Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining d... more Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining due to global warming-mediated changes in the oceans. Particularly for the Caribbean region, Acropora sp. stony corals have lost ∼80% of their original coverage, resulting in vast extensions of dead coral rubble. We analysed the microbial composition of biofilms that colonize and lithify dead A. palmata rubble in the Mexican Caribbean and identified the microbial assemblages that can persist under scenarios of global change, including high temperature and low pH. Lithifying biofilms have a mineral composition that includes aragonite and magnesium calcite (16 mole% MgCO3) and calcite, while the mineral phase corresponding to coral skeleton is basically aragonite. Microbial composition of the lithifying biofilms are different in comparison to surrounding biotopes, including a microbial mat, water column, sediments and live A. palmata microbiome. Significant shifts in biofilm composition wer...
Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining d... more Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining due to global warming mediated changes in the oceans. Particularly for the Caribbean region, Acropora sp. stony corals have lost ∼80% of their original coverage, resulting in vast extensions of dead coral rubble. We analyzed the microbial composition of biofilms that colonize and lithify dead A. palmata rubble in the Mexican Caribbean and identified the microbial assemblages that can persist under scenarios of global change, including high temperature and low pH. Lithifying biofilms have a mineral composition that includes aragonite and magnesium calcite (16 mole% MgCO3) and calcite, while the mineral phase corresponding to coral skeleton is basically aragonite. Microbial composition of the lithifying biofilms are different in comparison to surrounding biotopes, including a microbial mat, water column, sediments and live A. palmata microbiome. Significant shifts in biofilm composition wer...
Signaling pathways are fundamental for the establishment and maintenance of diverse symbioses. Th... more Signaling pathways are fundamental for the establishment and maintenance of diverse symbioses. The symbiosis of cnidarians and dinoflagellate algae is the foundation for the ecological success of coral reefs, involving the transfer of photosynthetic products from symbiont to host. However, signal transduction pathways for this symbiosis remain uncharacterized. Cultured and natural cnidarian symbionts can produce glycerol, one of the main translocated photosynthates. Here, we investigate whether a signal transduction pathway may be involved in inducing glycerol synthesis in cultured symbionts under an osmotic stress model. We evaluated the effect of specific inhibitors of the main transduction pathways, p38, JNK, and ERK 1/2 in Brevolium minutum, the symbiont of the Aiptasia model system. We found that glycerol production and the specific activity of the enzyme Gpdh were selectively inhibited by a p38 MAPK inhibitor. Additionally, the phosphorylation of a putative p38-like protein was rapidly detected. Finally, we studied the presence of each of the components of the p38 MAPK pathway in silico, in genomes and transcriptomes reported up to date for different symbiont types. We propose a model for the arrangement of this pathway in the family of dinoflagellate symbionts known as Symbiodiniaceae.
Background: Symbiotic dinoflagellates transfer a substantial amount of their photosynthetic produ... more Background: Symbiotic dinoflagellates transfer a substantial amount of their photosynthetic products to their animal hosts. This amount has been estimated to represent up to 90 % of the photosynthetically fixed carbon and can satisfy in some instances the full respiratory requirements of the host. Although in several cnidarian-dinoflagellate symbioses glycerol is the primary photosynthetic product translocated to the host, the mechanism for its production and release has not been demonstrated conclusively. Principal Findings: Using Symbiodinium cells in culture we were able to reproduce the synthesis and release of glycerol in vitro by employing an inductor for glycerol synthesis, osmotic up-shocks. Photosynthetic parameters and fluorescence analysis of photosystem II showed that the inductive conditions did not have a negative effect on photosynthetic performance, suggesting that the capacity for carbon fixation by the cells was not compromised. The demand for glycerol production r...
Coral bleaching—the loss of symbiotic dinoflagellates—is initiated when corals are exposed to sea... more Coral bleaching—the loss of symbiotic dinoflagellates—is initiated when corals are exposed to sea surface temperatures above the regional summer average and has been responsible for massive coral mortality episodes. The ability of symbionts to recolonize the host after bleaching may be critical in determining if a colony will recover or experience mortality. Here, following the serendipitous bleaching of specimens of Montastraea faveolata of known photosynthetic, spectroscopic, and genetic characteristics, we describe changes in photosynthesis and light utilization during the recovery process. Fully recovered M. faveolata had minimum quantum requirements (1/Wmax) that are very close to the theoretical minimum, indicating that symbiotic corals are not only one of the most efficient light collectors in nature, but also use this energy with maximum efficiency. Analyses of the photosynthetic responses of M. faveolata throughout the recovery process indicate that during the early stages,...
... from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense T... more ... from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense Trench from the acoel flatworm Haplodiscus sp. (from Belau) have been amplified by the polymerase chain reaction, cloned, and sequenced. ...
: In the yeasts Saccharomyces cerevisiae and Debaryomyces hansenii, glycerol-3-phosphate dehydrog... more : In the yeasts Saccharomyces cerevisiae and Debaryomyces hansenii, glycerol-3-phosphate dehydrogenase (NAD+-GPD) is believed to be the enzyme controlling the synthesis of glycerol in osmotically challenged cells. Consistent with this concept, immunoblot analyses demonstrated quantitative increases in NAD+-GPD 30 minutes after the addition of NaCl to actively growing D. hansenii cells; Northern blot analyses showed increases in GPD-messenger RNA within 15 minutes after the addition of salt. There appears to be an upper limit of salt concentration above which the cells appear to be less responsive. The results we describe support the concept of transcriptional regulation of NAD+-GPD in D. hansenii.
Cassiopea xamachana jellyfish are an attractive model system to study metamorphosis and/or cnidar... more Cassiopea xamachana jellyfish are an attractive model system to study metamorphosis and/or cnidarian-dinoflagellate symbiosis due to the ease of cultivation of their planula larvae and scyphistomae through their asexual cycle, in which the latter can bud new larvae and continue the cycle without differentiation into ephyrae. Then, a subsequent induction of metamorphosis and full differentiation into ephyrae is believed to occur when the symbionts are acquired by the scyphistomae. Although strobilation induction and differentiation into ephyrae can be accomplished in various ways, a controlled, reproducible metamorphosis induction has not been reported. Such controlled metamorphosis induction is necessary for an ensured synchronicity and reproducibility of biological, biochemical, and molecular analyses. For this purpose, we tested if differentiation could be pharmacologically stimulated as in Aurelia aurita, by the metamorphic inducers thyroxine, KI, NaI, Lugol's iodine, H2O2, i...
A gene homologous to GPD1, coding for glycerol-3-phosphate dehydrogenase (sn-glycerol 3-phosphate... more A gene homologous to GPD1, coding for glycerol-3-phosphate dehydrogenase (sn-glycerol 3-phosphate: NAD(+) oxidoreductase, EC 1.1.1.8), has been isolated from the halophilic yeast Debaryomyces hansenii by complementation of a Saccharomyces cerevisiae gpd1 Delta mutant. DNA sequencing of the complementing genomic clone indicated the existence of an open reading frame encoding a protein with 369 amino acids. Comparative analysis of the deduced amino acid sequence showed high similarity to homologous genes described for other eukaryotic GPD enzymes. The sequence has been submitted to the GenBank database under Accession No. AY333427.
World Journal of Microbiology and Biotechnology, 2015
Eukaryotic microorganisms possess mechanisms to detect osmotic variations in their surroundings, ... more Eukaryotic microorganisms possess mechanisms to detect osmotic variations in their surroundings, from specialized receptors and membrane transporters, to sophisticated systems such as two-component histidine kinases. Osmotic stimuli are transduced through conserved phosphorylation cascades that result in a rapid response to mitigate stress. This response allows for the maintenance of an optimal biochemical environment for cell functioning, as well as a suitable recovery in suboptimal environments that would otherwise endanger cell survival. The molecular basis of these responses has been largely studied in yeasts and bacteria. However, fewer studies have been published concerning the molecular basis of osmoregulation in other eukaryotic microorganisms such as protozoans and microalgae. Here, we review the main osmosensors reported in unicellular eukaryotic microorganisms (yeasts, microalgae and protozoa) and the pathways that maintain homeostasis in cells encountering hyperosmotic challenges.
Debaryomyces hansenii showed an increased growth in the presence of either 1 M, KCl or 1 M NaCl a... more Debaryomyces hansenii showed an increased growth in the presence of either 1 M, KCl or 1 M NaCl and a low acidification of the medium, higher for the cells grown in the presence of NaCl. These cells accumulated high concentrations of the cations, and showed a very fast capacity to exchange either Na+ or K+ for the opposite cation. They showed a rapid uptake of 86Rb+ and 22Na+. 86Rb+ transport was saturable, with K(m) and Vmax values higher for cells grown in 1 M NaCl. 22Na+ uptake showed a diffusion component, also higher for the cells grown with NaCl. Changes depended on growth conditions, and not on further incubation, which changed the internal ion concentration. K+ stimulated proton pumping produced a rapid extrusion of protons, and also a decrease of the membrane potential. Cells grown in 1 M KCl showed a higher fermentation rate, but significantly lower respiratory capacity. ATP levels were higher in cells grown in the presence of NaCl; upon incubation with glucose, those grown in the presence of KCl reached values similar to the ones grown in the presence of NaCl. In both, the addition of KCl produced a transient decrease of the ATP levels. As to ion transport mechanisms, D. hansenii appears to have (a) an ATPase functioning as a proton pump, generating a membrane potential difference which drives K+ through a uniporter; (b) a K+/H+ exchange system; and (c) a rapid cation/cation exchange system. Most interesting is that cells grown in different ionic environments change their studied capacities, which are not dependent on the cation content, but on differences in their genetic expression during growth.
The role for the gene encoding glycerol 3-phosphate dehydrogenase (DhGPD1) from the osmotolerant ... more The role for the gene encoding glycerol 3-phosphate dehydrogenase (DhGPD1) from the osmotolerant yeast Debaryomyces hansenii, in glycerol production and halotolerance, was studied through its heterologous expression in a Saccharomyces cerevisiae strain deficient in glycerol synthesis (gpd1Delta). The expression of the DhGPD1 gene in the gpd1Delta background restored glycerol production and halotolerance to wild type levels, corroborating its role in the salt-induced production of glycerol. Although the gene was functional in S. cerevisiae, its heterologous expression was not efficient, suggesting that the regulatory mechanism may not be shared by these two yeasts.
The highly halotolerant yeast Debaryomyces hansenii when grown in the presence of 2M NaCl, increa... more The highly halotolerant yeast Debaryomyces hansenii when grown in the presence of 2M NaCl, increased the expression of ARO4 which is involved in the biosynthesis of aromatic amino acids. The function of the isolated gene was verified by complementation of a Saccharomyces cerevisiae null mutant, aro4Delta, restoring the specific activity of the enzyme (a 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase) to wild-type levels. DhARO4 transcript expression under high salinity was stimulated at the beginning of the exponential growth phase. As the DhARO4 promoter region presents putative GCRE and CRE sequences, its expression was evaluated under conditions of NaCl stress and amino acid starvation, showing similar expression levels for either condition. The combined effect of both stressors resulted in a further increase in transcript levels over the singly added stressors, indicating independent stimulatory events. Our results support the hypothesis that high salinity and amino acid availability are physiologically interconnected.
Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining d... more Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining due to global warming-mediated changes in the oceans. Particularly for the Caribbean region, Acropora sp. stony corals have lost ∼80% of their original coverage, resulting in vast extensions of dead coral rubble. We analysed the microbial composition of biofilms that colonize and lithify dead A. palmata rubble in the Mexican Caribbean and identified the microbial assemblages that can persist under scenarios of global change, including high temperature and low pH. Lithifying biofilms have a mineral composition that includes aragonite and magnesium calcite (16 mole% MgCO3) and calcite, while the mineral phase corresponding to coral skeleton is basically aragonite. Microbial composition of the lithifying biofilms are different in comparison to surrounding biotopes, including a microbial mat, water column, sediments and live A. palmata microbiome. Significant shifts in biofilm composition wer...
Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining d... more Coral reefs are amongst the most productive ecosystems on the planet, but are rapidly declining due to global warming mediated changes in the oceans. Particularly for the Caribbean region, Acropora sp. stony corals have lost ∼80% of their original coverage, resulting in vast extensions of dead coral rubble. We analyzed the microbial composition of biofilms that colonize and lithify dead A. palmata rubble in the Mexican Caribbean and identified the microbial assemblages that can persist under scenarios of global change, including high temperature and low pH. Lithifying biofilms have a mineral composition that includes aragonite and magnesium calcite (16 mole% MgCO3) and calcite, while the mineral phase corresponding to coral skeleton is basically aragonite. Microbial composition of the lithifying biofilms are different in comparison to surrounding biotopes, including a microbial mat, water column, sediments and live A. palmata microbiome. Significant shifts in biofilm composition wer...
Signaling pathways are fundamental for the establishment and maintenance of diverse symbioses. Th... more Signaling pathways are fundamental for the establishment and maintenance of diverse symbioses. The symbiosis of cnidarians and dinoflagellate algae is the foundation for the ecological success of coral reefs, involving the transfer of photosynthetic products from symbiont to host. However, signal transduction pathways for this symbiosis remain uncharacterized. Cultured and natural cnidarian symbionts can produce glycerol, one of the main translocated photosynthates. Here, we investigate whether a signal transduction pathway may be involved in inducing glycerol synthesis in cultured symbionts under an osmotic stress model. We evaluated the effect of specific inhibitors of the main transduction pathways, p38, JNK, and ERK 1/2 in Brevolium minutum, the symbiont of the Aiptasia model system. We found that glycerol production and the specific activity of the enzyme Gpdh were selectively inhibited by a p38 MAPK inhibitor. Additionally, the phosphorylation of a putative p38-like protein was rapidly detected. Finally, we studied the presence of each of the components of the p38 MAPK pathway in silico, in genomes and transcriptomes reported up to date for different symbiont types. We propose a model for the arrangement of this pathway in the family of dinoflagellate symbionts known as Symbiodiniaceae.
Background: Symbiotic dinoflagellates transfer a substantial amount of their photosynthetic produ... more Background: Symbiotic dinoflagellates transfer a substantial amount of their photosynthetic products to their animal hosts. This amount has been estimated to represent up to 90 % of the photosynthetically fixed carbon and can satisfy in some instances the full respiratory requirements of the host. Although in several cnidarian-dinoflagellate symbioses glycerol is the primary photosynthetic product translocated to the host, the mechanism for its production and release has not been demonstrated conclusively. Principal Findings: Using Symbiodinium cells in culture we were able to reproduce the synthesis and release of glycerol in vitro by employing an inductor for glycerol synthesis, osmotic up-shocks. Photosynthetic parameters and fluorescence analysis of photosystem II showed that the inductive conditions did not have a negative effect on photosynthetic performance, suggesting that the capacity for carbon fixation by the cells was not compromised. The demand for glycerol production r...
Coral bleaching—the loss of symbiotic dinoflagellates—is initiated when corals are exposed to sea... more Coral bleaching—the loss of symbiotic dinoflagellates—is initiated when corals are exposed to sea surface temperatures above the regional summer average and has been responsible for massive coral mortality episodes. The ability of symbionts to recolonize the host after bleaching may be critical in determining if a colony will recover or experience mortality. Here, following the serendipitous bleaching of specimens of Montastraea faveolata of known photosynthetic, spectroscopic, and genetic characteristics, we describe changes in photosynthesis and light utilization during the recovery process. Fully recovered M. faveolata had minimum quantum requirements (1/Wmax) that are very close to the theoretical minimum, indicating that symbiotic corals are not only one of the most efficient light collectors in nature, but also use this energy with maximum efficiency. Analyses of the photosynthetic responses of M. faveolata throughout the recovery process indicate that during the early stages,...
... from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense T... more ... from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense Trench from the acoel flatworm Haplodiscus sp. (from Belau) have been amplified by the polymerase chain reaction, cloned, and sequenced. ...
: In the yeasts Saccharomyces cerevisiae and Debaryomyces hansenii, glycerol-3-phosphate dehydrog... more : In the yeasts Saccharomyces cerevisiae and Debaryomyces hansenii, glycerol-3-phosphate dehydrogenase (NAD+-GPD) is believed to be the enzyme controlling the synthesis of glycerol in osmotically challenged cells. Consistent with this concept, immunoblot analyses demonstrated quantitative increases in NAD+-GPD 30 minutes after the addition of NaCl to actively growing D. hansenii cells; Northern blot analyses showed increases in GPD-messenger RNA within 15 minutes after the addition of salt. There appears to be an upper limit of salt concentration above which the cells appear to be less responsive. The results we describe support the concept of transcriptional regulation of NAD+-GPD in D. hansenii.
Cassiopea xamachana jellyfish are an attractive model system to study metamorphosis and/or cnidar... more Cassiopea xamachana jellyfish are an attractive model system to study metamorphosis and/or cnidarian-dinoflagellate symbiosis due to the ease of cultivation of their planula larvae and scyphistomae through their asexual cycle, in which the latter can bud new larvae and continue the cycle without differentiation into ephyrae. Then, a subsequent induction of metamorphosis and full differentiation into ephyrae is believed to occur when the symbionts are acquired by the scyphistomae. Although strobilation induction and differentiation into ephyrae can be accomplished in various ways, a controlled, reproducible metamorphosis induction has not been reported. Such controlled metamorphosis induction is necessary for an ensured synchronicity and reproducibility of biological, biochemical, and molecular analyses. For this purpose, we tested if differentiation could be pharmacologically stimulated as in Aurelia aurita, by the metamorphic inducers thyroxine, KI, NaI, Lugol's iodine, H2O2, i...
A gene homologous to GPD1, coding for glycerol-3-phosphate dehydrogenase (sn-glycerol 3-phosphate... more A gene homologous to GPD1, coding for glycerol-3-phosphate dehydrogenase (sn-glycerol 3-phosphate: NAD(+) oxidoreductase, EC 1.1.1.8), has been isolated from the halophilic yeast Debaryomyces hansenii by complementation of a Saccharomyces cerevisiae gpd1 Delta mutant. DNA sequencing of the complementing genomic clone indicated the existence of an open reading frame encoding a protein with 369 amino acids. Comparative analysis of the deduced amino acid sequence showed high similarity to homologous genes described for other eukaryotic GPD enzymes. The sequence has been submitted to the GenBank database under Accession No. AY333427.
World Journal of Microbiology and Biotechnology, 2015
Eukaryotic microorganisms possess mechanisms to detect osmotic variations in their surroundings, ... more Eukaryotic microorganisms possess mechanisms to detect osmotic variations in their surroundings, from specialized receptors and membrane transporters, to sophisticated systems such as two-component histidine kinases. Osmotic stimuli are transduced through conserved phosphorylation cascades that result in a rapid response to mitigate stress. This response allows for the maintenance of an optimal biochemical environment for cell functioning, as well as a suitable recovery in suboptimal environments that would otherwise endanger cell survival. The molecular basis of these responses has been largely studied in yeasts and bacteria. However, fewer studies have been published concerning the molecular basis of osmoregulation in other eukaryotic microorganisms such as protozoans and microalgae. Here, we review the main osmosensors reported in unicellular eukaryotic microorganisms (yeasts, microalgae and protozoa) and the pathways that maintain homeostasis in cells encountering hyperosmotic challenges.
Debaryomyces hansenii showed an increased growth in the presence of either 1 M, KCl or 1 M NaCl a... more Debaryomyces hansenii showed an increased growth in the presence of either 1 M, KCl or 1 M NaCl and a low acidification of the medium, higher for the cells grown in the presence of NaCl. These cells accumulated high concentrations of the cations, and showed a very fast capacity to exchange either Na+ or K+ for the opposite cation. They showed a rapid uptake of 86Rb+ and 22Na+. 86Rb+ transport was saturable, with K(m) and Vmax values higher for cells grown in 1 M NaCl. 22Na+ uptake showed a diffusion component, also higher for the cells grown with NaCl. Changes depended on growth conditions, and not on further incubation, which changed the internal ion concentration. K+ stimulated proton pumping produced a rapid extrusion of protons, and also a decrease of the membrane potential. Cells grown in 1 M KCl showed a higher fermentation rate, but significantly lower respiratory capacity. ATP levels were higher in cells grown in the presence of NaCl; upon incubation with glucose, those grown in the presence of KCl reached values similar to the ones grown in the presence of NaCl. In both, the addition of KCl produced a transient decrease of the ATP levels. As to ion transport mechanisms, D. hansenii appears to have (a) an ATPase functioning as a proton pump, generating a membrane potential difference which drives K+ through a uniporter; (b) a K+/H+ exchange system; and (c) a rapid cation/cation exchange system. Most interesting is that cells grown in different ionic environments change their studied capacities, which are not dependent on the cation content, but on differences in their genetic expression during growth.
The role for the gene encoding glycerol 3-phosphate dehydrogenase (DhGPD1) from the osmotolerant ... more The role for the gene encoding glycerol 3-phosphate dehydrogenase (DhGPD1) from the osmotolerant yeast Debaryomyces hansenii, in glycerol production and halotolerance, was studied through its heterologous expression in a Saccharomyces cerevisiae strain deficient in glycerol synthesis (gpd1Delta). The expression of the DhGPD1 gene in the gpd1Delta background restored glycerol production and halotolerance to wild type levels, corroborating its role in the salt-induced production of glycerol. Although the gene was functional in S. cerevisiae, its heterologous expression was not efficient, suggesting that the regulatory mechanism may not be shared by these two yeasts.
The highly halotolerant yeast Debaryomyces hansenii when grown in the presence of 2M NaCl, increa... more The highly halotolerant yeast Debaryomyces hansenii when grown in the presence of 2M NaCl, increased the expression of ARO4 which is involved in the biosynthesis of aromatic amino acids. The function of the isolated gene was verified by complementation of a Saccharomyces cerevisiae null mutant, aro4Delta, restoring the specific activity of the enzyme (a 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase) to wild-type levels. DhARO4 transcript expression under high salinity was stimulated at the beginning of the exponential growth phase. As the DhARO4 promoter region presents putative GCRE and CRE sequences, its expression was evaluated under conditions of NaCl stress and amino acid starvation, showing similar expression levels for either condition. The combined effect of both stressors resulted in a further increase in transcript levels over the singly added stressors, indicating independent stimulatory events. Our results support the hypothesis that high salinity and amino acid availability are physiologically interconnected.
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