Calcification traits for cryptic species identification: Insights into coralline biomineralization.

Calcareous red algae are foundation species and ecosystem engineers with a global distribution. The principles governing their calcification pathways are still debated and the morphological characters are frequently unreliable for species segregation, as shown by molecular genetics. The recent description of the new species Lithophyllum pseudoracemus, previously undetected and morphologically confused with Lithophyllum racemus, offered a challenging opportunity to test the effectiveness of microanatomy and ultrastructural calcification traits as tools for the identification of these two species, for integrative taxonomy. High resolution SEM images of molecularly identified samples showed that the different size of the perithallial cells and the features of the asexual conceptacle chambers may contribute to the separation of the two species. The two species share the same crystallite morphology in the primary and secondary cell-wall calcification, as previously described in other species belonging to the same clade. However, the perithallial secondary calcification was significantly thicker in L. racemus than in L. pseudoracemus. We described a granular calcified layer in the innermost part of the cell wall, as a putative precursor phase in the biomineralization and formation of the secondary calcification. The hypothesis of different pathways for the formation of the primary and secondary calcification is supported by the observed cell elongation associated with thicker and higher Mg/Ca primary calcification, the inverse correlation of primary and secondary calcification thickness, and the absence of primary calcification in the newly formed wall cutting off an epithallial cell from the meristem.

Phototaxis of the Unicellular Red Alga Cyanidioschyzon merolae Is Mediated by Novel Actin-Driven Tentacles.

Phototaxis, which is the ability to move towards or away from a light source autonomously, is a common mechanism of unicellular algae. It evolved multiple times independently in different plant lineages. As of yet, algal phototaxis has been linked mainly to the presence of cilia, the only known locomotive organelle in unicellular algae. Red algae (Rhodophyta), however, lack cilia in all stages of their life cycle. Remarkably, multiple unicellular red algae like the extremophile Cyanidioschyzon merolae (C. merolae) can move towards light. Remarkably, it has remained unclear how C. merolae achieves movement, and the presence of a completely new mechanism has been suggested. Here we show that the basis of this movement are novel retractable projections, termed tentacles due to their distinct morphology. These tentacles could be reproducibly induced within 20 min by increasing the salt concentration of the culture medium. Electron microscopy revealed filamentous structures inside the tentacles that we identified to be actin filaments. This is surprising as C. merolae's single actin gene was previously published to not be expressed. Based on our findings, we propose a model for C. merolae's actin-driven but myosin-independent motility. To our knowledge, the described tentacles represent a novel motility mechanism.

Structural basis of energy transfer in Porphyridium purpureum phycobilisome.

Photosynthetic organisms have developed various light-harvesting systems to adapt to their environments1. Phycobilisomes are large light-harvesting protein complexes found in cyanobacteria and red algae2-4, although how the energies of the chromophores within these complexes are modulated by their environment is unclear. Here we report the cryo-electron microscopy structure of a 14.7-megadalton phycobilisome with a hemiellipsoidal shape from the red alga Porphyridium purpureum. Within this complex we determine the structures of 706 protein subunits, including 528 phycoerythrin, 72 phycocyanin, 46 allophycocyanin and 60 linker proteins. In addition, 1,598 chromophores are resolved comprising 1,430 phycoerythrobilin, 48 phycourobilin and 120 phycocyanobilin molecules. The markedly improved resolution of our structure compared with that of the phycobilisome of Griffithsia pacifica5 enabled us to build an accurate atomic model of the P. purpureum phycobilisome system. The model reveals how the linker proteins affect the microenvironment of the chromophores, and suggests that interactions of the aromatic amino acids of the linker proteins with the chromophores may be a key factor in fine-tuning the energy states of the chromophores to ensure the efficient unidirectional transfer of energy.

Prediction of Sequential Organelles Localization under Imbalance using A Balanced Deep U-Net.

Assessing the structure and function of organelles in living organisms of the primitive unicellular red algae Cyanidioschyzon merolae on three-dimensional sequential images demands a reliable automated technique in the class imbalance among various cellular structures during mitosis. Existing classification networks with commonly used loss functions were focused on larger numbers of cellular structures that lead to the unreliability of the system. Hence, we proposed a balanced deep regularized weighted compound dice loss (RWCDL) network for better localization of cell organelles. Specifically, we introduced two new loss functions, namely compound dice (CD) and RWCD by implementing multi-class variant dice and weighting mechanism, respectively for maximizing weights of peroxisome and nucleus among five classes as the main contribution of this study. We extended the Unet-like convolution neural network (CNN) architecture for evaluating the ability of our proposed loss functions for improved segmentation. The feasibility of the proposed approach is confirmed with three different large scale mitotic cycle data set with different number of occurrences of cell organelles. In addition, we compared the training behavior of our designed architectures with the ground truth segmentation using various performance measures. The proposed balanced RWCDL network generated the highest area under the curve (AUC) value in elevating the small and obscure peroxisome and nucleus, which is 30% higher than the network with commonly used mean square error (MSE) and dice loss (DL) functions. The experimental results indicated that the proposed approach can efficiently identify the cellular structures, even when the contour between the cells is obscure and thus convinced that the balanced deep RWCDL approach is reliable and can be helpful for biologist to accurately identify the relationship between the cell behavior and structures of cell organelles during mitosis.

Substrate specificity of plastid phosphate transporters in a non-photosynthetic diatom and its implication in evolution of red alga-derived complex plastids.

The triose phosphate transporter (TPT) is one of the prerequisites to exchange metabolites between the cytosol and plastids. In this study, we demonstrated that the four plastid TPT homologues in the non-photosynthetic diatom Nitzschia sp. NIES-3581 were highly likely integrated into plastid envelope membranes similar to counterparts in the model photosynthetic diatom Phaeodactylum tricornutum, in terms of target membranes and C-terminal orientations. Three of the four Nitzschia TPT homologues are capable of transporting various metabolites into proteo-liposomes including triose phosphates (TPs) and phosphoenolpyruvate (PEP), the transport substrates sufficient to support the metabolic pathways retained in the non-photosynthetic diatom plastid. Phylogenetic analysis of TPTs and closely related transporter proteins indicated that diatoms and other algae with red alga-derived complex plastids possess only TPT homologues but lack homologues of the glucose 6-phosphate transporter (GPT), xylulose 5-phosphate transporter (XPT), and phosphoenolpyruvate transporter (PPT). Comparative sequence analysis suggests that many TPT homologues of red alga-derived complex plastids potentially have the ability to transport mainly TPs and PEP. TPTs transporting both TPs and PEP highly likely mediate a metabolic crosstalk between a red alga-derived complex plastid and the cytosol in photosynthetic and non-photosynthetic species, which explains the lack of PPTs in all the lineages with red alga-derived complex plastids. The PEP-transporting TPTs might have emerged in an early phase of endosymbiosis between a red alga and a eukaryote host, given the broad distribution of that type of transporters in all branches of red alga-derived complex plastid-bearing lineages, and have probably played a key role in the establishment and retention of a controllable, intracellular metabolic connection in those organisms.

Coralline algal calcification: A morphological and process-based understanding.

RESEARCH PURPOSE AND FINDINGS: Coralline algae are key biological substrates of many carbonate systems globally. Their capacity to build enduring crusts that underpin the formation of tropical reefs, rhodolith beds and other benthic substrate is dependent on the formation of a calcified thallus. However, this important process of skeletal carbonate formation is not well understood. We undertook a study of cellular carbonate features to develop a model for calcification. We describe two types of cell wall calcification; 1) calcified primary cell wall (PCW) in the thin-walled elongate cells such as central medullary cells in articulated corallines and hypothallial cells in crustose coralline algae (CCA), 2) calcified secondary cell wall (SCW) with radial Mg-calcite crystals in thicker-walled rounded cortical cells of articulated corallines and perithallial cells of CCA. The distinctive banding found in many rhodoliths is the regular transition from PCW-only cells to SCW cells. Within the cell walls there can be bands of elevated Mg with Mg content of a few mol% higher than radial Mg-calcite (M-type), ranging up to dolomite composition (D-type). MODEL FOR CALCIFICATION: We propose the following three-step model for calcification. 1) A thin (< 0.5 µm) PCW forms and is filled with a mineralising fluid of organic compounds and seawater. Nanometer-scale Mg-calcite grains precipitate on the organic structures within the PCW. 2) Crystalline cellulose microfibrils (CMF) are extruded perpendicularly from the cellulose synthase complexes (CSC) in the plasmalemma to form the SCW. 3) The CMF soaks in the mineralising fluid as it extrudes and becomes calcified, retaining the perpendicular form, thus building the radial calcite. In Clathromorphum, SCW formation lags PCW creating a zone of weakness resulting in a split in the sub-surface crust. All calcification seems likely to be a bioinduced rather than controlled process. These findings are a substantial step forward in understanding how corallines calcify.

Selective loss of photosystem I and formation of tubular thylakoids in heterotrophically grown red alga Cyanidioschyzon merolae.

We previously found that glycerol is required for heterotrophic growth in the unicellular red alga Cyanidioschyzon merolae. Here, we analyzed heterotrophically grown cells in more detail. Sugars or other organic substances did not support the growth in the dark. The growth rate was 0.4 divisions day-1 in the presence of 400 mM glycerol, in contrast with 0.5 divisions day-1 in the phototrophic growth. The growth continued until the sixth division. Unlimited heterotrophic growth was possible in the medium containing DCMU and glycerol in the light. Light-activated heterotrophic culture in which cells were irradiated by intermittent light also continued without an apparent limit. In the heterotrophic culture in the dark, chlorophyll content drastically decreased, as a result of inability of dark chlorophyll synthesis. Photosynthetic activity gradually decreased over 10 days, and finally lost after 19 days. Low-temperature fluorescence measurement and immunoblot analysis showed that this decline in photosynthetic activity was mainly due to the loss of Photosystem I, while the levels of Photosystem II and phycobilisomes were maintained. Accumulated triacylglycerol was lost during the heterotrophic growth, while keeping the overall lipid composition. Observation by transmission electron microscopy revealed that a part of thylakoid membranes turned into pentagonal tubular structures, on which five rows of phycobilisomes were aligned. This might be a structure that compactly conserve phycobilisomes and Photosystem II in an inactive state, probably as a stock of carbon and nitrogen. These results suggest that C. merolae has a unique strategy of heterotrophic growth, distinct from those found in other red algae.

The cellular machineries responsible for the division of endosymbiotic organelles.

Chloroplasts (plastids) and mitochondria evolved from endosymbiotic bacteria. These organelles perform vital functions in photosynthetic eukaryotes, such as harvesting and converting energy for use in biological processes. Consistent with their evolutionary origins, plastids and mitochondria proliferate by the binary fission of pre-existing organelles. Here, I review the structures and functions of the supramolecular machineries driving plastid and mitochondrial division, which were discovered and first studied in the primitive red alga Cyanidioschyzon merolae. In the past decade, intact division machineries have been isolated from plastids and mitochondria and examined to investigate their underlying structure and molecular mechanisms. A series of studies has elucidated how these division machineries assemble and transform during the fission of these organelles, and which of the component proteins generate the motive force for their contraction. Plastid- and mitochondrial-division machineries have important similarities in their structures and mechanisms despite sharing no component proteins, implying that these division machineries evolved in parallel. The establishment of these division machineries might have enabled the host eukaryotic ancestor to permanently retain these endosymbiotic organelles by regulating their binary fission and the equal distribution of resources to daughter cells. These findings provide key insights into the establishment of endosymbiotic organelles and have opened new avenues of research into their evolution and mechanisms of proliferation.

Unique organization of photosystem I-light-harvesting supercomplex revealed by cryo-EM from a red alga.

Photosystem I (PSI) is one of the two photosystems present in oxygenic photosynthetic organisms and functions to harvest and convert light energy into chemical energy in photosynthesis. In eukaryotic algae and higher plants, PSI consists of a core surrounded by variable species and numbers of light-harvesting complex (LHC)I proteins, forming a PSI-LHCI supercomplex. Here, we report cryo-EM structures of PSI-LHCR from the red alga Cyanidioschyzon merolae in two forms, one with three Lhcr subunits attached to the side, similar to that of higher plants, and the other with two additional Lhcr subunits attached to the opposite side, indicating an ancient form of PSI-LHCI. Furthermore, the red algal PSI core showed features of both cyanobacterial and higher plant PSI, suggesting an intermediate type during evolution from prokaryotes to eukaryotes. The structure of PsaO, existing in eukaryotic organisms, was identified in the PSI core and binds three chlorophylls a and may be important in harvesting energy and in mediating energy transfer from LHCII to the PSI core under state-2 conditions. Individual attaching sites of LHCRs with the core subunits were identified, and each Lhcr was found to contain 11 to 13 chlorophylls a and 5 zeaxanthins, which are apparently different from those of LHCs in plant PSI-LHCI. Together, our results reveal unique energy transfer pathways different from those of higher plant PSI-LHCI, its adaptation to the changing environment, and the possible changes of PSI-LHCI during evolution from prokaryotes to eukaryotes.

Glycosyltransferase MDR1 assembles a dividing ring for mitochondrial proliferation comprising polyglucan nanofilaments.

Mitochondria, which evolved from a free-living bacterial ancestor, contain their own genomes and genetic systems and are produced from preexisting mitochondria by binary division. The mitochondrion-dividing (MD) ring is the main skeletal structure of the mitochondrial division machinery. However, the assembly mechanism and molecular identity of the MD ring are unknown. Multi-omics analysis of isolated mitochondrial division machinery from the unicellular alga Cyanidioschyzon merolae revealed an uncharacterized glycosyltransferase, MITOCHONDRION-DIVIDING RING1 (MDR1), which is specifically expressed during mitochondrial division and forms a single ring at the mitochondrial division site. Nanoscale imaging using immunoelectron microscopy and componential analysis demonstrated that MDR1 is involved in MD ring formation and that the MD ring filaments are composed of glycosylated MDR1 and polymeric glucose nanofilaments. Down-regulation of MDR1 strongly interrupted mitochondrial division and obstructed MD ring assembly. Taken together, our results suggest that MDR1 mediates the synthesis of polyglucan nanofilaments that assemble to form the MD ring. Given that a homolog of MDR1 performs similar functions in chloroplast division, the establishment of MDR1 family proteins appears to have been a singular, crucial event for the emergence of endosymbiotic organelles.

Structure of phycobilisome from the red alga Griffithsia pacifica.

Life on Earth depends on photosynthesis for its conversion of solar energy to chemical energy. Photosynthetic organisms have developed a variety of light-harvesting systems to capture sunlight. The largest light-harvesting complex is the phycobilisome (PBS), the main light-harvesting antenna in cyanobacteria and red algae. It is composed of phycobiliproteins and linker proteins but the assembly mechanisms and energy transfer pathways of the PBS are not well understood. Here we report the structure of a 16.8-megadalton PBS from a red alga at 3.5 Å resolution obtained by single-particle cryo-electron microscopy. We modelled 862 protein subunits, including 4 linkers in the core, 16 rod-core linkers and 52 rod linkers, and located a total of 2,048 chromophores. This structure reveals the mechanisms underlying specific interactions between linkers and phycobiliproteins, and the formation of linker skeletons. These results provide a firm structural basis for our understanding of complex assembly and the mechanisms of energy transfer within the PBS.

Are Thraustochytrids algae?

Thraustochytrids, a heterotrophic fungus-like clade of Stramenopiles, are becoming an increasingly important source of polyunsaturated fatty acids (PUFAs) for biotechnological industries. PUFA rich oils from these organisms are subsequently referred to in some literature and marketing sources as being derived from 'algae', in spite of their non-photosynthetic source organism. In this review, we attempt to disentangle the evolutionary relationship of the Thraustochytrids from other Protists, demonstrating that there is no scientific basis for the aforementioned misnomer. Some research has previously suggested that the ancestor of the Stramenopiles may have been photosynthetic, and subsequently lost their plastids in multiple lineages. The placement of the Thraustochytrids within the Stramenopiles and their possible plastid loss may have been a source of obfuscation. It is becoming increasingly evident that the common ancestor of the Stramenopiles was not photosynthetic, and that only the Ochrophyte lineage later engulfed a plastid via higher order endosymbiosis. Because all basal lineages of Stramenopiles are non-plastidial heterotrophs, including the Thraustochytrids, there remains no phylogenetic, biological, or ecological justification for the term 'algae' to be applied to Thraustochytrids or their products.

Multiple phases of mg-calcite in crustose coralline algae suggest caution for temperature proxy and ocean acidification assessment: lessons from the ultrastructure and biomineralization in Phymatolithon (Rhodophyta, Corallinales)1.

Magnesium content, strongly correlated with temperature, has been developed as a climate archive for the late Holocene without considering anatomical controls on Mg content. In this paper, we explore the ultrastructure and cellular scale Mg-content variations within four species of North Atlantic crust-forming Phymatolithon. The cell wall has radial grains of Mg-calcite, whereas the interfilament (middle lamella) has grains aligned parallel to the filament axis. The proportion of interfilament and cell wall carbonate varies by tissue and species. Three distinct primary phases of Mg-calcite were identified: interfilament Mg-calcite (mean 8.9 mol% MgCO3 ), perithallial cell walls Mg-calcite (mean 13.4 mol% MgCO3 ), and hypothallium Mg-calcite (mean 17.1 mol% MgCO3 ). Magnesium content for the bulk crust, an average of all phases present, showed a strongly correlated (R2  = 0.975) increase of 0.31 mol% MgCO3 per °C. Of concern for climate reconstructions is the potential for false warming signals from undetected postgrazing wound repair carbonate that is substantially enriched in Mg, unrelated to temperature. Within a single crust, Mg-content of component carbonates ranged from 8 to 20 mol% MgCO3 , representing theoretical thermodynamic stabilities from aragonite-equivalent to unstable higher-Mg-calcite. It is unlikely that existing current predictions of ocean acidification impact on coralline algae, based on saturation states calculated using average Mg contents, provide an environmentally relevant estimate.

Three-dimensional preservation of cellular and subcellular structures suggests 1.6 billion-year-old crown-group red algae.

The ~1.6 Ga Tirohan Dolomite of the Lower Vindhyan in central India contains phosphatized stromatolitic microbialites. We report from there uniquely well-preserved fossils interpreted as probable crown-group rhodophytes (red algae). The filamentous form Rafatazmia chitrakootensis n. gen, n. sp. has uniserial rows of large cells and grows through diffusely distributed septation. Each cell has a centrally suspended, conspicuous rhomboidal disk interpreted as a pyrenoid. The septa between the cells have central structures that may represent pit connections and pit plugs. Another filamentous form, Denaricion mendax n. gen., n. sp., has coin-like cells reminiscent of those in large sulfur-oxidizing bacteria but much more recalcitrant than the liquid-vacuole-filled cells of the latter. There are also resemblances with oscillatoriacean cyanobacteria, although cell volumes in the latter are much smaller. The wider affinities of Denaricion are uncertain. Ramathallus lobatus n. gen., n. sp. is a lobate sessile alga with pseudoparenchymatous thallus, "cell fountains," and apical growth, suggesting florideophycean affinity. If these inferences are correct, Rafatazmia and Ramathallus represent crown-group multicellular rhodophytes, antedating the oldest previously accepted red alga in the fossil record by about 400 million years.

The Chloroplast Tubulin Homologs FtsZA and FtsZB from the Red Alga Galdieria sulphuraria Co-assemble into Dynamic Filaments.

FtsZ is a homolog of eukaryotic tubulin and is present in almost all bacteria and many archaea, where it is the major cytoskeletal protein in the Z ring, required for cell division. Unlike some other cell organelles of prokaryotic origin, chloroplasts have retained FtsZ as an essential component of the division machinery. However, chloroplast FtsZs have been challenging to study because they are difficult to express and purify. To this end, we have used a FATT tag expression system to produce as soluble proteins the two chloroplast FtsZs from Galdieria sulphuraria, a thermophilic red alga. GsFtsZA and GsFtsZB assembled individually in the presence of GTP, forming large bundles of protofilaments. GsFtsZA also assembled in the presence of GDP, the first member of the FtsZ/tubulin superfamily to do so. Mixtures of GsFtsZA and GsFtsZB assembled protofilament bundles and hydrolyzed GTP at a rate approximately equal to the sum of their individual rates, suggesting a random co-assembly. GsFtsZA assembly by itself in limiting GTP gave polymers that remained stable for a prolonged time. However, when GsFtsZB was added, the co-polymers disassembled with enhanced kinetics, suggesting that the GsFtsZB regulates and enhances disassembly dynamics. GsFtsZA-mts (where mts is a membrane-targeting amphipathic helix) formed Z ring-like helices when expressed in Escherichia coli Co-expression of GsFtsZB (without an mts) gave co-assembly of both into similar helices. In summary, we provide biochemical evidence that GsFtsZA assembles as the primary scaffold of the chloroplast Z ring and that GsFtsZB co-assembly enhances polymer disassembly and dynamics.

Genetic and morphological variation in an ecosystem engineer, Lithophyllum byssoides (Corallinales, Rhodophyta).

Lithophyllum byssoides is a common coralline alga in the intertidal zone of Mediterranean coasts, where it produces biogenic concretions housing a high algal and invertebrate biodiversity. This species is an ecosystem engineer and is considered a target for conservation efforts, but designing effective conservation strategies currently is impossible due to lack of information about its population structure. The morphological and molecular variation of L. byssoides was investigated using morphoanatomy and DNA sequences (psbA and cox2,3) obtained from populations at 15 localities on the Italian and Croatian coasts. Lithophyllum byssoides exhibited a high number of haplotypes (31 psbA haplotypes and 24 cox2,3 haplotypes) in the central Mediterranean. The psbA and cox2,3 phylogenies were congruent and showed seven lineages. For most of these clades, the distribution was limited to one or a few localities, but one of them (clade 7) was widespread across the central Mediterranean, spanning the main biogeographic boundaries recognized in this area. The central Mediterranean populations formed a lineage separate from Atlantic samples; psbA pair-wise divergences suggested that recognition of Atlantic and Mediterranean L. byssoides as different species may be appropriate. The central Mediterranean haplotype patterns of L. byssoides were interpreted as resulting from past climatic events in the hydrogeological history of the Mediterranean Sea. The high haplotype diversity and the restricted spatial distribution of the seven lineages suggest that individual populations should be managed as independent units.

Supramolecular architecture of photosynthetic membrane in red algae in response to nitrogen starvation.

The availability of nitrogen is one of the most important determinants that can limit the growth of photosynthetic organisms including plants and algae; however, direct observations on the supramolecular architecture of photosynthetic membranes in response to nitrogen stress are still lacking. Red algae are an important evolutionary group of algae which contain phycobilisomes (PBSs) on their thylakoid membranes, as do cyanobacteria. PBSs function not only as light-harvesting antennae but also as nitrogen storage. In this report, alterations of the supramolecular architecture of thylakoid membranes from red alga Porphyridium cruentum during nitrogen starvation were characterized. The morphology of the intact thylakoid membrane was observed to be round vesicles. Thylakoid membranes were reduced in content and PBSs were degraded during nitrogen starvation. The size and density of PBSs were both found to be reduced. PBS size decreased by less than one-half after 20days of nitrogen starvation, but their hemispherical morphology was retained. The density of PBSs on thylakoid membranes was more seriously affected as time proceeded. Upon re-addition of nitrogen led to increasing of PBSs on thylakoid membranes. This work reports the first direct observation on alterations in the supramolecular architecture of thylakoid membranes from a photosynthetic organism in response to nitrogen stress.

The extraordinary joint material of an articulated coralline alga. II. Modeling the structural basis of its mechanical properties.

By incorporating joints into their otherwise rigid fronds, erect coralline algae have evolved to be as flexible as other seaweeds, which allows them to thrive - and even dominate space - on wave-washed shores around the globe. However, to provide the required flexibility, the joint tissue of Calliarthron cheilosporioides, a representative articulated coralline alga, relies on an extraordinary tissue that is stronger, more extensible and more fatigue resistant than that of other algae. Here, we used the results from recent experiments to parameterize a conceptual model that links the microscale architecture of cell walls to the adaptive mechanical properties of joint tissue. Our analysis suggests that the theory of discontinuous fiber-wound composite materials (with cellulose fibrils as the fibers and galactan gel as the matrix) can explain key aspects of the material's mechanics. In particular, its adaptive viscoelastic behavior can be characterized by two, widely separated time constants. We speculate that the short time constant (∼14 s) results from the viscous response of the matrix to the change in cell-wall shape as a joint is stretched, a response that allows the material both to remain flexible and to dissipate energy as a frond is lashed by waves. We propose that the long time constant (∼35 h), is governed by the shearing of the matrix between cellulose fibrils. The resulting high apparent viscosity ensures that joints avoid accumulating lethal deformation in the course of a frond's lifetime. Our synthesis of experimental measurements allows us to draw a chain of mechanistic inference from molecules to cell walls to fronds and community ecology.

Primitive red alga Cyanidioschyzon merolae accumulates storage glucan and triacylglycerol under nitrogen depletion.

Most microalgae accumulate neutral lipids, including triacylglycerol (TAG), into spherical structures called lipid bodies (LBs) under environmental stress conditions such as nutrient depletion. In green algae, starch accumulation precedes TAG accumulation, and the starch is thought to be a substrate for TAG synthesis. However, the relationship between TAG synthesis and the starch content in red algae, as well as how TAG accumulation is regulated, is unclear. In this study, we cultured the primitive red alga Cyanidioschyzon merolae under nitrogen-depleted conditions, and monitored the formation of starch granules (SGs) and LBs using microscopy. SGs stained with potassium iodide were observed at 24 h; however, LBs stained specifically with BODIPY 493/503 were observed after 48 h. Quantitative analysis of neutral sugar and cytomorphological semi-quantitative analysis of TAG accumulation also supported these results. Thus, the accumulation of starch occurred and preceded the accumulation of TAG in cells of C. merolae. However, TAG accumulation was not accompanied by a decrease in the starch content, suggesting that the starch is a major carbon storage sink, at least under nitrogen-depleted conditions. Quantitative real-time PCR revealed that the mRNA levels of genes involved in starch and TAG synthesis rarely changed during the culture period, suggesting that starch and TAG synthesis in C. merolae are not controlled through gene transcription but at other stages, such as translation and/or enzymatic activity.

Aragonite infill in overgrown conceptacles of coralline Lithothamnion spp. (Hapalidiaceae, Hapalidiales, Rhodophyta): new insights in biomineralization and phylomineralogy.

New empirical and quantitative data in the study of calcium carbonate biomineralization and an expanded coralline psbA framework for phylomineralogy are provided for crustose coralline red algae. Scanning electron microscopy (SEM) and energy dispersive spectrometry (SEM-EDS) pinpointed the exact location of calcium carbonate crystals within overgrown reproductive conceptacles in rhodolith-forming Lithothamnion species from the Gulf of Mexico and Pacific Panama. SEM-EDS and X-ray diffraction (XRD) analysis confirmed the elemental composition of these calcium carbonate crystals to be aragonite. After spore release, reproductive conceptacles apparently became overgrown by new vegetative growth, a strategy that may aid in sealing the empty conceptacle chamber, hence influencing the chemistry of the microenvironment and in turn promoting aragonite crystal growth. The possible relevance of various types of calcium carbonate polymorphs present in the complex internal structure and skeleton of crustose corallines is discussed. This is the first study to link SEM, SEM-EDS, XRD, Microtomography and X-ray microscopy data of aragonite infill in coralline algae with phylomineralogy. The study contributes to the growing body of literature characterizing and speculating about how the relative abundances of carbonate biominerals in corallines may vary in response to changes in atmospheric pCO2 , ocean acidification, and global warming.