Chlamydomonas Research Articles

Biological Activity of Chemical Constituents Isolated from Strain Chlamydomonas sp. KSF108 (Chlamydomonadaceae)

This study focused on investigation of the immunosuppressive inhibitory effect through determination of IL-2 production of nine compounds (1 - 9) isolated from Chlamydomonas sp. KSF108. Among them, compounds 1, 5, and 6 displayed moderately inhibitory effects on IL-2 production at a concentration of 100 μM. In addition, the related ones including cytotoxic, anti-inflammatory, and anti-oxidant activities were also elucidated. 6 further displayed cytotoxic activity against the MCF-7 cell line, with an IC50 value of 17.2 μM and 4, 6 - 7, and 9 possessed significant DPPH radical scavenging activity, with IC50 values ranging from 3.1 to 4.4 μM. To the best of our knowledge, this is the first report on the bioactivity of isolated chemical constituents from the genus Chlamydomonas. Compounds 1 and 5 investigated for the first time in the activity of immunosuppressivity and 6 may come to serve as the most important marker in broad-spectrum activities of the secondary metabolites identified from C. sp. KSF108.

CrABCA2 Facilitates Triacylglycerol Accumulation in Chlamydomonas reinhardtii under Nitrogen Starvation.

The microalga Chlamydomonas reinhardtii accumulates triacylglycerols (TAGs) in lipid droplets under stress conditions, such as nitrogen starvation. TAG biosynthesis occurs mainly at the endoplasmic reticulum (ER) and requires fatty acid (FA) substrates supplied from chloroplasts. How FAs are transferred from chloroplast to ER in microalgae was unknown. We previously reported that an Arabidopsis thaliana ATP-binding cassette (ABC) transporter, AtABCA9, facilitates FA transport at the ER during seed development. Here we identified a gene homologous to AtABCA9 in the C. reinhardtii genome, which we named CrABCA2. Under nitrogen deprivation conditions, CrABCA2 expression was upregulated, and the CrABCA2 protein level also increased. CrABCA2 knockdown lines accumulated less TAGs and CrABCA2 overexpression lines accumulated more TAGs than their untransformed parental lines. Transmission electron microscopy showed that CrABCA2 was localized in swollen ER. These results suggest that CrABCA2 transports substrates for TAG biosynthesis to the ER during nitrogen starvation. Our study provides a potential tool for increasing lipid production in microalgae.

The Cr(VI) Bioremediation Potential of Chlamydomonas Debaryana

Chromium (Cr) is an essential industrial component and has many applications in industrial manufacturing. Anthropogenic activities can lead to the release of hexavalent chromium [Cr(VI)] as well as trivalent chromium [Cr(III)]. Cr(VI) is 1,000 times more toxic than Cr(III), and very detrimental to most organisms even at low concentrations. Chlamydomonas debaryana, a freshwater algae, appeared to be Cr(VI) resistant and was able to survive in Cr(VI) concentrations of up to 80 mg/L for an extended period in packed aquifer media columns. This micro-green algae strain was isolated alongside Cr(VI) reducing bacteria (CRB) at a Cr(VI) contaminated site in South Africa. In this study, the growth response of C. debaryana in the presence of Cr(VI) was compared to that of Chlamydomonas reinhardtii, to assess the Cr(VI) tolerance of C. debaryana. At 50 mg/L Cr(VI) concentration the maximum chlorophyll a increase was 25.3 % for C. debaryana and 34.2 % for C. reinhardtii. However, after 3.5 d the chlorophyll a content decreased only 38.1 % for C. debaryana compared to a 100 % decrease for C. reinhardtii. Indicating that C. debaryana is only slightly more resistant than C. reinhardtii. Zero Cr(VI) removal was observed with the algae: C. debaryana could not reduce Cr(VI) to Cr(III) or adsorb Cr(VI) anions from the solution. C. debaryana as well as C. reinhardtii, algae cells are inhibited when directly exposed to Cr(VI) in continuously mixed batch reactors. C. debaryana might be better suited to use in cooperation with CRB for Cr(VI) treatment in a biofilm reactor. This research can lead to an improved treatment set up for Cr(VI) pollution that requires fewer chemical and energy inputs and produces less secondary waste.

Biological remediation of chromium (VI) in aquifer media columns

Hexavalent chromium (Cr(VI)) is highly soluble in water, but is widely used in industrial sectors for many purposes. The effluent from these industrial activities is often released to the environment posing a threat to aquatic life and to humans. Conventional methods of treating Cr(VI) often require the use of pump and treat methods followed by the use of harmful toxic chemicals that are hard to dispose of and usually are expensive. The study explores reduction of Cr(VI) to Cr(III) using biological means (a popular issue of bioremediation of contaminated aquifer) in columns; control with aquifer media, system 1 inoculated with CRB (chromium reducing bacteria) dried sludge, system 2 further amended with saw dust and system 3 amended with vegetative material from target site as carbon source. CRB are essential and were inoculated in the columns (system 1-3) as dried sludge, previously isolated (Enterococcus casseliflavus, Pseudomonas putida, C.istrobacter sedlakii, Enterobacter cloacae and Enterobacter hormaechei) and tested in a batch system. The column reactors were run for 60 days at concentrations 40, 60 and 80 mg/L. Culture isolates in the effluent of the reactors were isolated and identified. Complete reduction was observed from all columns under different concentrations, with some failures at certain periods before quasi steady state (determined after 40 d). At 40 mg/L more than 95 % of Cr(VI) was reduced across the spectrum. System 3 best reduced Cr(VI) compared to other treatment column reactors at high concentrations of 80 mg/L. System 2 carbon source didn’t enhance Cr(VI) reduction compared to Syetem 1 with no carbon material. Algae growth was observed in the columns after operating for 40 d at 40 mg/L. Using 18s rRNA the dominant algae was identified as Chlamydomonas debaryana. As Cr(VI) concentration was increased, both CRB and the algae were assumed to be in synergy as Cr(VI) was reduced at influent concentration as high as 80 mg/L (double the concentration from the site). Further investigation was done in the study to identify Cr(VI) reducing potential by algae. The use of vegetative material from the target site in the presence of algae proved to enhance Cr(VI) reduction by CRB even though saw dust did not perform as expected. This method has potential to be used in Cr polluted sites in South Africa with careful application.

Bioenergy production and nutrients removal by green microalgae with cultivation from agro-wastewater palm oil mill effluent (pome) - a review

Environmental pollution specifically wastewater is gaining attention both in the developed and developing countries. Malaysia is considered as one of the major palm oil producers in the world. Therefore, it is important to develop an environmental friendly and economic method to treat palm oil mill effluent (POME). The wastewater can serve as an economical nutrient source or substrate that can support the cultivation of microalgae. This can be a great nutrient for algal cultivation at the same time as remediating effluent and generating biomass. Nowadays, many microalgae species are being investigated to determine their potential and effectiveness for phytoremediation application, especially high growth rate. However, using synthetic media for growing microalgae in a mass scale is costly. It is acknowledged that POME (as nutrients enriched media) assisted enhanced microalgae growth under certain condition can considerably reduce the presence of organic and inorganic compounds. In this review, the potential of wide range of the predominant microalgae species with main focus on green microalgae (high removal efficiency): Chlamydomonas sp and Chlorella sp were investigated. Moreover, we discussed about the history, methods and future prospects in nutrients removal by green microalgae comprehensively. This review discusses several potential strategies for tackling the environmental issue generated by agro-waste water POME with enhancement of biomass productivity which can be used as an alternative for energy production.

Isolate new microalgal strain for biodiesel production and using ftir spectroscopy for assessment of pollutant removal from palm oil mill effluent (pome)

In tropical countries, the palm oil industry discharges a large amount of wastewater. The wastewater can serve as an economical nutrient source or substrate that can support the cultivation of microalgae. This study aimed to identify the local species of microalgae potentially existing in the industrial wastewater of palm oil mill effluent (POME). POME was selected as the key source of waste due to its higher potential in producing lipids from microalgae as biofuel substrate. A novel green microalgal strain was isolated from POME of Kahang- Johor west palm oil mill in Malaysia and was identified as Chlamydomonas sp. and subsequently named UTM 98 with Catalogue No. of KR349061. This study emphasised the effectiveness of POME as the main carbon source to maintain the growth of microalgae and simultaneously to increase the lipid content. In this study, Fourier Transform Infrared spectroscopy (FTIR) and Gas Chromatography (GC-FID) were used to identify andquantify lipids in the freshwater microalgae. Cultivation of microalgae were initially carried out in 250 mL Erlenmeyer flask containing 100 mL medium at ± 30 °C with continuous illumination (± 14 µmol-1 m-2 s-1) andup to 20 d of cultivations. Results demonstrated that on the chromatogram, the highest retention achieved is belong to palmitic acid (C16:0). Chlamydomonas incerta (C. incerta) species is found to contain shorter chain fatty acids, mainly 16 - 18 carbon length, which is ideal for biodiesel production. FTIR spectrum of POME treated biomass displayed the shifting of peak at 591 cm-1 and also removal of C-Cl stretching. The spectrum of POME effluent treated biomass revealed broad peak at 3,430 cm-1. The results of SEM micrographs showed that, after treating POME with C. incerta, the cells became slightly rough and corrugated textures and some particles were found on the surface of the cell wall. Using POME as a rich carbon and nutrient source is also a promising approach either as natural environment treatment or as high-lipid-content raw material for production of biofuel.

Growth of Chlamydomonas reinhardtii under Circadian Conditions

Growth of Chlamydomonas reinhardtii under Circadian Conditions

Expression and Optimization Strategy of Recombinant Proteins in Chlamydomonas Chloroplast

Expression and Optimization Strategy of Recombinant Proteins in Chlamydomonas Chloroplast

Ingestión y digestión de siete especies de microalgas por larvas de Strombus gigas (Mesogastropoda: Strombidae)

The potential nutritional value of seven microalgal diets as measured by their ingestibility and digestibility to queen conch Strombus gigas larvae was tested with 30 day old larvae reared at 28°C and fed at 1000 cells ml-1. The algae were Tetraselmis suecica, Tetraselmis chuii, Isochrysis aff. galbana, Dunaliella ter­tiolecta, Chlamydomonas coccoides, Chaetoceros sp. and Thalassiosira fluviatilis. Ingestion and digestion were measured by the four nutritional stages studied with epifluorescence microscopy with live larvae. Temporal and absolute indices showed that arvae fed Chaetoceros sp. and T. fluviatilis had lower ingestion and digestion levels. The other algae are recommend to feed S. gigas larvae.

Extraction, Optimization and Antimicrobial Activity of IWSP from Oleaginous Microalgae <i>Chlamydomonas</i> sp. YB-204

Extraction, Optimization and Antimicrobial Activity of IWSP from Oleaginous Microalgae <i>Chlamydomonas</i> sp. YB-204

Protein Phosphatase 2CInvolved in the Flagellar Transition Zone Structure of Chlamydomonas reinhardtii

Protein Phosphatase 2CInvolved in the Flagellar Transition Zone Structure of Chlamydomonas reinhardtii

2B35 クラミドモナスの分布に及ぼすエアレーションの影響(OS1-2:自然界のバイオメカニクス・バイオミメティクス(2))

2B35 クラミドモナスの分布に及ぼすエアレーションの影響(OS1-2:自然界のバイオメカニクス・バイオミメティクス(2))

미세조류 유래 Shinorine의 피부세포에서의 효능

오존층의 파괴와 증가하는 야외활동으로 인한 자외선의 노출을 최소화 하기위해 자외선 차단제는 필수적인 것이 되고 있다. 기존에 사용되는 자외선 차단제의 기능을 향상시키며 부작용을 줄이기 위해 천연식물과 같은 친환경적인 소재의 개발이 필요한 실정이다. 본 연구는 천연물을 활용한 자외선차단제 및 기능성 화장품 소재 개발을 위해 미세조류 (Chlamydomonas hedleyi.)로부터 Mycosporine-like amino acid (MAA)를 분리 정제하고 그 효능을 평가하고자 하였다. 미 세조류에서 분리 정제한 MAAs 중 shinorine을 분리하여 UV에 의한 피부세포의 손상을 막고, 염증과 관련 있는 단백질의 발현을 저해하는지를 평가하였다. 그 결과 shinorine이 피부세포가 UV에 의해 손상되는 것을 보호하여 cell viability가 10% 증가하였고, 염증관련 단백질인 COX2이 발현이 41%정도 감소하는 것을 확인하였다. 또한 콜라겐의 합성을 증가시켜 주름개 선 효능을 확인하였다. 이에 shinorine은 자외선 차단용 소재뿐 아니라 항노화 관련 제품의 소재로 가능성을 확인하였다.

Volvox, Rolling out from under the Shadow of Chlamydomonas with Support from the AGA

Volvox, Rolling out from under the Shadow of Chlamydomonas with Support from the AGA

Package and identification of replication deficient recombinant adenovirus expression vector of channelrhodopsin-2

Background Channelrhodopsin-2 (ChR2)is a cation channel isolated from the eyespot of Chlamydomonas algae and has been used to control neuron activity.The light stimulation is a more precise fashion whether space or time than that of electrical,magnetic and ultrasound stimulation. Objective This study was to construct a replication deficient recombinant adenovirus cxpression vector of ChR2 and to determine its function.Methods Human embryo kidney 293 (HEK293) cell line was cultured and passaged in DF12 medium containing 10％ fetal bovine serum(FBS).The ChR2 gene was cloned at the downstream of cytomegalovirus(CMV)promoter of the adenoviral shuttle plasmid pSB291 in sense direction,and the resultant recombinant plasmid pSB291-hChR2- GFP was transfected into HEK293 cell together with plasmid pBHG lox ( deltaE1,3 ) containing adenoviral genome,then small amounts replication deficient recombinant adenovirus expression vector of ChR2 (Ad-ChR2) was obtained.Through amplification gradient centrifugation and dialysis,pure Ad-ChR2 was obtained.Visual cortex cells derived from 4 1-day-old clean Long Evans rats were primary cultured with serum-free culture media and infected by AdChR2.When expressing green fluorescencc,those cells received the stimulated of blue light with 460 nm.Patch clamp technique was applied to record an action potential. Results After purification and concentration,the titer of AdhCHR2 reached 7.9×1010 PFU/ml.Twenty-four hours after transfect of Ad-ChR2,HEK293 cell membrane showed the green fluorescence for the recombinant plasmid with green fluorescence protein under the inversed fluorescence microscope.The HEK293 cells change their shape from flat to round 13 days after transfected.The primary cultured visual cortex cells exhibited the green fluorescence 3-5 days after infected by Ad-ChR2.The action potentials evoked by blue light stimulation were recorded with patch clamp on those cells expressing green fluorescence. Conclusions Ad-ChR2 expressing vector is constructed successfully in this study.It is verified that Ad-ChR2 expressing vector can infect visual cortex cells with visual function.This result is very important for visual plasticity study. Key words: Channelrhodopsin-2; Adenovirus; Patch clamp

The effects of nutritional restriction on neutral lipid accumulation in Chlamydomonas and Chlorella

Under nutrient starvation conditions, many microalgae are known to accumulate triacylglycerols (TAG) that can be used for biodiesel production. However, few studies have been performed to analyze the effect of deficiency in nutrient elements such as sulfur, phosphorus, potassium, iron, magnesium and calcium on oil production, particularly in Chlamydomonas and Chlorella. In this study, we investigated lipid content of Chlamydomonas reinhaditti CC124 and Chlorella vulgaris Y-019 grown in TAP, HSM, BG11, SE lacking optimal concentrations of these elements. Our results showed that, in high carbon HSM and TAP media, N and S starvation led to significant increase in cellular lipid content in both microalgae species. In addition, C. reinhaditti grown in TAP media without P, Fe, K, Ca or Mg or in HSM media without K, Ca or Mg also accumulated detectably higher neutral lipids. In contrast, in C. vulgaris, such accumulation was observed only in Mg-free and Fe-free HSM media. In low carbon SE and BG11 media, N starvation resulted in a moderate increase in the lipids content both in C. reinhaditti and C. vulgaris. On the other hand, P, S, K, Ca or Mg deficiency promoted neutral lipids accumulation in C. vulgaris. Finally, we analyzed and discussed the relationships among cell growth rate, lipid accumulation and nitrogen concentrations in C. reinhaditti. Key words: Nutrient limitation, lipid accumulation, nitrogen, sulfur, phosphorus, potassium, iron, magnesium, calcium.

The adaptability of three Arctic microalgae to different low temperatures

In order to study the adaptability of Arctic microalgae to different environmental temperatures, the growth curves and antioxidase system of three microalgae ( Skeletonema marinoi , Chlorella  sp. and Chlamydomonas sp.) that were separated from the Ny-Alesund, the high Arctic, at different low temperatures (0℃, 4℃ and 8℃) were determined. The result showed that the adaptability of the microalgae to temperatures depended on the species. The growth rate, SOD and CAT activities of Skeletonema marinoi were the highest at 4℃, but MDA content was the lowest. The growth rate and enzyme activity of Chlorella sp. were the highest at 8℃, while the lowest MDA content presented at 0℃. The growth of Chlamydomonas sp. at the different temperatures was not so significant, the lowest MDA content presented at 8℃. The change of antioxidase system also depended on species and temperatures. Three indexes of antioxidase system of Skeletonema marinoi between 0℃ and 4℃ showed extremely significant difference (p Skeletonema marinoi and Chlorella sp.between 0℃ and 8℃ showed significant difference (p Chlamydomonas sp. at the three temperatures differed insignificantly. In conclusion, the three microalgae had good adaptability to the three temperatures; their MDA content presented a low level, and had unique physiological mechanism to adapt to the environment with different low temperatures.

Evidence of monomeric photosystem I complexes and phosphorylation of chlorophyll a/c-binding polypeptides in Chroomonas sp. strain LT (Cryptophyceae).

Thylakoid membranes of the cryptophyte Chroomonas sp. strain LT were solubilized with dodecyl-beta-maltoside and subjected to sucrose density gradient centrifugation. The four pigment protein complexes obtained were subsequently characterized by absorption and fluorescence spectroscopy, SDS-PAGE, and Western immunoblotting using antisera against the chlorophyll a/c-binding proteins of the marine cryptophyte Cryptomonas maculata and the reaction-center protein D2 of photosystem II of maize. Band 1 consisted mainly of free pigments, phycobiliproteins, and chlorophyll-a/c-binding proteins. Band 2 represented a major chlorophyll a/c-binding protein fraction. A mixture of photosystem II and photosystem I proteins comprised band 3, whereas band 4 was enriched in proteins of photosystem I. Western immunoblotting demonstrated the presence of chlorophyll a/c-binding proteins and their association with photosystem I in band 4. Phosphorylation experiments showed that chlorophyll a/c-binding proteins became phosphorylated. Negative staining electron microscopy of band B4 revealed photosystem I particles with dimensions of 22 nm. Our work showed that PSI-LHCI complexes of cryptophytes are similar to those of Chlamydomonas rheinhardtii, the diatom Phaeodactylum tricornutum, and higher plants.

A molecular characterisation of PsaN: a nuclear-encoded subunit of the photosystem I complex of Chlamydomonas reinhardtii.

Photosystem I (PSI) is a membrane protein complex composed of a large number of polypeptide subunits, designated PsaA to PsaN. In eukaryotes, the genes for these subunits are distributed between the nuclear and chloroplast genomes with the core subunits encoded by chloroplast genes. The function of many of the small nuclear-encoded subunits is unknown. Primarily, we are investigating the role of a nuclear-encoded subunit PsaN, which is absent in prokaryotic PSI complexes. PsaN is an extrinsic subunit located entirely on the lumenal side of the thylakoid membrane. We have isolated and sequenced the psaN gene in Chlamydomonas and shown that it is a single copy gene with an unusual exon structure. We are currently generating nuclear PSI mutants by random mutagenesis, in order to isolate tagged Chlamydomonas mutants disrupted in psaN or the other nuclear genes. The isolation of such mutants will provide an insight into their function and a valuable null background for structure-function studies, in which site-directed variants of the gene are re-introduced into the nuclear genome. In addition, we are analysing an existing psaF-deficient mutant of Chlamydomonas reinhardtii [1] in order to determine the function of the C-terminal region of the PsaF subunit. [1] Farah, J. et al. (1995). EMBO J. 14: 4976-4984.

Redox regulation of PSI nuclear genes through photosynthetic electron flow

Light is one of the most important environmental cues that regulate photosynthesis genes. Recently, significance of the redox regulatory system has been suggested in the light regulation of photosynthesis genes, in cyanobacteria, eukaryotic algae, and also in higher plants. In this study, we examined how PSI nuclear genes of Chlamydomonas reinhardtii are regulated in response to light. When Chlamydomonas cells were transferred from the darkness to the light, psaE mRNA level was drastically increased. This light induction was inhibited by the photosynthetic electron transport inhibitors, DCMU and DBMIB, but not by an uncoupler, CCCP. These demonstrate that, in this induction, photosynthetic electron flow plays a pivotal role but photophosphorylation is not necessary. Subsequently, we examined whether redox state of intersystem electron carrier is responsible for this induction. For this sake, we irradiated the Chlamydomonas cells with PSI-light (695nm) or PSII-light (644nm), which makes plastoquinone pool oxidative and reductive, respectively. However, both monochromes induced the psaE mRNA to similar extent, indicating that the redox state of intersystem electron carriers, e.g. plastoquinone pool, is not responsible for this light induction. We are currently examining how the photosynthesis-derived signal causes this response, and several hints obtained by our recent experiments will be discussed.