RESUMEN

Cellular and physiological cycles are driven by endogenous pacemakers, the diurnal and circadian rhythms. Key functions such as cell cycle progression and cellular metabolism are under rhythmic regulation, thereby maintaining physiological homeostasis. The photoreceptors phytochrome and cryptochrome, in response to light cues, are central input pathways for physiological cycles in most photosynthetic organisms. However, among Archaeplastida, red algae are the only taxa that lack phytochromes. Current knowledge about oscillatory rhythms is primarily derived from model species such as Arabidopsis thaliana and Chlamydomonas reinhardtii in the Viridiplantae, whereas little is known about these processes in other clades of the Archaeplastida, such as the red algae (Rhodophyta). We used genome-wide expression profiling of the red seaweed Gracilariopsis chorda and identified 3,098 rhythmic genes. Here, we characterized possible cryptochrome-based regulation and photosynthetic/cytosolic carbon metabolism in this species. We found a large family of cryptochrome genes in G. chorda that display rhythmic expression over the diurnal cycle and may compensate for the lack of phytochromes in this species. The input pathway gates regulatory networks of carbon metabolism which results in a compact and efficient energy metabolism during daylight hours. The system in G. chorda is distinct from energy metabolism in most plants, which activates in the dark. The green lineage, in particular, land plants, balance water loss and CO2 capture in terrestrial environments. In contrast, red seaweeds maintain a reduced set of photoreceptors and a compact cytosolic carbon metabolism to thrive in the harsh abiotic conditions typical of intertidal zones.

Asunto(s)

Arabidopsis , Rhodophyta , Algas Marinas , Algas Marinas/genética , Criptocromos/metabolismo , Rhodophyta/genética , Ritmo Circadiano/genética , Arabidopsis/genética

RESUMEN

In this study, we studied the bioactive peptides produced by thermolysin hydrolysis of a water-soluble protein (WSP) from the red alga Gracilariopsis chorda, whose major components are phycobiliproteins and Ribulose-1,5-bisphosphate carboxylase-oxygenase (RuBisCo). The results showed that WSP hydrolysate exhibited significantly higher ACE inhibitory activity (92% inhibition) compared to DPP-IV inhibitory activity and DPPH scavenging activity. The phycobiliproteins and RuBisCo of G. chorda contain a high proportion of hydrophobic (31.0-46.5%) and aromatic (5.1-46.5%) amino acid residues, which was considered suitable for the formation of peptides with strong ACE inhibitory activity. Therefore, we searched for peptides with strong ACE inhibitory activity and identified two novel peptides (IDHY and LVVER). Then, their interaction with human ACE was evaluated by molecular docking, and IDHY was found to be a promising inhibitor. In silico analysis was then performed on the structural factors affecting ACE inhibitory peptide release, using the predicted 3D structures of phycobiliproteins and RuBisCo. The results showed that most of the ACE inhibitory peptides are located in the highly solvent accessible α-helix. Therefore, it was suggested that G. chorda is a good source of bioactive peptides, especially ACE-inhibitory peptides.

Asunto(s)

Rhodophyta , Ribulosa-Bifosfato Carboxilasa , Humanos , Simulación del Acoplamiento Molecular , Péptidos/química , Rhodophyta/metabolismo , Ficobiliproteínas , Peptidil-Dipeptidasa A/química

RESUMEN

Carbonic anhydrase (CA; EC 4.2.2.1) is a Zn-binding metalloenzyme that catalyzes the reversible hydration of CO2. Recently, CAs have gained a great deal of attention as biocatalysts for capturing CO2 from industrial flue gases owing to their extremely fast reaction rates and simple reaction mechanism. However, their general application for this purpose requires improvements to stability at high temperature and under in vitro conditions, and reductions in production and scale-up costs. In the present study, we developed a strategy for producing GcCAα3, a CA isoform from the red alga Gracilariopsis chorda, in Nicotiana benthamiana. To achieve high-level expression and facile purification of GcCAα3, we designed various constructs by incorporating various domains such as translation-enhancing M domain, SUMO domain and cellulose-binding domain CBM3. Of these constructs, MC-GcCAα3 that had the M and CBM3 domains was expressed at high levels in N. benthamiana via agroinfiltration with a yield of 1.0 g/kg fresh weight. The recombinant protein was targeted to the endoplasmic reticulum (ER) for high-level accumulation in plants. Specific and tight CBM3-mediated binding of recombinant GcCAα3 proteins to microcrystalline cellulose beads served as a means for both protein purification from total plant extracts and protein immobilization to a solid surface for increased stability, facilitating multiple rounds of use in CO2 hydration reactions.

RESUMEN

KEY MESSAGE: Red alga, Gracilariopsis chorda, contains seven carbonic anhydrases that can be grouped into α-, ß- and γ-classes. Carbonic anhydrases (CAHs) are metalloenzymes that catalyze the reversible hydration of CO2. These enzymes are present in all living organisms and play roles in various cellular processes, including photosynthesis. In this study, we identified seven CAH genes (GcCAHs) from the genome sequence of the red alga Gracilariopsis chorda and characterized them at the molecular, cellular and biochemical levels. Based on sequence analysis, these seven isoforms were categorized into four α-class, one ß-class, and two γ-class isoforms. RNA sequencing revealed that of the seven CAHs isoforms, six genes were expressed in G. chorda in light at room temperature. In silico analysis revealed that these seven isoforms localized to multiple subcellular locations such as the ER, mitochondria and cytosol. When expressed as green fluorescent protein fusions in protoplasts of Arabidopsis thaliana leaf cells, these seven isoforms showed multiple localization patterns. The four α-class GcCAHs with an N-terminal hydrophobic leader sequence localized to the ER and two of them were further targeted to the vacuole. GcCAHß1 with no noticeable signal sequence localized to the cytosol. The two γ-class GcCAHs also localized to the cytosol, despite the presence of a predicted presequence. Based on these results, we propose that the red alga G. chorda also employs multiple CAH isoforms for various cellular processes such as photosynthesis.

Asunto(s)

Arabidopsis/genética , Anhidrasas Carbónicas/metabolismo , Rhodophyta/enzimología , Anhidrasas Carbónicas/clasificación , Simulación por Computador , Retículo Endoplásmico/metabolismo , Regulación Enzimológica de la Expresión Génica , Glicosilación , Aparato de Golgi/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Filogenia , Plantas Modificadas Genéticamente , Transporte de Proteínas , Protoplastos/metabolismo , Fracciones Subcelulares/metabolismo , Vacuolas/metabolismo

RESUMEN

Exogenous neurotrophins can induce neuronal differentiation, outgrowth, survival, and synaptic function in the central nervous system. In primary cultures of rat hippocampal neurons, an ethanol extract of the red alga Gracilariopsis chorda (GCE) and its active compound arachidonic acid (AA) significantly increased the densities of dendritic filopodia and spines, promoted the expression of presynaptic vesicle protein 2 (SV2) and postsynaptic density protein 95 (PSD-95), induced robust synaptogenesis, and increased the expression of cell division control protein 42 (CDC42) and actin-related protein 2 (ARP2), which are important for actin organization in dendritic protrusions, and facilitated presynaptic plasticity by increasing the size of the synaptic vesicle pool at presynaptic nerve terminals. In addition, oral administration of GCE and AA for 10 days, at concentrations of 1 mg/g and 2.2 µg/g body weight, respectively, significantly protected against scopolamine-induced memory impairment in mice by increasing the latency time in the passive avoidance test. These results provide strong scientific evidence that these natural products can be used as neurotrophic substances and/or dietary supplements for the prevention and treatment of memory-related neurological disorders via the reconstruction of axo-dendrites and its synapses.

Asunto(s)

Ácido Araquidónico/farmacología , Gracilaria/química , Hipocampo/efectos de los fármacos , Plasticidad Neuronal/efectos de los fármacos , Neuronas/efectos de los fármacos , Seudópodos/efectos de los fármacos , Proteína 2 Similar a la Angiopoyetina , Proteínas Similares a la Angiopoyetina/genética , Proteínas Similares a la Angiopoyetina/metabolismo , Animales , Células Cultivadas , Cognición/efectos de los fármacos , Hipocampo/citología , Trastornos de la Memoria/inducido químicamente , Trastornos de la Memoria/tratamiento farmacológico , Ratones , Ratones Endogámicos ICR , Neurogénesis/efectos de los fármacos , Neuronas/citología , Extractos Vegetales/farmacología , Proteínas/genética , Proteínas/metabolismo , Ratas , Ratas Sprague-Dawley , Escopolamina/toxicidad

RESUMEN

The edible red seaweed Gracilariopsis chorda (Holmes) Ohmi is known for its extensive medicinal benefits and its use as a food ingredient in Korea, Japan, and China. In a previous study, an ethanol extract of G. chorda (GCE) showed potential neuroprotective effects in cultured hippocampal neurons. In this study, we further examined the ability of GCE to promote neurite extension in primary rat hippocampal neurons. Neurons were stained with the lipophilic dye DiO or immunostained to visualize the neuronal morphology. The results indicated that GCE concentration-dependently increased neurite outgrowth, with an optimal concentration of 30 µg/mL. GCE significantly promoted early neuronal differentiation (i.e., polarity and process number) and enhanced axonal and dendritic arborization in a time-responsive manner. In addition, arachidonic acid, which was previously identified and quantified as a major neuroprotective component of GCE, significantly accelerated neurite outgrowth similar to GCE. Our findings suggest that G. chorda and its active component, arachidonic acid, may be useful for developing medicinal food or pharmaceuticals in the prevention and treatment of neurological disorders.

Asunto(s)

Axones/efectos de los fármacos , Gracilaria/química , Hipocampo/ultraestructura , Plasticidad Neuronal/efectos de los fármacos , Neuronas/ultraestructura , Extractos Vegetales/farmacología , Animales , Ácido Araquidónico/farmacología , Axones/ultraestructura , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Femenino , Hipocampo/embriología , Embarazo , Ratas , Ratas Sprague-Dawley , República de Corea

RESUMEN

Age-related neurological disorders are of growing concern among the elderly, and natural products with neuroprotective properties have been attracting increasing attention as candidates for the prevention or treatment of neurological disorders induced by oxidative stress. In an effort to explore natural resources, we collected some common marine seaweed from the Korean peninsula and Indonesia and screened them for neuroprotective activity against hypoxia/reoxygenation (H/R)-induced oxidative stress. Of the 23 seaweeds examined, the ethanol extract of Gracilariopsis chorda (GCE) provided maximum neuroprotection at an optimum concentration of 15 µg/mL, followed by Undaria pinnatifida. GCE increased cell viability after H/R, decreased the formation of reactive oxygen species (measured by 2',7'-dichlorodihydrofluorescein diacetate [DCF-DA] staining), and inhibited the double-stranded DNA breaks (measured by H2AX immunocytochemistry), apoptosis (measured by Annexin V/propidium iodide staining), internucleosomal DNA fragmentation (measured by DNA laddering), and dissipation of mitochondrial membrane potential (measured by JC-1 staining). Using reverse-phase high-pressure liquid chromatography, we quantitated the arachidonic acid (AA) in GCE, which provides neuroprotection against H/R-induced oxidative stress. This neuroprotective effect of AA was comparable to that of GCE. These findings suggest that the neuroprotective effect of GCE against H/R-induced neuronal death is due, at least in part, to the AA content that suppresses neuronal apoptosis.

Asunto(s)

Ácido Araquidónico/farmacología , Hipocampo/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Estrés Oxidativo/efectos de los fármacos , Daño por Reperfusión/metabolismo , Rhodophyta/química , Algas Marinas/química , Animales , Apoptosis/efectos de los fármacos , Ácido Araquidónico/análisis , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Fragmentación del ADN/efectos de los fármacos , Hipocampo/citología , Hipocampo/metabolismo , Hipoxia , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiología , Neuronas/efectos de los fármacos , Oxígeno/metabolismo , Fitoterapia , Extractos Vegetales/química , Extractos Vegetales/farmacología , Plantas Comestibles , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Daño por Reperfusión/tratamiento farmacológico , Undaria

RESUMEN

Abstract We sequenced the complete mitochondrial genome of Gracilariopsis chorda (Gracilariales, Rhodophyta), which is an agar-producing economic red algal species distributed in the northwest Pacific. The mitogenome is 26,534 bp in length with 27.7% GC content that consists of 52 genes including 26 protein-coding, 2 rRNA and 24 tRNA genes. We compared G. chorda mitogenome with that of recently published G. lemaneiformis. Nucleotide sequence similarity between these two mitogenomes was 99.82%, however, there was significant difference in length caused by trnR trnS and trnY genes missing in G. lemaneiformis.

Asunto(s)

Agar/metabolismo , Genoma Mitocondrial , Rhodophyta/citología , Rhodophyta/genética , Composición de Base , ADN de Algas/análisis , ADN Mitocondrial/análisis , Evolución Molecular , Genes Mitocondriales , Rhodophyta/clasificación , Rhodophyta/metabolismo , Análisis de Secuencia de ADN