Alessandra Norici

An Anaplerotic Role for Mitochondrial Carbonic Anhydrase in Chlamydomonas reinhardtii

Previous studies of the mitochondrial carbonic anhydrase (mtCA) of Chlamydomonas reinhardtii showed that expression of the two genes encoding this enzyme activity required photosynthetically active radiation and a low CO2 concentration. These studies suggested that the mtCA was involved in the inorganic carbon-concentrating mechanism. We have now shown that the expression of the mtCA at low CO2 concentrations decreases when the external NH4+ concentration decreases, to the point of being undetectable when NH4+ supply restricts the rate of photoautotrophic growth. The expression of mtCA can also be induced at supra-atmospheric partial pressure of CO2 by increasing the NH4+ concentration in the growth medium. Conditions that favor mtCA expression usually also stimulate anaplerosis. We therefore propose that the mtCA is involved in supplying HCO3- for anaplerotic assimilation catalyzed by phosphoenolpyruvate carboxylase, which provides C skeletons for N assimilation under some circumstances.

FTIR spectra of algal species can be used as physiological fingerprints to assess their actual growth potential

Fourier transform infrared (FTIR) spectra were measured from cells of Microcystis aeruginosa and Protoceratium reticulatum, whose growth rates were manipulated by the availability of nutrients or light. As expected, the macromolecular composition changed in response to the treatments. These changes were species-specific and depended on the type of perturbation applied to the growth regime. Microcystis aeruginosa showed an increase in the carbohydrate-to-protein ratio with decreased growth rates, under nutrient limitation, whereas light limitation induced a decrease of the carbohydrate-to-protein ratio with decreasing proliferation rates. The macromolecular pools of P. reticulatum showed a higher degree of compositional homeostasis. Only when the lowest light irradiance and nutrient availability were supplied, an increase of the carbohydrate-to-protein FTIR absorbance ratio was observed. A species-specific partial least squares (PLS) model was developed using the whole FTIR spectra. This model afforded a very high correlation between the predicted and the measured growth rates, regardless of the growth conditions. On the contrary, the prediction based on absorption band ratios generally used in FTIR studies would strongly depend on growth conditions. This new computational method could constitute a substantial improvement in the early warning systems of algal blooms and, in general, for the study of algal growth, e.g. in biotechnology. Furthermore, these results confirm the suitability of FTIR spectroscopy as a tool to map complex biological processes like growth under different environmental conditions.

Impact of irradiance on the C allocation in the coastal marine diatom Skeletonema marinoi Sarno and Zingone

Elemental stoichiometry and organic composition were investigated in an Adriatic strain of Skeletonema marinoi, cultured at 25 [low light (LL)] and 250 [high light (HL)]µmol photon m⁻² s⁻¹. Inorganic carbon acquisition, fixation and allocation, and silicic acid and orthophosphate uptake were also studied. The C:P ratio was below the Redfield ratio, especially at LL. In HL cells, N quota was halved, C quota was similar, silica quota was lower, growth rate and long-term net primary productivity were almost doubled, relative to LL cells. The HL:LL cell quota ratios were 6 for lipid, 0.5 for protein and 0.4 for carbohydrate. Phosphoenolpyruvate carboxylase (PEPc) and glutamine synthetase (GS) activities were unaffected by the growth irradiance; phosphoenolpyruvate carboxykinase (PEPck) was 2.5-fold more active in LL cells. This suggests that in S. marinoi, C₄ photosynthesis is unlikely, PEPc is anaplerotic and PEPck may be involved in the conversion of lipid C to carbohydrates, especially in LL cells. Because about 50% of the cost for the production of an HL cell is caused by lipid biosynthesis, we propose that the preferential allocation of C to lipid at HL takes advantage of the relatively high volume-based energy content of lipids, in an organism that reduces its size at each vegetative cell division.

Sulfur and primary production in aquatic environments: an ecological perspective.

Sulfur is one of the critical elements in living matter, as it participates in several structural, metabolic and catalytic activities. Photosynthesis is an important process that entails the use of sulfur during both the light and carbon reactions. Nearly half of global photosynthetic carbon fixation is carried out by phytoplankton in the aquatic environment. Aquatic environments are very different from one another with respect to sulfur content: while in the oceans sulfate concentration is constantly high, freshwaters are characterized by daily and seasonal variations and by a wide range of sulfur concentration. The strategies that algal cells adopt for energy and resource allocation often reflect these differences. In the oceans, the amount and chemical form of sulfur has changed substantially during the course of the Earths history; it is possible that sulfur availability played a role in the evolution of marine phytoplankton communities and it may continue to have appreciable effects on global biogeochemistry and ecology. Phytoplankton is also the main biogenic source of sulfur; sulfur can be released into the atmosphere by algal cells as dimethylsulfide, with possibly important repercussions on global climate. These and related matters are discussed in this review.

Physiological responses of the green alga Dunaliella parva (Volvocales, Chlorophyta) to controlled incremental changes in the N source

This work is aimed at obtaining information on the acclimation processes of the green flagellate Dunaliella parva Lerche to gradual changes in the N source from NO3- to NH4+, in continuous cultures. Photosynthesis, dark respiration, and light-independent carbon fixation (LICF) rates, chlorophyll a fluorescence, RUBISCO and phosphoenolpyruvate carboxylase (PEPc) activities, plasmalemma electrical potential difference, cell volume, and absolute or relative amounts of major cell constituents were measured. Two phases characterised the response to the transition from NO3- to NH4+: (1) an initial phase in which photosynthesis and anaplerosis were stimulated and protein increased; (2) a subsequent phase in which most parameters reached new values that were close to those at the beginning of the experiment (100% NO3-). The only exceptions were PEPc activity and LICF, whose rates remained at least 2-fold higher than at 100% NO3-, when NH4+ was the sole N source. The results are indicative of a tendency to re-establish homeostasis, after an initial perturbation of the intracellular composition. The roles of different metabolic processes during acclimation are discussed.

Taxonomy and growth conditions concur to determine the energetic suitability of algal fatty acid complements

We hypothesize that, in algae, carbon allocation to lipids depends on the combined effects of the metabolic constraints imposed by the genotype (i.e., species-specific differences in composition) and on the acclimation responses (phenotype) to changes in the stoichiometry of available inorganic carbon (for photosynthesis) and nitrogen (primarily for amino acids, protein, and nucleic acid synthesis). We thus cultured three taxonomically distinct algae, the green alga Dunaliella salina, the diatom Thalassiosira pseudonana, and the dinoflagellate Protoceratium reticulatum, at four nitrate concentrations and constant inorganic carbon. Since energy availability also directly impacts carbon partitioning, we studied the effect of irradiance on the oil quality of P. reticulatum. We used Fourier transform infrared (FTIR) spectroscopy to study carbon allocation and biomass reduction level and gas chromatography for fatty acid analysis. The fatty acid complements of the three species were different; within each species, growth conditions substantially altered oil quality. We ranked the oils in terms of their suitability as biodiesels, using international standards as reference. We believe that this approach may help to identify the appropriate combination of taxa and culture conditions for algal biodiesel production and in general offers insight on carbon allocation to fatty acids.

Herbivory in the soft coral Sinularia flexibilis (Alcyoniidae)

Our work provides strong support for the hypothesis that Sinularia flexibilis ingests diatoms such as Thalassiosira pseudonana. We assessed algal ingestion by S. flexibilis through estimates of algal removal, histological analyses, scanning electron microscopy observations, and gene expression determination (18S and silicon transporter 1) by real time PCR. Cell counts are strongly suggestive of algal removal by the coral; light and scanning microscopy provide qualitative evidence for the ingestion of T. pseudonana by S. flexibilis, while molecular markers did not prove to be sufficiently selective/specific to give clear results. We thus propose that previous instances of inability of corals to ingest algae are reconsidered using different technical approach, before concluding that coral herbivory is not a general feature.

Estimating and correcting interference fringes in infrared spectra in infrared hyperspectral imaging

Short-term acclimation response of individual cells of Thalassiosira weissflogii was monitored by Synchrotron FTIR imaging over the span of 75 minutes. The cells, collected from batch cultures, were maintained in a constant flow of medium, at an irradiance of 120 μmol m-2 s-1 and at 20 °C. Multiple internal reflections due to the micro fluidic channel were modeled, and showed that fringes are additive sinusoids to the pure absorption of the other components of the system. Preprocessing of the hyperspectral cube (x, y, Abs(λ)) included removing spectral fringe using an EMSC approach. Principal component analysis of the time series of hyperspectral cubes showed macromolecular pool variations (carbohydrates, lipids and DNA/RNA) of less than 2% after fringe correction.