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Hybrids have been made between species of Flaveria exhibiting varying levels of C4 photosynthesis. The degree of C4 photosynthesis expressed in four interspecific hybrids (Flaveria trinervia [C4] × F. linearis [C3-C4], F. brownii [C4-like] × F. linearis, and two three-species hybrids from F. trinervia × [F. brownii × F. linearis]) was estimated by inhibiting phosphoenolpyruvate carboxylase in vivo with 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate (DCDP). The inhibitor was fed to detached leaves at a concentration of 4 mm, and apparent photosynthesis was measured at atmospheric levels of CO2 and at 20 and 210 mL L−1 of O2. Photosynthesis at 210 mL L−1 of O2 was inhibited 32% by DCDP in F. linearis, by 60% in F. brownii, and by 87% in F. trinervia. Inhibition in the hybrids ranged from 38 to 52%. The inhibition of photosynthesis by 210 mL L−1 of O2 was increased when DCDP was used, except in the C4 species, F. trinervia, in which photosynthesis was insensitive to O2. Except for F. trinervia, control plants with less O2 sensitivity (more C4-like) exhibited a progressively greater change in O2 inhibition of photosynthesis when treated with DCDP. This increased O2 inhibition probably resulted from decreased CO2 concentrations in bundle sheath cells due to inhibition of phosphoenolpyruvate carboxylase. The inhibition of photosynthesis by DCDP is concluded to underestimate the degree of C4 photosynthesis in the interspecific hybrids because increased direct assimilation of atmospheric CO2 by ribulose bisphosphate carboxylase may compensate for inhibition of phosphoenolpyruvate carboxylase.

The degree of C4 photosynthesis was assessed in four hybrids among C4, C4-like, and C3-C4 species in the genus Flaveria using 14C labeling, CO2 exchange, 13C discrimination, and C4 enzyme activities. The hybrids incorporated from 57 to 88% of the 14C assimilated in a 10-s exposure into C4 acids compared with 26% for the C3-C4 species Flaveria linearis, 91% for the C4 species Flaveria trinervia, and 87% for the C4-like Flaveria brownii. Those plants with high percentages of 14C initially fixed into C4 acids also metabolized the C4 acids quickly, and the percentage of 14C in 3-phosphoglyceric acid plus sugar phosphates increased for at least a 30-s exposure to 12CO2. This indicated a high degree of coordination between the carbon accumulation and reduction phases of the C4 and C3 cycles. Synthesis and metabolism of C4 acids by the species and their hybrids were highly and linearly correlated with discrimination against 13C. The relationship of 13C discrimination or 14C metabolism to O2 inhibition of photosynthesis was curvilinear, changing more rapidly at C4-like values of 14C metabolism and 13C discrimination. Incorporation of initial 14C into C4 acids showed a biphasic increase with increased activities of phosphoenolpyruvate carboxylase and NADP-malic enzyme (steep at low activities), but turnover of C4 acids was linearly related to NADP-malic enzyme activity. Several other traits were closely related to the in vitro activity of NADP-malic enzyme but not phosphoenolpyruvate carboxylase. The data indicate that the hybrids have variable degrees of C4 photosynthesis but that the carbon accumulation and reduction portions of the C4 and C3 cycles are well coordinated.

Here it is reported that sucrose synthase can be readily measured in growing wild tomato fruits (Lycopersicon chmielewskii) when suitable methods are adopted during fruit extraction. The enzyme also was present in fruit pericarp tissues, in seeds, and in flowers. To check for novel characteristics, the wild tomato fruit sucrose synthase was purified, by (NH(4))(2)SO(4) fraction and chromatography with DE-32, Sephadex G-200, and PBA-60, to one major band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The following characteristics were obtained: native protein relative molecular weight 380,000; subunit relative molecular weight 89,000; K(m) values with: sucrose 53 millimolar, UDP 18.9 micromolar, UDP-glucose 88 micromolar, fructose 8.4 millimolar; pH optima between 6.2 to 7.3 for sucrose breakdown and 7 to 9 for synthesis; and temperature optima near 50 degrees C. The enzyme exhibited a high affinity and a preference for uridylates. The enzyme showed more sensitivity to divalent cations in the synthesis of sucrose than in its breakdown. Sink strength in tomato fruits also was investigated in regard to sucrose breakdown enzyme activities versus fruit weight gain. Sucrose synthase activity was consistently related to increases in fruit weight (sink strength) in both wild and commercial tomatoes. Acid and neutral invertases were not, because the published invertase activity values were too variable for quantitative analyses regarding the roles of invertases in tomato fruit development. In rapidly growing fruits of both wild and commercially developed tomato plants, the activity of sucrose synthase per growing fruit, i.e. sucrose synthase peak activity X fruit size, was linearly related to final fruit size; and the activity exceeded fruit growth and carbon import rates by at least 10-fold. In mature, nongrowing fruits, sucrose synthase activities approached nil values. Therefore, sucrose synthase can serve as an indicator of sink strength in growing tomato fruits.

An analog of phosphoenolpyruvate, 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate (DCDP), was used to inhibit phosphoenolpyruvate carboxylase and, therefore, assess the contribution of the C4 pathway to photosynthesis in detached leaves of several C3-C4 intermediate species. There was no effect of 4 millimolar DCDP on apparent photosynthesis or on inhibition of apparent photosynthesis by 210 milliliters per liter of O2 for the C3-C4 species Panicum milioides, Moricandia arvensis, and Neurachne minor or the C3 species Flaveria pringlei. However, when leaves of Flaveria linearis (C3-C4), Flaveria brownii (C4-like), and Flaveria trinervia (C4) were fed 4 millimolar DCDP, photosynthesis was reduced 32, 60, and 90%, respectively. Photosynthetic inhibition by 210 milliliter per liter of O2 was also significantly increased in 4 millimolar DCDP-fed leaves of F. linearis and F. brownii but not in F. trinervia when compared with control values. These results with DCDP clearly demarcate C3-C4 species into species including Panicum, Moricandia, and Neurachne whose reduced photorespiration occurs without any C4 photosynthetic involvement and species of Flaveria in which C4 photosynthesis contributes to CO2 assimilation.

A plasma membrane-enriched fraction was isolated from various tissues of developing lima bean seedlings, Phaseolus lunatus var Cangreen, to study β-1,3-glucan synthase activity changes. All tissues contained an active β-glucan synthase, including the cotyledons that will be senescent in mature lima bean plants. Young primary leaves exhibited a very active β-glucan synthase; but this activity dropped markedly, about fivefold, as the leaves gained weight and became photosynthetic. Some tissues, such as the hypocotyl and young stem, exhibited an increase in β-glucan synthase activity as the tissues were growing and a decrease as the growth rate slowed. Roots exhibited a high activity early in development that only decreased slightly, about 30%, as root growth increased. Surprisingly the senescent cotyledons contained an activity equivalent to some other tissues that was maintained over our measurement time of 21 days. Perhaps this callose synthesis activity is related to translocation processes as the cotyledons transfer their reserves to the growing seedling. We concluded that β-glucan synthase was not a good indicator of sink strength in these lima bean tissues. The plasma membrane fractions also were tested for other enzymes that might be present because an electron microscope study revealed a low contamination by other types of membranes. The membrane fractions had low but detectable activities of sucrose synthase, UDPglucose pyrophosphorylase, UDPase, alkaline invertase, and a general phosphatase; but these enzymes exhibited no consistent pattern(s) of activity change with plant development.