Carbon Dioxide Fixation by Detached Cereal Caryopses1

Immature detached cereal caryopses from barley (Hordeum vulgare L. var distichum cv Midas) and wheat (Triticum aestivum L. cv Sicco) were shown to be capable of fixing externally supplied"'CO2 in the light or dark. Green cross cells and the testa contained the majority of the 14Clabeled material. Some "4C-labeled material was also found in the outer, or transparent, layer and in the endosperm/embryo fraction. More 14C was recovered from caryopses when they were incubated in '4CO2 without the transparent layer, thus suggesting that this layer is a barrier to the uptake of CO2. In all cases, significant amounts of 14C-labeled material were found in caryopses after dark incubation with '4CO2. Interestingly, CO2 fixation in the chlorophyll-less mutant Albino lemma was significantly greater in the light than in the dark. The results indicate that intact caryopses have the ability to translocate "4C-labeled assimilate derived from external CO2 to the endosperm/embryo. Carboxylating activity in the transparent layer appears to be confined to phosphoenolpyruvate carboxylase activity but that in the chloroplast-containing cross-cells may be accounted for by both ribulose-1,5-bisphosphate carboxylase-oxygenase and phosphoenolpyruvate carboxylase activity. Depending on a number of assumptions, the amount of CO2 fixed is sufricient to account for about 2% of the weight of starch found in the mature caryopsis. The main source of assimilates for grain-filling in cereals is thpught to be photosynthesis by the ear, stem, and flag leaf after pollination. In barley, and possibly other cereals, material stored in the stem before pollination may make a significant contribution to final grain yield (3). The proportion of total photosynthetic activity met by ear photosynthesis depends on a number of factors, including species, cultivar, environmental conditions, developmental stage and methods used in the determination of rates of ear photosynthesis. Thus, estimates of the contribution made by ear photosynthesis to final grain weight varies between as little as 13% and as much as 76% (8). Certainly, all the green tissues of the wheat ear are capable of photosynthesis (10) and therefore may be potential sources of assimilate for grain filling. The Chl-containing cells of the immature pericarp of barley, wheat, rye, rice, and oats are the site of caryopsis photosynthesis (8). These form a continuous band (known as the green layer) on the outside of the testa and in turn are covered by a layer of nonchlorenchymatous cells (known as the transparent layer). Together these two layers constitute the pericarp. The Chl-containing cells of the barley caryopsis are capable of high rates of light-dependent oxygen evolution (6), and there is evidence to suggest that in oats and wheat as well as in barley, these cells have some capacity for C4 metabolism (11, 12). In barley pericarps for example, the first-formed product of photosynthesis is ' Support from the Agricultural and Food Research Council, London, is gratefully acknowledged. the C4-acid malate which is subsequently rapidly converted to sucrose (11). Evans and Rawson (10) have found by infrared gas analysis that wheat grains can assimilate CO2 at rates almost equal to rates of dark respiration until the late stages of grain development. However, we do not know whether or not atmospheric CO2 fixed by the caryopsis can be translocated to the endosperm/ embryo. Nor do we know what contribution pericarp photosynthesis may make to the overall economy of grain-filling. The present work is an attempt to answer these questions. MATERIALS AND METHODS Plant Material. Barley (Hordeum vulgare L. var hexastichum Albino lemma, and Hordeum vulgare L. var distichum cv Midas) and wheat (Triticum aestivum L. cv Sicco) were grown from seed in pots of 180 mm diameter. Six plants per pot were grown in Levington's peat-based potting compost (Fisons Ltd, Horticulture Division, Bramford, Ipswich, UK) under glasshouse conditions. Natural daylength was extended to 20 h with 400 W mercury vapor lamps and the mean daily temperature was 18°C. The ages of the barley cv Midas caryopses were determined using a developmental time scale of 60 'days' from anthesis to harvestripeness (5). Wheat caryopses were aged using a similar developmental time scale based on field-grown wheat harvested in 1981. A. lemma is a six-row barley and is a chemically induced mutant. The pericarp, lemma, and palea of the grain are albino with the exception of small green areas at the tips of the lemma and palea. However, the awns, sterile glumes, and remainder of the plant contain Chl, and the plant grows well in a glasshouse after vernalization. The age of each ear of A. lemma was estimated by ageing an ear of cv Midas which reached anthesis on the same day as that of the A. lemma ear. In this way, comparison can be made at similar stages of development in caryopses from different species. All caryopses were 25 d after anthesis since at this stage Chl content and grain growth rates are maximal (6). "4CO2 Fixation by Detached Caryopses. The method used was a modification of the method of Nutbeam and Duffus (11). The sterile glumes, lemma, and palea were dissected from 4 grains, and the caryopses were placed on filter paper discs previously soaked in 330 mm sorbitol, 50 mm Tricine-KOH (pH 7.5). In a number of experiments, the transparent layer of the pericarp was also removed. The detached caryopses on moist discs were placed in an airtight Perspex chamber (volume 11 cm3) and 0.02 cm3 of 17 mm sodium [14C]carbonate (specific activity 2.16 x 106 Bq moll) was injected through a rubber seal into a well containing 0.15 cm3 of 13 M lactic acid. The final concentration of CO2 in the chamber was 0.1% by volume. The chamber was illuminated with a 1000 W tunsten halogen lamp (photon flux density in the chamber was 710 ,tmol m-2 s1) for varying periods of time. A Perspex box (dimensions 250 x 250 x 60 mm) with cold water circulating through it was positioned between the lamp and the chamber to maintain the chamber at 21°C. The light-dependence of 14C02 fixation was