Reduced Expression of Succinyl-Coenzyme A Ligase Can Be Compensated for by Up-Regulation of the γ-Aminobutyrate Shunt in Illuminated Tomato Leaves

Growth phenotype of SCoAL transgenic tomato plants. Transgenic plants showed reduced fruit yield and slightly reduced leaf biomass. A, Photograph showing representative plants after 8 weeks of growth. B, Biomass (in g/DW) of various plant organs on plant maturity. Wild type, Black bar; AL18, light gray bar; RL40, dark gray bar; RL25, white bar. Values are presented as mean ± se of determination on six individual plants per line; an asterisk indicates values that were determined by the t test to be significantly different (P < 0.05) from the wild type. SCoAL activity of the lines determined 2 weeks prior to these experiments were 162.4 ± 10.7, 134.6 ± 3.4, 139.7 ± 7.5, and 12.4 ± 2.9 nmol⁻¹ min⁻¹ g⁻¹ FW for wild type, AL18, RL40, and RL25, respectively.

Effect of decreased SCoAL activity on photosynthesis. A, Photosynthetic carbon assimilation and partitioning at the onset of illumination. Leaf discs were cut from six separate plants of each genotype, after 6 weeks of growth, at the end of the night and illuminated at 150 μmol photons m⁻² s⁻¹ of photosynthetically active radiation in an oxygen electrode chamber containing air saturated with ¹⁴CO₂. After 30 min, the leaf discs were extracted and fractionated. Wild type, Black bars; AL18, light gray bars; RL40, dark gray bars; RL25, white bars. B, In vivo fluorescence emission was measured as an indicator of the electron transport rates by use of a PAM fluorometer at PFDs ranging from 0 to 1,000 μmol photons m⁻² s⁻¹. C, Assimilation rate as a function of PFD. Wild type, Black circles; RL40, white triangles; RL25, black squares. Values are presented as mean ± se of determinations on six individual plants per line. Asterisk indicates values that were determined by the t test to be significantly different (P < 0.05) from the wild type. SCoAL activity of the lines determined in parallel to these experiments was identical to that described in the legend for Figure 2.

Respiratory parameters in leaves of the SCoAL transgenic lines. ¹⁴CO₂ evolution from isolated leaf discs, after 6 weeks of growth in the light. Leaf discs were taken from 10-week-old plants and were incubated in 10 mm MES-KOH solution, pH 6.5, 0.3 mm Glc supplemented with [1-¹⁴C], [2-¹⁴C], [3:4-¹⁴C], or [6-¹⁴C]Glc (at a specific activity of 7.5 MBq mmol⁻¹). The ¹⁴CO₂ liberated was captured, at hourly intervals, in a KOH trap and subsequently quantified by liquid scintillation counting. Values are presented as means ± se of six determinants per line. SCoAL activity of the lines determined in parallel to these experiments was identical to that described in the legend for Figure 2.

Relative metabolite content of fully expanded leaves from 6-week-old plants of the SCoAL transgenic lines. Metabolites were determined as described in the “Materials and Methods.” The full dataset can be accessed at our Web site (www.mpimp-golm.mpg.de/fernie). Data are normalized with respect to the mean response calculated for the wild type. Values are presented as mean ± se of determinations on six individual plants per line. Asterisk indicates values that were significantly different from wild type when assessed by t tests (P < 0.05). Wild type, Black bars; AL18, light gray bars; RL40, dark gray bars; RL25, white bars. SCoAL activity of the lines determined in parallel to these experiments was identical to that described in the legend for Figure 2.

Differences in transcript levels between leaves of the well-characterized SCoAL line RL25 and wild type for genes associated with metabolism (A) and regulation (B). Both sets of material were harvested after 6 weeks of growth in the middle of the day. Red and blue represent a decrease and an increase of expression, respectively, in the SCoAL transgenic line with respect to the wild type. The color scale used is reproduced in the figure. This figure and all data point annotations are best viewed at http://gabi.rzpd.de/projects/MapMan (see “Materials and Methods”). SCoAL activity of the lines determined in parallel to these experiments was identical to that described in the legend for Figure 2.

Estimation of GABA shunt activity. A, Redistribution of label following feeding of leaves [U-¹³C]Glu via the petiole (see “Materials and Methods”). Values are presented as the accumulation of micromolar C1 equivalents per hour per gram FW. B, Flux through the GABA shunt was determined by measuring ¹⁴CO₂ release from leaf discs incubated in [1-¹⁴C]Glu. Incubation was performed as described in “Materials and Methods.” Values are presented as means ±se of six determinants per line. Wild type, Black bars; AL18, light gray bars; RL40, dark gray bars; RL25, white bars. For experiment A, SCoAL activity of the lines was identical to that described in the legend for Figure 2. For experiment B, the measurements of additional lines, RL34 and AL10, as well as SCoAL activity of the lines are provided in Supplemental Table S4.