Nejvíce citovaný článek - PubMed ID 21573815
Chlamydomonas reinhardtii: duration of its cell cycle and phases at growth rates affected by temperature
This work represents a detailed guide for commitment point analysis in microalgae dividing by multiple fission. The method is based on allowing the committed cells to divide in favorable conditions in the dark. This protocol offers a strategy to monitor cell cycle progression, both in control cultures and cultures treated with compounds affecting cell cycle length and/or progression. As the variety of such compounds is wide, our aim was to make the protocol easily modifiable to various research aims. The technique is easy to follow, low-cost, does not require any special equipment and offers reliable results in a reasonable time. The protocol offers step-by-step instructions, explains the theory behind these steps and offers solutions to some of the problems that may arise during the procedure.
- Klíčová slova
- Cell cycle, Cell division, Chlamydomonas, Commitment point, Green algae, Multiple fission,
- MeSH
- buněčné dělení * MeSH
- buněčný cyklus * MeSH
- Viridiplantae MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
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.
- Klíčová slova
- Chlamydomonas reinhardtii, cell cycle, diclofenac, non-steroidal anti-inflammatory drug,
- MeSH
- antiflogistika nesteroidní toxicita MeSH
- buněčné dělení účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- Chlamydomonas reinhardtii účinky léků genetika růst a vývoj MeSH
- diklofenak toxicita MeSH
- DNA rostlinná biosyntéza genetika MeSH
- fotosyntéza účinky léků MeSH
- replikace DNA účinky léků MeSH
- velikost buňky účinky léků MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antiflogistika nesteroidní MeSH
- diklofenak MeSH
- DNA rostlinná 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.
- Klíčová slova
- Parachlorella kessleri, cell cycle, deuterated lipid, deuterated starch, deuterium, energy reserves, growth processes, microalgae, reproduction events, starch, supra-optimal temperature,
- MeSH
- biomasa MeSH
- buněčné dělení fyziologie MeSH
- Chlorophyta růst a vývoj MeSH
- lipidy MeSH
- metabolismus lipidů fyziologie MeSH
- mikrořasy metabolismus MeSH
- škrob metabolismus MeSH
- teplota * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- lipidy MeSH
- škrob MeSH
The extremophilic unicellular red microalga Galdieria sulphuraria (Cyanidiophyceae) is able to grow autotrophically, or mixo- and heterotrophically with 1% glycerol as a carbon source. The alga divides by multiple fission into more than two cells within one cell cycle. The optimal conditions of light, temperature and pH (500 µmol photons m-2 s-1, 40 °C, and pH 3; respectively) for the strain Galdieria sulphuraria (Galdieri) Merola 002 were determined as a basis for synchronization experiments. For synchronization, the specific light/dark cycle, 16/8 h was identified as the precondition for investigating the cell cycle. The alga was successfully synchronized and the cell cycle was evaluated. G. sulphuraria attained two commitment points with midpoints at 10 and 13 h of the cell cycle, leading to two nuclear divisions, followed subsequently by division into four daughter cells. The daughter cells stayed in the mother cell wall until the beginning of the next light phase, when they were released. Accumulation of glycogen throughout the cell cycle was also described. The findings presented here bring a new contribution to our general understanding of the cell cycle in cyanidialean red algae, and specifically of the biotechnologically important species G. sulphuraria.
- Klíčová slova
- Galdieria, cell cycle, cell division, growth, light intensity, red algae, synchronization, temperature, trophic regimes,
- MeSH
- buněčný cyklus fyziologie MeSH
- heterotrofní procesy fyziologie MeSH
- kultivované buňky MeSH
- mikrořasy cytologie růst a vývoj MeSH
- Rhodophyta cytologie růst a vývoj MeSH
- teplota MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Extensive in vivo replacement of hydrogen by deuterium, a stable isotope of hydrogen, induces a distinct stress response, reduces cell growth and impairs cell division in various organisms. Microalgae, including Chlamydomonas reinhardtii, a well-established model organism in cell cycle studies, are no exception. Chlamydomonas reinhardtii, a green unicellular alga of the Chlorophyceae class, divides by multiple fission, grows autotrophically and can be synchronized by alternating light/dark regimes; this makes it a model of first choice to discriminate the effect of deuterium on growth and/or division. Here, we investigate the effects of high doses of deuterium on cell cycle progression in C. reinhardtii. Synchronous cultures of C. reinhardtii were cultivated in growth medium containing 70 or 90% D2O. We characterize specific deuterium-induced shifts in attainment of commitment points during growth and/or division of C. reinhardtii, contradicting the role of the "sizer" in regulating the cell cycle. Consequently, impaired cell cycle progression in deuterated cultures causes (over)accumulation of starch and lipids, suggesting a promising potential for microalgae to produce deuterated organic compounds.
- Klíčová slova
- Chlamydomonas reinhardtii, cell cycle, cell division, commitment point, deuterium, heavy water, multiple fission, stress,
- MeSH
- buněčné dělení účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- Chlamydomonas reinhardtii růst a vývoj metabolismus MeSH
- deuterium škodlivé účinky chemie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- deuterium MeSH
An increase in temperature can have a profound effect on the cell cycle and cell division in green algae, whereas growth and the synthesis of energy storage compounds are less influenced. In Chlamydomonas reinhardtii, laboratory experiments have shown that exposure to a supraoptimal temperature (39 °C) causes a complete block of nuclear and cellular division accompanied by an increased accumulation of starch. In this work we explore the potential of supraoptimal temperature as a method to promote starch production in C. reinhardtii in a pilot-scale photobioreactor. The method was successfully applied and resulted in an almost 3-fold increase in the starch content of C. reinhardtii dry matter. Moreover, a maximum starch content at the supraoptimal temperature was reached within 1-2 days, compared with 5 days for the control culture at the optimal temperature (30 °C). Therefore, supraoptimal temperature treatment promotes rapid starch accumulation and suggests a viable alternative to other starch-inducing methods, such as nutrient depletion. Nevertheless, technical challenges, such as bioreactor design and light availability within the culture, still need to be dealt with.
- Klíčová slova
- Chlamydomonas reinhardtii, cell cycle, microalgae, pilot-scale production, starch, supraoptimal temperature,
- MeSH
- biomasa * MeSH
- bioreaktory MeSH
- buněčný cyklus MeSH
- Chlamydomonas reinhardtii metabolismus MeSH
- fotobioreaktory * MeSH
- kultivační média MeSH
- mikrořasy MeSH
- průmyslová mikrobiologie metody MeSH
- škrob metabolismus MeSH
- světlo MeSH
- teplota MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kultivační média MeSH
- škrob 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.
- Klíčová slova
- Chlamydomonas reinhardtii, DNA replication, cell cycle arrest, cell size, cyclin-dependent kinase, starch accumulation, supraoptimal temperature, synchronized cultures,
- MeSH
- bílkoviny řas metabolismus MeSH
- buněčné kultury metody MeSH
- Chlamydomonas reinhardtii cytologie fyziologie MeSH
- cyklin-dependentní kinasy metabolismus MeSH
- down regulace MeSH
- fyziologický stres MeSH
- kontrolní body buněčného cyklu * MeSH
- regulace genové exprese u rostlin MeSH
- vysoká teplota MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- bílkoviny řas MeSH
- cyklin-dependentní kinasy MeSH
Most cells divide into two daughter cells; however, some green algae can have different division patterns in which a single mother cell can sometimes give rise to up to thousands of daughter cells. Although such cell cycle patterns can be very complex, they are governed by the same general concepts as the most common binary fission. Moreover, cell cycle progression appears to be connected with size, since cells need to ensure that their size after division will not drop below the limit required for survival. Although the exact mechanism that lets cells measure cell size remains largely unknown, there have been several prominent hypotheses that try to explain it.
