Toshio Sakamoto - Academia.edu (original) (raw)

Papers by Toshio Sakamoto

Plant Physiology, 1997

Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and t... more Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and the main site of the damage is the D1 protein of the photosystem II (PSII) complex. Here we describe that transformation of Synechococcus sp. PCC 7942 with the desA gene for a [delta]12 desaturase increased unsaturation of membrane lipids and enhanced tolerance to strong light. To our knowledge, this is the first report of the successful genetic enhancement of tolerance to strong light. Analysis of the light-induced inactivation and of the subsequent recovery of the activity of the PSII complex revealed that the recovery process was markedly accelerated by the genetic transformation. Labeling experiments with [35S]L-methionine also revealed that the synthesis of the D1 protein de novo at low temperature, which was a prerequisite for the restoration of the PSII complex, was much faster in the transformed cells than in the wild-type cells. These findings demonstrate that the ability of memb...

Plant and Cell Physiology, Mar 1, 1995

Plant and Cell Physiology, Mar 1, 1996

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the ω3 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the ω3 desaturase

Plant and Cell Physiology, 1997

A cyaC gene encoding an adenylate cyclase of the filamentous cyanobacterium Spirulina platensis w... more A cyaC gene encoding an adenylate cyclase of the filamentous cyanobacterium Spirulina platensis was sequenced. The predicted amino acid sequence of the C-terminal region of cyaC is similar to the catalytic domains of adenylate cyclases in other cyanobacteria and eukaryotes. The sequences of other regions are similar to those of proteins consisting of the bacterial two-component signal transduction system: the sensory kinase and the response regulator. The predicted gene product of cyaC contains, from the N-terminal end, a receiver domain of the response regulator protein (Rl), a domain similar to the ETR1 of Arabidopsis thaliana, a transmitter domain of the sensory kinase protein, a receiver domain of the response regulator protein (R2), and a catalytic domain of adenylate cyclase. The cyaC gene was expressed as an affinity-tagged protein in Escherichia coli, and the recombinant protein was purified. The purified protein had adenylate cyclase activity which was activated by Mn 2+. The results of Western blotting using an anti-CyaC antiserum and the S. platensis cell extract confirmed that cyaC gene is expressed in S. platensis.

Current Opinion in Microbiology, 2002

The expression of cold-inducible genes is regulated by a two-component system in Synechocystis an... more The expression of cold-inducible genes is regulated by a two-component system in Synechocystis and Bacillus subtilis. The cold sensors are membrane-bound histidine kinases and it seems likely that they sense and transduce changes in the fluidity of membranes. Desaturation of fatty acids in membrane lipids has been implicated in tolerance to cold and salt stress.

Biochemical Journal, 2006

Acyl-lipid desaturases are enzymes that convert a C–C single bond into a C=C double bond in fatty... more Acyl-lipid desaturases are enzymes that convert a C–C single bond into a C=C double bond in fatty acids that are esterified to membrane-bound glycerolipids. Four types of acyl-lipid desaturase, namely DesA, DesB, DesC, and DesD, acting at the Δ12, Δ15, Δ9, and Δ6 positions of fatty acids respectively, have been characterized in cyanobacteria. These enzymes are specific for fatty acids bound to the sn−1 position of glycerolipids. In the present study, we have cloned two putative genes for a Δ9 desaturase, designated desC1 and desC2, from Nostoc species. The desC1 gene is highly similar to the desC gene that encodes a Δ9 desaturase that acts on C18 fatty acids at the sn−1 position. Homologues of desC2 are found in genomes of cyanobacterial species in which Δ9-desaturated fatty acids are esterified to the sn−2 position. Heterologous expression of the desC2 gene in Synechocystis sp. PCC 6803, in which a saturated fatty acid is found at the sn−2 position, revealed that DesC2 could desatu...

Archives of Microbiology, 1997

Cyanobacteria acclimate to low temperature by desaturating their membrane lipids. Mutant strains ... more Cyanobacteria acclimate to low temperature by desaturating their membrane lipids. Mutant strains of Synechococcus sp. PCC 7002 containing insertionally inactivated desA (Delta12 acyl-lipid desaturase) and desB (omega3 acyl-lipid desaturase) genes were produced, and their low-temperature susceptibility was characterized. The desA mutant synthesized no linoleic acid or alpha-linolenic acid, and the desB mutant did not produce alpha-linolenic acid. The desA mutant grew more slowly than the wild-type at 22 degrees C and could not grow at 15 degrees C. The desB mutant could not continuously grow at 15 degrees C, although no observable phenotype appeared at higher temperatures. It has been shown that expression of the desA gene occurs at 38 degrees C and is up-regulated at 22 degrees C, and that the desB gene is only expressed at 22 degrees C. These results indicate that the expression of the desA and desB genes occurs at higher temperatures than those at which a significant decline in physiological activities is caused by the absence of their products. The temperature dependency of photosynthesis was not affected by these mutations. Since chlorosis and inability to grow at 15 degrees C with nitrate was suppressed by the substitution of urea as a nitrogen source, it is very likely that the chilling susceptibility of the desaturase mutants is attributable to nutrient limitation.

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the 093 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the o93 desaturase from the cyanobacterium, Synechocystis sp. PCC 6803. The desB gene encodes a protein a 359 amino-acid residues with molecular mass of 41.9 kDa. The desB gene is transcribed as a monocistronic operon that produced a single transcript of 1.4 kb. The level of the desB transcript in cells grown at 22 °C was 10 times higher than that in cells grown at 34 ° C. In order to manipulate the fatty-acid unsaturation of membrane lipids, the desB gene in Synechocystis sp. PCC 6803 was mutated by insertion of a kanamycin-resistance gene cartridge. The resultant mutant was unable to desaturate fatty acids at the 093 position. The desA gene, which encodes the A12 desaturase of Synechocystis sp. PCC 6803, and the desB gene were introduced into Synechococcus sp. PCC 7942. Whilst the parent cyanobacterium can only desaturate membrane lipids at the A9 position of fatty acids, the resultant transformant was able to desaturate fatty acids of membrane lipids at the A9, A12 and o93 positions. These results confirm the function of the desB gene and demonstrate that it is possible to genetically manipulate the fatty-acid unsaturation of membrane lipids in cyanobacteria.

Plant physiology

Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and t... more Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and the main site of the damage is the D1 protein of the photosystem II (PSII) complex. Here we describe that transformation of Synechococcus sp. PCC 7942 with the desA gene for a A12 desaturase increased unsaturation of membrane lipids and enhanced tolerance to strong light. To our knowledge, this is the first report of the successful genetic enhancement of tolerance to strong light.

Applied and environmental microbiology, 1998

Laboratory conditions have been identified that cause the rapid death of cultures of cyanobacteri... more Laboratory conditions have been identified that cause the rapid death of cultures of cyanobacteria producing urease. Once the death phase had initiated in the stationary growth phase, cells were rapidly bleached of all pigmentation. Null mutations in the ureC gene, encoding the alpha subunit of urease, were constructed, and these mutants were no longer sensitive to growth in the presence of urea. High levels of peroxides, including lipid peroxides, were detected in the bleaching cells. Exogenously added polyunsaturated fatty acids triggered a similar death response. Vitamin E suppressed the formation of peroxides and delayed the onset of cell bleaching. The results suggest that these cyanobacterial cells undergo a metabolic imbalance that ultimately leads to oxidative stress and lipid peroxide formation. These observations may provide insights into the mechanism of sudden cyanobacterial bloom disappearance in nature.

Photosynthesis research, 2002

The effect of polyunsaturated fatty acids on photosynthesis and the growth of the marine cyanobac... more The effect of polyunsaturated fatty acids on photosynthesis and the growth of the marine cyanobacterium Synechococcus sp. PCC 7002 was examined using wild-type and Delta12 fatty acid desaturase mutant strains. Under a light intensity of 250 mumol m(-2) s(-1), wild-type cells could grow exponentially in a temperature range of 20-38 degrees C, but growth was non-exponential below 20 degrees C and ceased at 12 degrees C. The Delta12 desaturase mutant cells lacking polyunsaturated fatty acids had the same growth rate as wild-type cells in a temperature range of 25-38 degrees C but grew slowly at 22 degrees C, and no cell growth took place below 18 degrees C. Under a very high-light intensity of 2.5 mmol m(-2) s(-1), wild-type cells could grow exponentially in a temperature range of 30-38 degrees C, although the high-light grown cells became chlorotic because of nitrogen limitation. The temperature sensitive phenotype in the Delta12 desaturase mutant was enhanced in cells grown under hig...

Plant Molecular Biology, 1994

Cyanobacterial genes for enzymes that desaturate fatty acids at the A12 position, designated desA... more Cyanobacterial genes for enzymes that desaturate fatty acids at the A12 position, designated desA, were isolated from Synechocystis PCC6714, Synechococcus PCC7002 and Anabaena variabilis by crosshybridization with a DNA probe derived from the desA gene of Synechocystis PCC6803. The genes of Synechocystis PCC6714, Synechococcus PCC7002 and A. variabilis encode proteins of 349, 347 and 350 amino acid residues, respectively. The transformation of Synechococcus PCC7942 with the desA genes from Synechocystis PCC6714, Synechococcus PCC7002 and A. variabilis was associated with the ability to introduce a second double bond at the A12 position of fatty acids. The amino acid sequence of the products of the desA genes revealed the presence of four conserved domains. Since one of the conserved domains was also found in the amino acid sequences of o93 desaturases ofBrassica napus and mung bean, this domain may play an essential role in the introduction of a double bond into fatty acids bound to membrane lipids.

Plant Molecular Biology, 1996

A cyaA gene, encoding an adenylate cyclase, was isolated from a filamentous cyanobacterium, Spiru... more A cyaA gene, encoding an adenylate cyclase, was isolated from a filamentous cyanobacterium, Spirulina platensis, by functional complementation of a cya mutant of Escherichia coli, defective in adenylate cyclase activity. The predicted gene product of cyaA contains a signal peptide-like domain, a putative sensor domain similar to the gene product of vsrA of Pseudomonas solanacearum, a putative membrane-spanning domain and an adenylate cyclase-like catalytic domain. Two other positive clones that complemented the E. coli mutant were isolated from the same cyanobacterium, suggesting that several cya genes are functioning in S. platensis.

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the ω3 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the ω3 desaturase from the cyanobacterium, Synechocystis sp. PCC 6803. The desB gene encodes a protein a 359 amino-acid residues with molecular mass of 41.9 kDa. The desB gene is transcribed as a monocistronic operon that produced a single transcript of 1.4 kb. The level of the desB transcript in cells grown at 22°C was 10 times higher than that in cells grown at 34°C. In order to manipulate the fatty-acid unsaturation of membrane lipids, the desB gene in Synechocystis sp. PCC 6803 was mutated by insertion of a kanamycin-resistance gene cartridge. The resultant mutant was unable to desaturate fatty acids at the ω3 position. The desA gene, which encodes the Δ12 desaturase of Synechocystis sp. PCC 6803, and the desB gene were introduced into Synechococcus sp. PCC 7942. Whilst the parent cyanobacterium can only desaturate membrane lipids at the Δ9 position of fatty acids, the resultant transformant was able to desaturate fatty acids of membrane lipids at the Δ9, Δ12 and ω3 positions. These results confirm the function of the desB gene and demonstrate that it is possible to genetically manipulate the fatty-acid unsaturation of membrane lipids in cyanobacteria.

Phycological Research, 2009

Changes in photosynthetic activity and trehalose levels in field-isolated, natural colonies of th... more Changes in photosynthetic activity and trehalose levels in field-isolated, natural colonies of the terrestrial cyanobacterium Nostoc commune responding to desiccation and salt stress were investigated. As the water content decreased in N. commune colonies during desiccation, photosynthetic O2-evolving activity decreased and no activity was detected in desiccated colonies. A high level of O2 evolution was restored in the colonies as they absorbed atmospheric moisture, indicating that only a small amount of water is required for reactivation of photosynthesis. No detectable trehalose was found in fully hydrated N. commune colonies; however, trehalose accumulation occurred in response to water loss during desiccation and high levels of trehalose were detected in the airdried colonies. Moreover, a 0.2 M NaCl treatment also induced trehalose accumulation to a level equivalent to that by desiccation. Photosynthetic O2 evolution was inhibited by 0.2 M NaCl, indicating that N. commune can tolerate only low levels of salt. These results suggest that cessation of photosynthesis and trehalose accumulation occur in response to both matric water stress (desiccation) and osmotic water stress (high salt concentration), and that while trehalose may be a less effective osmoprotective compound than others, it is important for the extreme tolerance to desiccation observed in terrestrial cyanobacterium.

Photosynthesis Research, 2007

Ferredoxin-NADP(+) oxidoreductase (FNR) catalyzing the terminal step of the linear photosynthetic... more Ferredoxin-NADP(+) oxidoreductase (FNR) catalyzing the terminal step of the linear photosynthetic electron transport was purified from the cyanobacterium Spirulina platensis and the red alga Cyanidium caldarium. FNR of Spirulina consisted of three domains (CpcD-like domain, FAD-binding domain, and NADP(+)-binding domain) with a molecular mass of 46 kDa and was localized in either phycobilisomes or thylakoid membranes. The membrane-bound FNR with 46 kDa was solublized by NaCl and the solublized FNR had an apparent molecular mass of 90 kDa. FNR of Cyanidium consisted of two domains (FAD-binding domain and NADP(+)-binding domain) with a molecular mass of 33 kDa. In Cyanidium, FNR was found on thylakoid membranes, but there was no FNR on phycobilisomes. The membrane-bound FNR of Cyanidium was not solublized by NaCl, suggesting the enzyme is tightly bound in the membrane. Although both cyanobacteria and red algae are photoautotrophic organisms bearing phycobilisomes as light harvesting complexes, FNR localization and membrane-binding characteristics were different. These results suggest that FNR binding to phycobilisomes is not characteristic for all phycobilisome retaining oxygenic photosynthetic organisms, and that the rhodoplast of red algae had possibly originated from a cyanobacterium ancestor, whose FNR lacked the CpcD-like domain.

Molecular Microbiology, 1997

Cyanobacteria acclimate to low-temperature conditions by desaturating their membrane lipids. The ... more Cyanobacteria acclimate to low-temperature conditions by desaturating their membrane lipids. The desB (w3 desaturase) and desC (A9 desaturase) genes of Synechococcus sp. strain PCC 7002 were cloned and characterized, and the expression of the desA (A12 desaturase), desB and desC genes was studied as a function of temperature. The steady-state mRNA abundance for the desA gene was threefold higher in cells grown at 22°C than in cells grown at 38°C. desB transcripts were not detected at 38C, but were abundant in cells grown at 22°C. Levels of desC mRNA were similar at both growth temperatures. The mRNA levels of each desaturase gene increased within 15min of a temperature shift-down to 22"C, and mRNA levels recovered within 15min after a shift-up to 38°C. The cold-induced accumulation of transcripts from the desAand desB genes was suppressed by the addition of chloramphenicol, but the transient elevation of the desC transcript levels at 22°C was not affected by chloramphenicol. The half-lives of the desA and desB mRNAs were significantly longer in cells grown at 22°C than in cells grown at 38"C, but the desC mRNA had a similar half-life at both temperatures. These studies reveal three patterns of temperature regulation for the desaturase genes, whose expression is tightly controlled by a combination of mRNA synthesis and stabilization. These studies demonstrate that elevation of desaturase mRNA levels is not the rate-limiting event during the low-temperature acclimation of cyanobacteria.

Journal of Photochemistry and Photobiology B: Biology, 2011

1 Mycosporine-like amino acids (MAAs) are UV absorbing pigments, and structurally distinct 2 MAAs... more 1 Mycosporine-like amino acids (MAAs) are UV absorbing pigments, and structurally distinct 2 MAAs have been identified in taxonomically diverse organisms. Two novel MAAs were 3 purified from the cyanobacterium Nostoc commune, and their chemical structures were 4 characterized. An MAA with an absorption maximum at 335 nm was identified as a 5 pentose-bound porphyra-334 derivative with a molecular mass of 478 Da. Another identified 6 MAA had double absorption maxima at 312 and 340 nm and a molecular mass of 1050 Da. Its 7 unique structure consisted of two distinct chromophores of 3-aminocyclohexen-1-one and 8 1,3-diaminocyclohexen and two pentose and hexose sugars. These MAAs had radical 9 scavenging activity in vitro; the 1050-Da MAA contributed approximately 27% of the total 10 radical scavenging activities in a water extract of N. commune. These results suggest that 11 these glycosylated MAAs have multiple roles as a UV protectant and an antioxidant relevant 12 to anhydrobiosis in N. commune. 13 14 High lights 15 > Two novel glycosylated MAAs were found in Nostoc commune. > The 478-Da MAA was 16 identified as a pentose-bound porphyra-334 derivative. > The 1050-Da MAA consisted of two 17 distinct chromophores, pentose and hexose. > These MAAs had radical scavenging activity. 18 19

FEMS Microbiology Letters, 2006

Plant Physiology, 1997

Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and t... more Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and the main site of the damage is the D1 protein of the photosystem II (PSII) complex. Here we describe that transformation of Synechococcus sp. PCC 7942 with the desA gene for a [delta]12 desaturase increased unsaturation of membrane lipids and enhanced tolerance to strong light. To our knowledge, this is the first report of the successful genetic enhancement of tolerance to strong light. Analysis of the light-induced inactivation and of the subsequent recovery of the activity of the PSII complex revealed that the recovery process was markedly accelerated by the genetic transformation. Labeling experiments with [35S]L-methionine also revealed that the synthesis of the D1 protein de novo at low temperature, which was a prerequisite for the restoration of the PSII complex, was much faster in the transformed cells than in the wild-type cells. These findings demonstrate that the ability of memb...

Plant and Cell Physiology, Mar 1, 1995

Plant and Cell Physiology, Mar 1, 1996

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the ω3 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the ω3 desaturase

Plant and Cell Physiology, 1997

A cyaC gene encoding an adenylate cyclase of the filamentous cyanobacterium Spirulina platensis w... more A cyaC gene encoding an adenylate cyclase of the filamentous cyanobacterium Spirulina platensis was sequenced. The predicted amino acid sequence of the C-terminal region of cyaC is similar to the catalytic domains of adenylate cyclases in other cyanobacteria and eukaryotes. The sequences of other regions are similar to those of proteins consisting of the bacterial two-component signal transduction system: the sensory kinase and the response regulator. The predicted gene product of cyaC contains, from the N-terminal end, a receiver domain of the response regulator protein (Rl), a domain similar to the ETR1 of Arabidopsis thaliana, a transmitter domain of the sensory kinase protein, a receiver domain of the response regulator protein (R2), and a catalytic domain of adenylate cyclase. The cyaC gene was expressed as an affinity-tagged protein in Escherichia coli, and the recombinant protein was purified. The purified protein had adenylate cyclase activity which was activated by Mn 2+. The results of Western blotting using an anti-CyaC antiserum and the S. platensis cell extract confirmed that cyaC gene is expressed in S. platensis.

Current Opinion in Microbiology, 2002

The expression of cold-inducible genes is regulated by a two-component system in Synechocystis an... more The expression of cold-inducible genes is regulated by a two-component system in Synechocystis and Bacillus subtilis. The cold sensors are membrane-bound histidine kinases and it seems likely that they sense and transduce changes in the fluidity of membranes. Desaturation of fatty acids in membrane lipids has been implicated in tolerance to cold and salt stress.

Biochemical Journal, 2006

Acyl-lipid desaturases are enzymes that convert a C–C single bond into a C=C double bond in fatty... more Acyl-lipid desaturases are enzymes that convert a C–C single bond into a C=C double bond in fatty acids that are esterified to membrane-bound glycerolipids. Four types of acyl-lipid desaturase, namely DesA, DesB, DesC, and DesD, acting at the Δ12, Δ15, Δ9, and Δ6 positions of fatty acids respectively, have been characterized in cyanobacteria. These enzymes are specific for fatty acids bound to the sn−1 position of glycerolipids. In the present study, we have cloned two putative genes for a Δ9 desaturase, designated desC1 and desC2, from Nostoc species. The desC1 gene is highly similar to the desC gene that encodes a Δ9 desaturase that acts on C18 fatty acids at the sn−1 position. Homologues of desC2 are found in genomes of cyanobacterial species in which Δ9-desaturated fatty acids are esterified to the sn−2 position. Heterologous expression of the desC2 gene in Synechocystis sp. PCC 6803, in which a saturated fatty acid is found at the sn−2 position, revealed that DesC2 could desatu...

Archives of Microbiology, 1997

Cyanobacteria acclimate to low temperature by desaturating their membrane lipids. Mutant strains ... more Cyanobacteria acclimate to low temperature by desaturating their membrane lipids. Mutant strains of Synechococcus sp. PCC 7002 containing insertionally inactivated desA (Delta12 acyl-lipid desaturase) and desB (omega3 acyl-lipid desaturase) genes were produced, and their low-temperature susceptibility was characterized. The desA mutant synthesized no linoleic acid or alpha-linolenic acid, and the desB mutant did not produce alpha-linolenic acid. The desA mutant grew more slowly than the wild-type at 22 degrees C and could not grow at 15 degrees C. The desB mutant could not continuously grow at 15 degrees C, although no observable phenotype appeared at higher temperatures. It has been shown that expression of the desA gene occurs at 38 degrees C and is up-regulated at 22 degrees C, and that the desB gene is only expressed at 22 degrees C. These results indicate that the expression of the desA and desB genes occurs at higher temperatures than those at which a significant decline in physiological activities is caused by the absence of their products. The temperature dependency of photosynthesis was not affected by these mutations. Since chlorosis and inability to grow at 15 degrees C with nitrate was suppressed by the substitution of urea as a nitrogen source, it is very likely that the chilling susceptibility of the desaturase mutants is attributable to nutrient limitation.

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the 093 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the o93 desaturase from the cyanobacterium, Synechocystis sp. PCC 6803. The desB gene encodes a protein a 359 amino-acid residues with molecular mass of 41.9 kDa. The desB gene is transcribed as a monocistronic operon that produced a single transcript of 1.4 kb. The level of the desB transcript in cells grown at 22 °C was 10 times higher than that in cells grown at 34 ° C. In order to manipulate the fatty-acid unsaturation of membrane lipids, the desB gene in Synechocystis sp. PCC 6803 was mutated by insertion of a kanamycin-resistance gene cartridge. The resultant mutant was unable to desaturate fatty acids at the 093 position. The desA gene, which encodes the A12 desaturase of Synechocystis sp. PCC 6803, and the desB gene were introduced into Synechococcus sp. PCC 7942. Whilst the parent cyanobacterium can only desaturate membrane lipids at the A9 position of fatty acids, the resultant transformant was able to desaturate fatty acids of membrane lipids at the A9, A12 and o93 positions. These results confirm the function of the desB gene and demonstrate that it is possible to genetically manipulate the fatty-acid unsaturation of membrane lipids in cyanobacteria.

Plant physiology

Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and t... more Strong light leads to damage to photosynthetic machinery, particularly at low temperatures, and the main site of the damage is the D1 protein of the photosystem II (PSII) complex. Here we describe that transformation of Synechococcus sp. PCC 7942 with the desA gene for a A12 desaturase increased unsaturation of membrane lipids and enhanced tolerance to strong light. To our knowledge, this is the first report of the successful genetic enhancement of tolerance to strong light.

Applied and environmental microbiology, 1998

Laboratory conditions have been identified that cause the rapid death of cultures of cyanobacteri... more Laboratory conditions have been identified that cause the rapid death of cultures of cyanobacteria producing urease. Once the death phase had initiated in the stationary growth phase, cells were rapidly bleached of all pigmentation. Null mutations in the ureC gene, encoding the alpha subunit of urease, were constructed, and these mutants were no longer sensitive to growth in the presence of urea. High levels of peroxides, including lipid peroxides, were detected in the bleaching cells. Exogenously added polyunsaturated fatty acids triggered a similar death response. Vitamin E suppressed the formation of peroxides and delayed the onset of cell bleaching. The results suggest that these cyanobacterial cells undergo a metabolic imbalance that ultimately leads to oxidative stress and lipid peroxide formation. These observations may provide insights into the mechanism of sudden cyanobacterial bloom disappearance in nature.

Photosynthesis research, 2002

The effect of polyunsaturated fatty acids on photosynthesis and the growth of the marine cyanobac... more The effect of polyunsaturated fatty acids on photosynthesis and the growth of the marine cyanobacterium Synechococcus sp. PCC 7002 was examined using wild-type and Delta12 fatty acid desaturase mutant strains. Under a light intensity of 250 mumol m(-2) s(-1), wild-type cells could grow exponentially in a temperature range of 20-38 degrees C, but growth was non-exponential below 20 degrees C and ceased at 12 degrees C. The Delta12 desaturase mutant cells lacking polyunsaturated fatty acids had the same growth rate as wild-type cells in a temperature range of 25-38 degrees C but grew slowly at 22 degrees C, and no cell growth took place below 18 degrees C. Under a very high-light intensity of 2.5 mmol m(-2) s(-1), wild-type cells could grow exponentially in a temperature range of 30-38 degrees C, although the high-light grown cells became chlorotic because of nitrogen limitation. The temperature sensitive phenotype in the Delta12 desaturase mutant was enhanced in cells grown under hig...

Plant Molecular Biology, 1994

Cyanobacterial genes for enzymes that desaturate fatty acids at the A12 position, designated desA... more Cyanobacterial genes for enzymes that desaturate fatty acids at the A12 position, designated desA, were isolated from Synechocystis PCC6714, Synechococcus PCC7002 and Anabaena variabilis by crosshybridization with a DNA probe derived from the desA gene of Synechocystis PCC6803. The genes of Synechocystis PCC6714, Synechococcus PCC7002 and A. variabilis encode proteins of 349, 347 and 350 amino acid residues, respectively. The transformation of Synechococcus PCC7942 with the desA genes from Synechocystis PCC6714, Synechococcus PCC7002 and A. variabilis was associated with the ability to introduce a second double bond at the A12 position of fatty acids. The amino acid sequence of the products of the desA genes revealed the presence of four conserved domains. Since one of the conserved domains was also found in the amino acid sequences of o93 desaturases ofBrassica napus and mung bean, this domain may play an essential role in the introduction of a double bond into fatty acids bound to membrane lipids.

Plant Molecular Biology, 1996

A cyaA gene, encoding an adenylate cyclase, was isolated from a filamentous cyanobacterium, Spiru... more A cyaA gene, encoding an adenylate cyclase, was isolated from a filamentous cyanobacterium, Spirulina platensis, by functional complementation of a cya mutant of Escherichia coli, defective in adenylate cyclase activity. The predicted gene product of cyaA contains a signal peptide-like domain, a putative sensor domain similar to the gene product of vsrA of Pseudomonas solanacearum, a putative membrane-spanning domain and an adenylate cyclase-like catalytic domain. Two other positive clones that complemented the E. coli mutant were isolated from the same cyanobacterium, suggesting that several cya genes are functioning in S. platensis.

Plant Molecular Biology, 1994

Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids... more Cyanobacteria respond to a decrease in temperature by desaturating fatty acids of membrane lipids to compensate for the decrease in membrane fluidity. Among various desaturation reactions in cyanobacteria, the desaturation of the ω3 position of fatty acids is the most sensitive to the change in temperature. In the present study, we isolated a gene, designated desB, for the ω3 desaturase from the cyanobacterium, Synechocystis sp. PCC 6803. The desB gene encodes a protein a 359 amino-acid residues with molecular mass of 41.9 kDa. The desB gene is transcribed as a monocistronic operon that produced a single transcript of 1.4 kb. The level of the desB transcript in cells grown at 22°C was 10 times higher than that in cells grown at 34°C. In order to manipulate the fatty-acid unsaturation of membrane lipids, the desB gene in Synechocystis sp. PCC 6803 was mutated by insertion of a kanamycin-resistance gene cartridge. The resultant mutant was unable to desaturate fatty acids at the ω3 position. The desA gene, which encodes the Δ12 desaturase of Synechocystis sp. PCC 6803, and the desB gene were introduced into Synechococcus sp. PCC 7942. Whilst the parent cyanobacterium can only desaturate membrane lipids at the Δ9 position of fatty acids, the resultant transformant was able to desaturate fatty acids of membrane lipids at the Δ9, Δ12 and ω3 positions. These results confirm the function of the desB gene and demonstrate that it is possible to genetically manipulate the fatty-acid unsaturation of membrane lipids in cyanobacteria.

Phycological Research, 2009

Changes in photosynthetic activity and trehalose levels in field-isolated, natural colonies of th... more Changes in photosynthetic activity and trehalose levels in field-isolated, natural colonies of the terrestrial cyanobacterium Nostoc commune responding to desiccation and salt stress were investigated. As the water content decreased in N. commune colonies during desiccation, photosynthetic O2-evolving activity decreased and no activity was detected in desiccated colonies. A high level of O2 evolution was restored in the colonies as they absorbed atmospheric moisture, indicating that only a small amount of water is required for reactivation of photosynthesis. No detectable trehalose was found in fully hydrated N. commune colonies; however, trehalose accumulation occurred in response to water loss during desiccation and high levels of trehalose were detected in the airdried colonies. Moreover, a 0.2 M NaCl treatment also induced trehalose accumulation to a level equivalent to that by desiccation. Photosynthetic O2 evolution was inhibited by 0.2 M NaCl, indicating that N. commune can tolerate only low levels of salt. These results suggest that cessation of photosynthesis and trehalose accumulation occur in response to both matric water stress (desiccation) and osmotic water stress (high salt concentration), and that while trehalose may be a less effective osmoprotective compound than others, it is important for the extreme tolerance to desiccation observed in terrestrial cyanobacterium.

Photosynthesis Research, 2007

Ferredoxin-NADP(+) oxidoreductase (FNR) catalyzing the terminal step of the linear photosynthetic... more Ferredoxin-NADP(+) oxidoreductase (FNR) catalyzing the terminal step of the linear photosynthetic electron transport was purified from the cyanobacterium Spirulina platensis and the red alga Cyanidium caldarium. FNR of Spirulina consisted of three domains (CpcD-like domain, FAD-binding domain, and NADP(+)-binding domain) with a molecular mass of 46 kDa and was localized in either phycobilisomes or thylakoid membranes. The membrane-bound FNR with 46 kDa was solublized by NaCl and the solublized FNR had an apparent molecular mass of 90 kDa. FNR of Cyanidium consisted of two domains (FAD-binding domain and NADP(+)-binding domain) with a molecular mass of 33 kDa. In Cyanidium, FNR was found on thylakoid membranes, but there was no FNR on phycobilisomes. The membrane-bound FNR of Cyanidium was not solublized by NaCl, suggesting the enzyme is tightly bound in the membrane. Although both cyanobacteria and red algae are photoautotrophic organisms bearing phycobilisomes as light harvesting complexes, FNR localization and membrane-binding characteristics were different. These results suggest that FNR binding to phycobilisomes is not characteristic for all phycobilisome retaining oxygenic photosynthetic organisms, and that the rhodoplast of red algae had possibly originated from a cyanobacterium ancestor, whose FNR lacked the CpcD-like domain.

Molecular Microbiology, 1997

Cyanobacteria acclimate to low-temperature conditions by desaturating their membrane lipids. The ... more Cyanobacteria acclimate to low-temperature conditions by desaturating their membrane lipids. The desB (w3 desaturase) and desC (A9 desaturase) genes of Synechococcus sp. strain PCC 7002 were cloned and characterized, and the expression of the desA (A12 desaturase), desB and desC genes was studied as a function of temperature. The steady-state mRNA abundance for the desA gene was threefold higher in cells grown at 22°C than in cells grown at 38°C. desB transcripts were not detected at 38C, but were abundant in cells grown at 22°C. Levels of desC mRNA were similar at both growth temperatures. The mRNA levels of each desaturase gene increased within 15min of a temperature shift-down to 22"C, and mRNA levels recovered within 15min after a shift-up to 38°C. The cold-induced accumulation of transcripts from the desAand desB genes was suppressed by the addition of chloramphenicol, but the transient elevation of the desC transcript levels at 22°C was not affected by chloramphenicol. The half-lives of the desA and desB mRNAs were significantly longer in cells grown at 22°C than in cells grown at 38"C, but the desC mRNA had a similar half-life at both temperatures. These studies reveal three patterns of temperature regulation for the desaturase genes, whose expression is tightly controlled by a combination of mRNA synthesis and stabilization. These studies demonstrate that elevation of desaturase mRNA levels is not the rate-limiting event during the low-temperature acclimation of cyanobacteria.

Journal of Photochemistry and Photobiology B: Biology, 2011

1 Mycosporine-like amino acids (MAAs) are UV absorbing pigments, and structurally distinct 2 MAAs... more 1 Mycosporine-like amino acids (MAAs) are UV absorbing pigments, and structurally distinct 2 MAAs have been identified in taxonomically diverse organisms. Two novel MAAs were 3 purified from the cyanobacterium Nostoc commune, and their chemical structures were 4 characterized. An MAA with an absorption maximum at 335 nm was identified as a 5 pentose-bound porphyra-334 derivative with a molecular mass of 478 Da. Another identified 6 MAA had double absorption maxima at 312 and 340 nm and a molecular mass of 1050 Da. Its 7 unique structure consisted of two distinct chromophores of 3-aminocyclohexen-1-one and 8 1,3-diaminocyclohexen and two pentose and hexose sugars. These MAAs had radical 9 scavenging activity in vitro; the 1050-Da MAA contributed approximately 27% of the total 10 radical scavenging activities in a water extract of N. commune. These results suggest that 11 these glycosylated MAAs have multiple roles as a UV protectant and an antioxidant relevant 12 to anhydrobiosis in N. commune. 13 14 High lights 15 > Two novel glycosylated MAAs were found in Nostoc commune. > The 478-Da MAA was 16 identified as a pentose-bound porphyra-334 derivative. > The 1050-Da MAA consisted of two 17 distinct chromophores, pentose and hexose. > These MAAs had radical scavenging activity. 18 19

FEMS Microbiology Letters, 2006