 350 mu bar CO2) by the hemizy gous leaves was reduced to 40% of that of the controls without materia l effect on stomatal conductance, chlorophyll content or other photosy nthetic components. Leaf soluble protein was reduced commensurately wi th the reduction in Rubisco. CO2 assimilation rate in the hemizygous l eaves remained limited by Rubisco activity at all, even very high, CO2 concentrations. This led to a simple, hyperbolic response of photosyn thesis to intraplastid CO2 concentration from which the in vivo cataly tic properties of Rubisco were inferred and compared with those of iso lated Rubisco in vitro. Using a similar approach, the content of Rubis co activase was suppressed by incorporating a partial cDNA for activas e into the tobacco genome in the antisense orientation with respect to a cauliflower mosaic virus 35S promoter. The progeny of a primary tra nsformant with two anti-activase inserts had from <1% to 20% of the ac tivase content of control plants. Quite severe suppression of activase , to less than 5% of the amount present in control leaves, was require d before effects on photosynthesis and growth became apparent, indicat ing that one activase tetramer must be able to service, continuously, as many as 200 Rubisco octamers. Plants with lower activase contents c ould not grow unless the atmosphere was enriched with CO2. Their Rubis co was less carbamylated and they had lower CO2 assimilation rates tha n the controls. The rate of release of 2'-carboxyarabinitol-1-phosphat e from Rubisco after illumination of the anti-activase leaves was also impaired. Older anti-activase plants accumulated increasing amounts o f Rubisco in their younger leaves, but were unable to carbamylate it. The photosynthetic rate per carbamylated Rubisco active site in the st rongly suppressed anti-activase leaves was only approximately 25% of t hat seen in control leaves, suggesting that activase may not only prom ote carbamylation of uncarbamylated Rubisco sites, but also accelerate turnover at carbamylated sites.