The aim of the study was to verify the hypothesis that a potential cause of the phytotoxicity of diclofenac (DCF, a non-steroidal anti-inflammatory drug) is an effect of cell cycle progression. This research was conducted using synchronous cultures of a model organism, green alga Chlamydomonas reinhardtii. The project examined DCF effects on selected parameters that characterize cell cycle progression, such as cell size, attainment of commitment points, DNA replication, number of nuclei formed during cells division and morphology of cells in consecutive stages of the cell cycle, together with the physiological and biochemical parameters of algae cells at different stages. We demonstrated that individual cell growth remained unaffected, whereas cell division was delayed in the DCF-treated groups grown in continuous light conditions, and the number of daughter cells from a single cell decreased. Thus, the cell cycle progression is a target affected by DCF, which has a similar anti-proliferative effect on mammalian cells.

• Keywords
• Chlamydomonas reinhardtii, cell cycle, diclofenac, non-steroidal anti-inflammatory drug,
• MeSH
• Anti-Inflammatory Agents, Non-Steroidal toxicity   MeSH
• Cell Division drug effects   MeSH
• Cell Cycle drug effects   MeSH
• Chlamydomonas reinhardtii drug effects   genetics   growth & development   MeSH
• Diclofenac toxicity   MeSH
• DNA, Plant biosynthesis   genetics   MeSH
• Photosynthesis drug effects   MeSH
• DNA Replication drug effects   MeSH
• Cell Size drug effects   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Anti-Inflammatory Agents, Non-Steroidal MeSH
• Diclofenac MeSH
• DNA, Plant MeSH

Green algae are fast-growing microorganisms that are considered promising for the production of starch and neutral lipids, and the chlorococcal green alga Parachlorella kessleri is a favorable model, as it can produce both starch and neutral lipids. P. kessleri commonly divides into more than two daughter cells by a specific mechanism-multiple fission. Here, we used synchronized cultures of the alga to study the effects of supra-optimal temperature. Synchronized cultures were grown at optimal (30 °C) and supra-optimal (40 °C) temperatures and incident light intensities of 110 and 500 μmol photons m-2 s-1. The time course of cell reproduction (DNA replication, cellular division), growth (total RNA, protein, cell dry matter, cell size), and synthesis of energy reserves (net starch, neutral lipid) was studied. At 40 °C, cell reproduction was arrested, but growth and accumulation of energy reserves continued; this led to the production of giant cells enriched in protein, starch, and neutral lipids. Furthermore, we examined whether the increased temperature could alleviate the effects of deuterated water on Parachlorella kessleri growth and division; results show that supra-optimal temperature can be used in algal biotechnology for the production of protein, (deuterated) starch, and neutral lipids.

• Keywords
• Parachlorella kessleri, cell cycle, deuterated lipid, deuterated starch, deuterium, energy reserves, growth processes, microalgae, reproduction events, starch, supra-optimal temperature,
• MeSH
• Biomass MeSH
• Cell Division physiology   MeSH
• Chlorophyta growth & development   MeSH
• Lipids MeSH
• Lipid Metabolism physiology   MeSH
• Microalgae metabolism   MeSH
• Starch metabolism   MeSH
• Temperature * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Lipids MeSH
• Starch MeSH

Temperature is one of the key factors affecting growth and division of algal cells. High temperature inhibits the cell cycle in Chlamydomonas reinhardtii. At 39 °C, nuclear and cellular divisions in synchronized cultures were blocked completely, while DNA replication was partly affected. In contrast, growth (cell volume, dry matter, total protein, and RNA) remained unaffected, and starch accumulated at very high levels. The cell cycle arrest could be removed by transfer to 30 °C, but a full recovery occurred only in cultures cultivated up to 14 h at 39 °C. Thereafter, individual cell cycle processes began to be affected in sequence; daughter cell release, cell division, and DNA replication. Cell cycle arrest was accompanied by high mitotic cyclindependent kinase activity that decreased after completion of nuclear and cellular division following transfer to 30 °C. Cell cycle arrest was, therefore, not caused by a lack of cyclin-dependent kinase activity but rather a blockage in downstream processes.

• Keywords
• Chlamydomonas reinhardtii, DNA replication, cell cycle arrest, cell size, cyclin-dependent kinase, starch accumulation, supraoptimal temperature, synchronized cultures,
• MeSH
• Algal Proteins metabolism   MeSH
• Cell Culture Techniques methods   MeSH
• Chlamydomonas reinhardtii cytology   physiology   MeSH
• Cyclin-Dependent Kinases metabolism   MeSH
• Down-Regulation MeSH
• Stress, Physiological MeSH
• Cell Cycle Checkpoints * MeSH
• Gene Expression Regulation, Plant MeSH
• Hot Temperature MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Algal Proteins MeSH
• Cyclin-Dependent Kinases MeSH

DNA damage is a threat to genomic integrity in all living organisms. Plants and green algae are particularly susceptible to DNA damage especially that caused by UV light, due to their light dependency for photosynthesis. For survival of a plant, and other eukaryotic cells, it is essential for an organism to continuously check the integrity of its genetic material and, when damaged, to repair it immediately. Cells therefore utilize a DNA damage response pathway that is responsible for sensing, reacting to and repairing damaged DNA. We have studied the effect of 5-fluorodeoxyuridine, zeocin, caffeine and combinations of these on the cell cycle of the green alga Scenedesmus quadricauda. The cells delayed S phase and underwent a permanent G2 phase block if DNA metabolism was affected prior to S phase; the G2 phase block imposed by zeocin was partially abolished by caffeine. No cell cycle block was observed if the treatment with zeocin occurred in G2 phase and the cells divided normally. CDKA and CDKB kinases regulate mitosis in S. quadricauda; their kinase activities were inhibited by Wee1. CDKA, CDKB protein levels were stabilized in the presence of zeocin. In contrast, the protein level of Wee1 was unaffected by DNA perturbing treatments. Wee1 therefore does not appear to be involved in the DNA damage response in S. quadricauda. Our results imply a specific reaction to DNA damage in S. quadricauda, with no cell cycle arrest, after experiencing DNA damage during G2 phase.

• MeSH
• Bleomycin pharmacology   MeSH
• Cell Cycle drug effects   genetics   MeSH
• Chlorophyta MeSH
• Floxuridine pharmacology   MeSH
• G2 Phase genetics   MeSH
• Caffeine pharmacology   MeSH
• DNA Repair drug effects   MeSH
• DNA Damage physiology   MeSH
• Cell Cycle Proteins MeSH
• Scenedesmus cytology   genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bleomycin MeSH
• Floxuridine MeSH
• Caffeine MeSH
• Cell Cycle Proteins MeSH
• Zeocin MeSH Browser

We developed an alternative method of staining cell nuclei and chloroplast nucleoids of algal cells using SYBR Green I (the fluorescent dye used commonly for detecting dsDNA in agarose and polyacrylamide gels as an alternative to highly mutagenic ethidium bromide and for DNA staining of viruses and bacteria followed by flow cytometry, digital image analysis or real-time PCR), which enabled routine staining in vivo. Cells do not need to be fixed or treated chemically or physically before staining, thus the shape, size and position of DNA-containing structures are not affected. The fluorescence signal is sharp and reproducible. Examples of application of the method are shown in color microphotographs for representatives of eukaryotic algae from the taxa Chlorophyta, Rhodophyta and the prokaryotic Cyanophyta. The method is also useful for studying progress of the cell cycle in algal cells dividing by multiple fission, as shown by observation of changes in nuclear number during the cell cycle of the green alga Chlamydomonas reinhardtii and Scenedesmus quadricauda. Staining with SYBR Green I can be recommended as a fast, safe and efficient method for the detection of DNA-containing structures in vivo.

• MeSH
• Staining and Labeling methods   MeSH
• Benzothiazoles MeSH
• Cell Cycle MeSH
• Quinolines MeSH
• Chlorophyta genetics   physiology   ultrastructure   MeSH
• Diamines MeSH
• DNA, Algal analysis   MeSH
• Fluorescent Dyes metabolism   MeSH
• Organic Chemicals metabolism   MeSH
• Cell Membrane Permeability MeSH
• Image Processing, Computer-Assisted MeSH
• Polymerase Chain Reaction MeSH
• Flow Cytometry MeSH
• Reproducibility of Results MeSH
• Rhodophyta genetics   physiology   ultrastructure   MeSH
• Cyanobacteria genetics   physiology   ultrastructure   MeSH
• Publication type
• Journal Article MeSH
• Evaluation Study MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• Benzothiazoles MeSH
• Quinolines MeSH
• Diamines MeSH
• DNA, Algal MeSH
• Fluorescent Dyes MeSH
• Organic Chemicals MeSH
• SYBR Green I MeSH Browser

The aim of the study was to elucidate the effect of cadmium ions on the arrangement of the actin and tubulin cytoskeleton, as well as the relationships between cytoskeletal changes and growth processes in the green filamentous alga Spirogyra decimina. Batch cultures of algae were carried out under defined conditions in the presence of various cadmium concentrations. In control cells, the cytoskeleton appeared to be a transversely oriented pattern of both microtubules and actin filaments of various thickness in the cell cortex; colocalization of cortical microtubules and actin filaments was apparent. Microtubules were very sensitive to the presence of cadmium ions. Depending on the cadmium concentration and the time of exposure, microtubules disintegrated into short rod-shaped fragments or they completely disappeared. A steep increase in cell width and a decrease in growth rate accompanied (and probably ensued) a very rapid disintegration of microtubules. Actin filaments were more stable because they were disturbed several hours later than microtubules at any cadmium concentration used. When cadmium ions were washed out, the actin cytoskeleton was rebuilt even in cells in which actin filaments were completely disintegrated at higher cadmium concentrations (40 or 100 microM). The much more sensitive microtubules were regenerated after treatment with lower cadmium concentrations (10 or 15 microM) only.

• MeSH
• Cell Division drug effects   MeSH
• Time Factors MeSH
• Chlorophyta drug effects   growth & development   metabolism   MeSH
• Cytoskeleton drug effects   ultrastructure   MeSH
• Microscopy, Fluorescence MeSH
• Interphase MeSH
• Cadmium pharmacology   MeSH
• Actin Cytoskeleton drug effects   ultrastructure   MeSH
• Microtubules drug effects   ultrastructure   MeSH
• Metals, Heavy pharmacology   MeSH
• Dose-Response Relationship, Drug MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Cadmium MeSH
• Metals, Heavy MeSH

Methods for determining the points of commitment for cell division are described for species of green algae dividing by multiple fission, both forming coenobia (Scenedesmus quadricauda) and releasing single daughter cells (Chlamydomonas eugametos, Scenedesmus armatus). The timing of commitment points was followed in detail in synchronous cultures of S. quadricauda grown under various light intensities, illumination regimes, and temperatures. The pre-commitment periods were rate limiting, while the post-commitment periods remained more or less constant under various light intensity. Temperature, on the other hand, affected both periods in a similar manner and they were prolonged with decreasing temperature.

• MeSH
• Cell Division * physiology   radiation effects   MeSH
• Cell Cycle * MeSH
• Chlamydomonas cytology   growth & development   MeSH
• Culture Media MeSH
• Scenedesmus cytology   growth & development   MeSH
• Light MeSH
• Temperature MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Evaluation Study MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Culture Media MeSH

The courses of rRNA accumulation, DNA replication, and nuclear division were followed both in the chloroplast and the nucleocytosolic compartments during the cell cycle in synchronized populations of the chlorococcal alga Scenedesmus quadricauda. Control and nalidixic acid-treated cultures were compared. Nalidixic acid (150 mg/L) was added either at the beginning of the cell cycle or consecutively during the cell cycle to subcultures transferred into the dark. If the inhibitor was applied at the beginning of the cell cycle, chloroplast DNA did not replicate and nucleoids did not divide. Chloroplast division, however, was coordinated in a timely fashion with cytokinesis even under conditions of blocked chloroplast DNA replication. While the growth rate was slowed down, the courses of reproductive processes in the nucleocytosolic compartment were not affected and their timing and the number of rounds were coordinated with growth rate as in the control culture. The rate of cytosolic rRNA synthesis was lower but no apparent effect was seen on the amount of rRNA that accumulated during the cell cycle. In contrast, lower levels of chloroplast rRNA were found at the end of the cell cycle compared with the control culture. Experiments in which cells were transferred to the dark during the cell cycle showed that the inhibitor affected none of the reproductive events in the nucleocytosolic compartment. In the chloroplast compartment, DNA replication was inhibited in inhibitor-treated cultures, but was unaffected in controls. The chloroplast nucleoids themselves divided even in the presence of the inhibitor, reducing their DNA content to a level which corresponded to that in freshly formed control daughter cells.

• MeSH
• Chloroplasts drug effects   MeSH
• Nalidixic Acid pharmacology   MeSH
• Scenedesmus drug effects   growth & development   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Nalidixic Acid MeSH