Events Surrounding the Early Development of Euglena Chloroplasts: I Cellular Origins of Chloroplast Enzymes in Euglena*

Abstract

S U M M A R YDuring chloroplast development, the large increases in ribulose diphosphate carboxylase (RUDPCase) activity and cytochrome 5 5 2 concentration follow the pattern of chlorophyll synthesis, in that the formation of these two enzymes is inhibited by streptomycin (Sm) and by chloramphenicol (Cm) beyond 12 h of development. Neither enzyme can be detected in W,BUL, a mutant of Euglena in which chloroplasts and chloroplast DNA are undetectable. In contrast, the NADP-linked triose phosphate dehydrogenase (NADP-TPDase), another plastidlocalized enzyme, increases in activity without the 12 h lag normally observed for chlorophyll synthesis; this increase in activity is not inhibited by Sm and Cm, but is inhibited by cycloheximide, an antibiotic which acts on 87 S cytoplasmic ribosomes. NADP-TPDase activity is present at the same level in W,BUL as in the dark-grown wild-type organisms. These data are interpreted to mean that NADP-TPDase is coded in the nuclear DNA, and is translated on 8 7 s cytoplasmic ribosomes. The sensitivity of the increase in cytochrome 552 and RUDPCase activities to Sm and Cm indicates that they are translated, at least in part, on the 68 S ribosomes of the chloroplast. Thus, chloroplast differentiation in Euglena is dependent upon information and synthetic machinery from both the plastid and the rest of the cell. Since total cellular protein does not change significantly during chloroplast development in resting cells, we conclude that protein turnover probably occurs.

Events Surrounding the Early Development of Euglena Chloroplasts: Experiments with Streptomycin in Non-dividing Cells

“…Strep fomycin and plus f ids in resf ing Euglenu 61 development of the photosynthetic apparatus that we have measured are not affected by streptomycin, but some are strongly inhibited thereafter (Bovarnick et al 1974). This has been confirmed by us with fluorescence microscopy, and by electron microscopy (BenShaul, Silman & Ophir, 1972) where the structure of the developing plastid after 72 h in the presence of streptomycin resembles the normal plastid at 12 to 14 h development.…”

Nutritional Regulation of Organelle Biogenesis in Euglena

“…The 24-h lag period for the photoinduction of glycolate dehydrogenase in resting Euglena (Fig. 4) differs from the lag periods (0-12 h) previously reported for other light-induced enzymes (3,14,15). In resting cells, glycolate dehydrogenase synthesis appears to begin only after the cells become photosynthetically competent (27) supporting Merrett's proposal (20) that glycolate dehydrogenase synthesis is controlled by a product of photosynthesis rather than directly by light exposure.…”

Light-induced Enzyme Formation in a Chlorophyll-less Mutant of Euglena gracilis

“…Do, a measure of sensitivity of the UV targets (17), was 4.5 sec for the wild type organism and 3.5 sec for Y9 cells. Exposure of (4,5,11,24,26) we observed the gradual appearance of these enzyme activities in dark-grown wild type cells exposed to light. The time course for the appearance of these activities over a 48-to 72-hr period and the finding that cycloheximide (15 ,g/ml) inhibited 80 to 95% of the light-induced formation of each of the four proteins strongly support the notion that each enzyme is synthesized de novo during chloroplast development in Euglena.…”

“…The kinetics of light-induced synthesis of NADP glyceraldehyde-3-P dehydrogenase in regreening cells of Euglena has been measured by several investigators (3,4,11,26). Brawerman and Konigsberg (5) advanced the hypothesis that photosynthetically generated NADPH might serve as an inducer of this enzyme in Euglena, and more recently Ben-Amotz and Avron (2) demonstrated that disalicylidenepropanediamine (DSPD), a potent antagonist of ferredoxin, prevented the light-induced formation of NADPH glyceraldehyde-3-P dehydrogenase in Euglena.…”