Identification and characterisation of a novel inorganic carbon acquisition gene, CIA7, from an insertional mutant of Chlamydomonas reinhardtii

2008 ◽  
Vol 35 (5) ◽  
pp. 373 ◽  
Author(s):  
Ruby A. Ynalvez ◽  
James V. Moroney
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Metal Ion ◽  
Inorganic Carbon ◽  
Antibiotic Resistance Gene ◽  
Inverse Pcr ◽  
Chloroplast Targeting ◽  
Calculated Molecular Mass ◽  
Prediction Program ◽  
Metal Transporter ◽  
Low Co2

Chlamydomonas reinhardtii is a unicellular eukaryotic alga which possesses a CO2-concentrating mechanism (CCM) that enables it to grow at low CO2 concentrations. Previously, insertional mutants were generated to enable isolation of inorganic carbon transporters and other proteins that might be essential for a functional CCM. These mutants have an antibiotic resistance gene that encodes a protein that binds to Zeocin inhibiting Zeocin’s DNA strand cleavage activity. The DNA flanking the BleR insert of one of the high CO2 requiring strains, named cia7, was cloned with inverse-PCR and sequenced. Sequence analysis showed homology to conserved bacterial proteins of unknown function, but there were no ESTs in this region of the genome. However, the presence of a gene was established by PCR and RLM-RACE. CIA7 was found to have four exons and the BleR insert was in the fourth exon. CIA7 encodes a protein of 104 amino acids with a calculated molecular mass of 11.3 kDa. Based on the ChloroP prediction program, the protein is predicted to have a chloroplast targeting signal. Complementation analyses results showed possible partially rescued mutants, and RNAi showed several transformants with a sick on low CO2 phenotype with reduced expression of CIA7. These results suggest that CIA7 is a gene that facilitates growth in C. reinhardtii under low CO2 conditions. One possible role of CIA7 would be in the delivery or storage of a metal ion. It may play a potential role as either a domain of a metal transporter or as a metallochaperone.

A mutant of Chlamydomonas reinhardtii that cannot acclimate to low CO2 conditions has an insertion in the Hdh1 gene

2005 ◽  
Vol 32 (1) ◽  
pp. 55 ◽  
Author(s):  
James E. Adams ◽  
Sergio L. Colombo ◽  
Catherine B. Mason ◽  
Ruby A. Ynalvez ◽  
Baran Tural ◽  
...  
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Inorganic Carbon ◽  
Single Copy ◽  
Wild Type ◽  
Poor Growth ◽  
Altered Expression ◽  
Haloacid Dehalogenase ◽  
Low Co2 ◽  
Photosynthetic Microorganisms ◽  
Zeocin Resistance

Photosynthetic microorganisms must acclimate to environmental conditions, such as low CO2 environments or high light intensities, which may lead to photo-oxidative stress. In an effort to understand how photosynthetic microorganisms acclimate to these conditions, Chlamydomonas reinhardtii was transformed using the BleR cassette, selected for Zeocin resistance and screened for colonies that showed poor growth at low CO2 levels. One of the insertional mutants obtained, named slc-230, was shown to have a BleR insert in the first exon of Hdh1, a novel, single copy gene. The predicted Hdh1 gene product has similarity to bacterial haloacid dehalogenase-like proteins, a protein family that includes phosphatases and epoxide hydrolases. In addition, Hdh1 is predicted to be localised to the chloroplast or mitochondria in C. reinhardtii. It was found that a genomic copy of wild type Hdh1 can complement slc-230. Physiological studies were conducted to determine the effects of the altered expression of Hdh1 in slc-230. slc-230 grows slowly autotrophically in low CO2, exhibits a lower affinity for inorganic carbon, a decreasing photosynthetic rate over time and a lower content of chlorophylls and quenching xanthophylls than wild type cells. Some possible roles of Hdh1 in the acclimation to low CO2 conditions are discussed.

scholarly journals Expression of a Low CO2–Inducible Protein, LCI1, Increases Inorganic Carbon Uptake in the Green Alga Chlamydomonas reinhardtii

2010 ◽  
Vol 22 (9) ◽  
pp. 3105-3117 ◽  
Author(s):  
Norikazu Ohnishi ◽  
Bratati Mukherjee ◽  
Tomoki Tsujikawa ◽  
Mari Yanase ◽  
Hirobumi Nakano ◽  
...  
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Green Alga ◽  
Inorganic Carbon ◽  
Carbon Uptake ◽  
Low Co2 ◽  
Inducible Protein ◽  
Inorganic Carbon Uptake

Use of the bleomycin resistance gene to generate tagged insertional mutants of Chlamydomonas reinhardtii that require elevated CO2 for optimal growth

2002 ◽  
Vol 29 (3) ◽  
pp. 231 ◽  
Author(s):  
Sergio L. Colombo ◽  
Steve V. Pollock ◽  
Karla A. Eger ◽  
Ashley C. Godfrey ◽  
James E. Adams ◽  
...  
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Elevated Co2 ◽  
Insertional Mutagenesis ◽  
Optimal Growth ◽  
Inverse Pcr ◽  
Transformation Protocol ◽  
Low Co2 ◽  
Co2 Concentrations ◽  
Polymerase Chain ◽  
Tail Pcr

Chlamydomonas reinhardtii Dangeard possesses a CO2 concentrating mechanism (CCM) that enables it to grow at very low CO2 concentrations. In previous studies, insertional mutagenesis was successfully used to identify genes required for growth at low CO2 in C. reinhardtii. These earlier studies used the C. reinhardtii genes, Nit1 and Arg7 to complement nit1– or arg7– strains, thereby randomly inserting a second copy of Nit1 or Arg7 into the genome. Because these genes are already present in the C. reinhardtii genome, it was often difficult to identify the location of the inserted DNA and the gene disrupted by the insertion. We have developed a transformation protocol using the BleR gene, which confers resistance to the antibiotic Zeocin. The insertion of this gene allows one to use a variety of existing polymerase chain reaction (PCR) methodologies to identify the disrupted gene. In this study the D66 strain (nit2–, cw15, mt+) was transformed by electroporation using a plasmid containing the BleR gene. Primary transformants (42 000) were obtained after growth in the dark on acetate plus Zeocin medium. Colonies were then tested for their ability to grow photosynthetically on elevated CO2 or low levels of CO2 (100 ppm). About 120 mutants were identified which grew on elevated CO2 but were unable to grow well at low CO2 concentrations. About 50% of these mutants had low affinities for inorganic carbon as assessed by K0.5(CO2), indicating a potential defect in the CCM. The location of the inserted DNA is being determined using inverse PCR (iPCR) and thermal asymmetric interlaced (TAIL) PCR. Using these methods, one can rapidly locate the inserted DNA in the genome and identify the gene that has been disrupted by the insertion.

scholarly journals Rubisco Activase Is Required for Optimal Photosynthesis in the Green Alga Chlamydomonas reinhardtii in a Low-CO2 Atmosphere

2003 ◽  
Vol 133 (4) ◽  
pp. 1854-1861 ◽  
Author(s):  
Steve V. Pollock ◽  
Sergio L. Colombo ◽  
Davey L. Prout ◽  
Ashley C. Godfrey ◽  
James V. Moroney
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Green Alga ◽  
Rubisco Activase ◽  
Low Co2 ◽  
Co2 Atmosphere

Properties of Selenium-Induced Glutathione Peroxidase in Low-CO2-Grown Chlamydomonas reinhardtii

1990 ◽  
pp. 3409-3412 ◽  
Author(s):  
Shigeru Shigeoka ◽  
Toru Takeda ◽  
Tsuyoshi Hanaoka ◽  
Akiho Yokota ◽  
Shozaburo Kitaoka ◽  
...  
Keyword(s):  
Glutathione Peroxidase ◽  
Chlamydomonas Reinhardtii ◽  
Low Co2

Photosynthetic utilisation of inorganic carbon and its regulation in the marine diatom Skeletonema costatum

2004 ◽  
Vol 31 (10) ◽  
pp. 1027 ◽  
Author(s):  
Xiongwen Chen ◽  
Kunshan Gao
Keyword(s):  
Inorganic Carbon ◽  
Dissolved Inorganic Carbon ◽  
Skeletonema Costatum ◽  
Co2 Concentration ◽  
Marine Diatom ◽  
Disulfonic Acid ◽  
High Co2 ◽  
Low Co2 ◽  
Diatom Skeletonema Costatum ◽  
Very High

Photosynthetic uptake of inorganic carbon and regulation of photosynthetic CO2 affinity were investigated in Skeletonema costatum (Grev.) Cleve. The pH independence of K1/2(CO2) values indicated that algae grown at either ambient (12 μmol L–1) or low (3 μmol L–1) CO2 predominantly took up CO2 from the medium. The lower pH compensation point (9.12) and insensitivity of photosynthetic rate to di-isothiocyanatostilbene disulfonic acid (DIDS) indicated that the alga had poor capacity for direct HCO3– utilisation. Photosynthetic CO2 affinity is regulated by the concentration of CO2 rather than HCO3–, CO32– or total dissolved inorganic carbon (DIC) in the medium. The response of photosynthetic CO2 affinity to changes in CO2 concentration was most sensitive within the range 3–48 μmol L–1 CO2. Light was required for the induction of photosynthetic CO2 affinity, but not for its repression, when cells were shifted between high (126 μmol L–1) and ambient (12 μmol L–1) CO2. The time needed for cells grown at high CO2 (126 μmol L–1) to fully develop photosynthetic CO2 affinity at ambient CO2 was approximately 2 h, but acclimation to low or very low CO2 levels (3 and 1.3 μmol L–1, respectively) took more than 10 h. Cells grown at low CO2 (3 μmol L–1) required approximately 10 h for repression of all photosynthetic CO2 affinity when transferred to ambient or high CO2 (12 or 126 μmol L–1, respectively), and more than 10 h at very high CO2 (392 μmol L–1).

The metal transporter CrNRAMP1 is involved in zinc and cobalt transports in Chlamydomonas reinhardtii

2020 ◽  
Vol 523 (4) ◽  
pp. 880-886 ◽  
Author(s):  
Peng Chang ◽  
Hua Yin ◽  
Tadayuki Imanaka ◽  
Yasuo Igarashi ◽  
Nannan Li ◽  
...  
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Metal Transporter
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