Absence of fluorescence quenching in a photosynthetic mutant of Euglena gracilis.

A photosynthetic mutant of Euglena gracilis, Z strain, thought to be blocked in the electron transport chain between the two photosystems and to have a missing or nonfunctional primary acceptor for photosystem II, was further studied and characterized. The data from low temperature fluorescence spectra, delayed light emission, and electron paramagnetic resonance support the previous work.The mutant was shown to lack plastoquinone A and possibly cytochrome b5m but to possess plastoquinone B and a higher complement of carotenoids, especially xanthophylls, than the wild type.The results are consistent with the postulated role of plastoquinone as the primary electron acceptor for photosystem II and as responsible for the electron paramagnetic resonance signal II. The abundance of xanthophylls in the mutant can be explained by the protective role of carotenoids against photosensitized reactions.Russell and Lyman (23)

Section: Resultssupporting

confidence: 82%

“…In the light of evidence from the kinetics of fluorescence induction in the mutant P4, Russell et al (23) hypothesized that the primary acceptor for photosystem II (also known as the quencher or Q) was missing or nonfunctional. Our data from delayed light emission support this suggestion.…”

Section: Discussionmentioning

confidence: 99%

Biochemical and Biophysical Characteristics of a Photosynthetic Mutant of Euglena gracilis Blocked in Photosystem II

Schwelitz¹,

Dilley²,

Crane³

1972

“…Even after extensive growth in the dark in the presence of Nal, only a small fraction of the population ever shows inhibition of green colony-forming ability (9). The role of light in enhancing the Nal effect was investigated using P4ZUL, a mutant of Euglena, which has a block in photosynthetic electron transport (14,15) and DCMU, an inhibitor of photosynthetic electron transport. PZUL, while capable of making chloroplasts in the light, is unable to photosynthesize.…”

Section: Resultsmentioning

confidence: 99%

Action of Nalidixic Acid on Chloroplast Replication in Euglena gracilis

Lyman¹,

Jupp²,

Larrinua³

1975

The role of light in nalidixic acid bleaching of Euglena gracilis var. bacillaris was investigated. The kinetics of loss of the chloroplast-associated DNA and the sensitivity of chloroplast replication to ultraviolet light was followed during treatment with nalidixic acid. By using the mutant P4ZUL, and 3-(3,4-dichlorophenyl)-, 1-dimethylurea, it was demonstrated that the requirement for light was a functioning photosynthetic electron transport system. Ultracentifugal analysis showed a substantial decrease in chloroplast-associated DNA after 6 hours of treatment with nalidixic acid. Ultraviolet target analysis revealed that the number of chloroplast genomes per cell had been reduced. The possible role of light and implications of the reduction in chloroplast genomes for chloroplast replication are discussed.Chloroplast replication in Euglena gracilis is specifically inhibited by Na12 (5, 6, 9, 13), a potent inhibitor of DNA synthesis (2,7,8 and grown for experiments on a defined organic medium at pH 3.6 (18). Incubation was carried out in gyrotary water bath shakers (New Brunswick) at 26 C ± 0.5 C. The cultures were illuminated with a bank of four 40-w cool white Westingthouse fluorescent bulbs supplemented by cool beam tungsten 150-and 75-w bulbs (General Electric) to give an intensity of 3.6 X 10, ergs/ cm2 sec.Green colony-forming ability was assessed on plates using the above medium (18).Preparation of Nalidixic Acid. A fresh stock solution of Nal' (5 mg/ml) was prepared for each experiment as follows: 100 mg of Nal powder were dissolved in 0.8 ml of 1 N NaOH and distilled H20 added to 100 ml. The solution was sterilized by Millipore fitration. The final concentration of Nal for all experiments was 50 ,ug/ml of culture.Preparation of DCMU. For the stock solution DCMU was dissolved in 95% ethanol. The final concentration for all experiments was 10-sM.DNA Isolation. A total of 3 to 4 X 10' log phase cells was harvested by centrifugation at 3,000 rpm in a Sorvall refrigerated centrifuge at 4 C. The resultant pellet was washed once with distilled H.O followed by a second wash with saline-EDTA. Those cells not extracted immediately were fast frozen in Dry Ice and acetone then stored at -5 C.DNA was isolated using a modification of the Marmur method (12). The pellet was suspended in a 5-ml cold saline-EDTA (0.15 M NaCl, 0.1 M Na2EDTA, pH 8). SLS (25%'o, freshly prepared) was added to a final concentration of 0.2 mg/ml of suspension and lysis was determined by microscopic examination. If lysis was incomplete, the tube was warmed with the hands and/or a small additional amount of SLS added. The material was then incubated with 1 mg/ml pronase (stock solution 10 mg/ml, predigested) at 37 C. The lysate was placed in an ice bath, and 5 M sodium perchlorate was added to a final concentration of 1 M. One volume of chloroform-isoamnl alcohol (24: 1, v/v) was added, and the suspension was placed in an ice bath for 30 min. During this time the emulsion was agitated frequently with a gentle reciprocal motion. Followi...

“…A number of mutants that were isolated previously in our laboratory (31,32,35) have been used in this study. These mutants have blocks in chloroplast development and have been used to study the regulation of chloroplast DNA synthesis.…”

Section: Cellmentioning

confidence: 99%

Photomorphogenic Regulation of Chloroplast Replication in Euglena

Srinivas

Lyman²

1980

Dark-grown cells of Euglena contain proplastids which, upon exposure to light, differentiate into chloroplasts. During this process, structural and functional changes occur, including the synthesis of pigments, enzymes of photosynthetic electron transport, enzymes of CO2 reduction, chloroplast membrane proteins, and lipids and chloroplast ribosomes (36).Holowinsky and Schiff (17) In addition to the increase of plastid DNA during chloroplast development, Manning and Richards (24) noted that chloroplast DNA undergoes one and one-half rounds of DNA replication per one round of nuclear DNA replication, yet the amount of chloroplast DNA per cell remains constant. They postulated that a mechanism must exist to remove the "extra" DNA. Egan, Dorsky and Schiff (12) reported the light activation of a chloroplast associated DNase. This enzyme was further characterized by Small and Sturgen (40). The action spectrum for the light activation of the nuclease is similar to the action spectrum for the synthesis of Chl and some chloroplast-associated enzymes. Schmidt and Lyman (unpublished data) found that the amount of chloroplast DNA per cell increases in high intensities of blue light. All this evidence suggests the operation of a mechanism to regulate the amount of plastid DNA during chloroplast development as well as during growth. Regulation seems to be operating to keep the amount of chloroplast DNA constant because there is more DNA synthesized per chloroplast division than is needed for merely doubling the DNA content (24).In this paper, we propose that such regulation may occur through a restriction-type nuclease which degrades a fraction of the chloroplast DNA. We also propose that the putative nuclease is activated by the red-blue photomorphogenic system which mediates, with the blue chromophore, chloroplast development.Chloroplast DNA in Euglena constitutes 3 to 5% of the total cellular DNA (39