13H1-and 8-14CIAminolevulinic acids were incorporated into the chlorophylls of Skektonema costatwn, a marine plankton diatom. In the stationary phase of growth, the tetrapyrrole-based pigments reached steadystate labeling after 10 hours. Under conditions of exponential cell division and chlorophyll accumulation, 3H was rapidly lost from the labeled chlorophylls and was replaced with 14C derived from acid. The kinetics of isotope dilution suggests recycling of tetrapyrrole precursors and/or two pigment pools, containing both chlorophyll a and chlorophyllide c, one which turns over rapidly (10 hours) and another which turns over more slowly (100 hours). Calculation of turnover times varied from 3 to 10 hours for chlorophyll a and from 7 to 26 hours for chlorophyllide c. The data suggest the dynamics of chlorophyll metabolism in S. costatum and explain the diatom's ability to undergo light-shade adaptation within a generation time.Light available for photosynthesis is highly variable in aquatic environments. As a result of mixing, wave action, scattering, and heterogeneous extinction, unicellular planktonic algae are potentially exposed to changes in light intensity of one or two orders of magnitude within a period of hours. These algae have evolved diverse and complex light-harvesting pigment systems, some of which are capable of adapting to variations in incident light. Changes in intracellular photosynthetic pigments, such as Chl a (light-shade adaptation), or carotenoids (chromatic adaptation) are often observed within a short time after exposure to a new light regime (5, 15). Such rapid and reversible responses in pigmentation suggest that the metabolism of Chl and accessory pigments is highly dynamic, and imply that pigments may turn over rapidly in vivo.In batch cultures, cell constituents are diluted as a consequence ofgrowth, and the significance of turnover as a means of regulating the steady-state level of many metabolites has been overlooked. We have attempted to examine the turnover of Chl a and chlide c in a common neritic diatom, Skeletonema costatum, as a means of understanding the regulation of light-shade adaptation.The [3,194 mCi/mmol) were purchased from New England Nuclear.Culture Conditions. S. costatum (Grev.) Cleve, Woods Hole clone SKEL, was grown on a 14:10 light-dark cycle at 15 C as previously described (10). The cultures were continuously mixed by stirring, and gently bubbled with sterile air by negative pressure. After 4 to 5 days of growth (cell density of about 2.5 x 106 cells/ml) a 500-ml aliquot was inoculated into 3 liters of fresh media containing 0.5 to 0.6 fLM [3,5-3H]ALA. The reinoculation resulted in a shift-up (12) in the permeability of the cell to ALA, allowing for enhanced uptake of ALA (see under "Results"). Following ALA uptake in isotope dilution and dual label experiments the cultures were continuously centrifuged at 12,000 g at 15 C with a flow rate of about 200 ml/min. The packed cells were washed with filtered seawater and resuspended in fresh media c...

The accumulation of 8-aminolevulinic acid and activities of 8-aminolevulinic acid dehydrase were examined in the marine diatom, Skektonema costatum, grown in the presence of levulinic acid. Levuinic acid concentrations greater than 10 mm affect growth and morphology, and inhibit chlorophyll synthesis. The algae recover from the effects of levulinic acid after 48 hours of exposure. The recovery is characterized by increased cellular cholorphyll content, decreased 8-aminolevulinic acid accumulation, decreased 3-(3,4-dchiorophenyl)-1,1-dimethylurea-enhanced in vivo fluorescence, and the induction of a levulinic acid-activated 8-aminolevuinic acid dehydrase which does not follow Michaelis-Menten kinetics. The data indicate that levuinic acid blocks may be ineffective in vivo, and that 8-aminolevulHlnc acid is metabolized to amino and dicarboxylc acids. 8-Aminolevulnc acid dehydrase activities are used to estimate the capacity for chlorophyll synthesis. Results suggest this diatom may be capable of rapid chlorophyll turnover, which would allow the plant to Light-shade adapt on the time scales appropriate to vertical mixing rates in the sea.In the sea, light limits primary production to the upper tens of meters. Many species of marine phytoplankton maximize their photosynthetic capacity by acclimating to variations in the ambient light intensity through changes in cellular Chl content. As the cells are mixed by physical processes through the euphotic zone, light-shade adaptation on the time scales of vertical mixing rates would be greatly facilitated by rapid Chi metabolism. The potential to regulate Chl degradation and synthesis rapidly makes marine phytoplankton ideal organisms for studies of Chl metabolism and light-shade adaptation.Chl shares a common biosynthetic pathway with porphyrins and other tetrapyrroles (11). ALA4 is the first unique intermediate in the pathway, and its synthesis apparently has a major role in the regulation of ChM synthesis (6,7, 23 bacteria (27) and a variety of animal tissues (13). This enzyme has not been conclusively demonstrated in algae or higher plants (3).The labeling studies of Beale (3), Wellburn (29), and others imply that glutamate, a-ketoglutarate, and [1,5-'4C]citrate are more readily incorporated into ALA than succinyl-CoA or glycine. At least four alternative pathways leading to ALA have been proposed, but the most accepted appears to be via the transamination of 4,5-dioxovaleric acid (3, 16).The second enzyme in the porphyrin pathway is ALAD which appears to be ubiquitous. Beale (1, 2) found that treatment of cells or tissue with LEV, a competitive inhibitor of ALAD, results in an accumulation of ALA in vivo. LEV concentrations between 10 and 50 mm inhibited Chl a synthesis by as much as 80% without obvious deleterious effects. The LEV block has been extensively used in studies of ALA and Chl synthesis in algae and greening tissues of higher plants (4,5,15,(19)(20)(21).ALA does not accumulate in untreated plant cells (10), and its condensation to PBG is not known ...