Shota Kato

Plant-based agricultural systems rely heavily on inorganic nitrogen (N) fertilizers to increase yields and ensure food security for the rapidly growing global population. However, the production and overuse of synthetic fertilizers lead to significant amounts of greenhouse gas emissions, causing a critical need for the development of alternative and sustainable organic N fertilizers. Here, we demonstrate the effective use of lysed and dried bacterial biomass from the marine purple photosynthetic bacterium Rhodovulum sulfidophilum as an alternative source of N fertilizer for the cultivation of Japanese mustard spinach (komatsuna, Brassica rapa var. perviridis). To assess the suitability of this processed bacterial biomass, containing approximately 11% N, as a N fertilizer, we examined the effects of its application on plant germination and growth (measured by leaf chlorophyll, maximum leaf length, and dry weight) under two different temperature regimes, comparing it to that of a conventional N-containing mineral fertilizer. Application at rates up to four times that of the mineral fertilizer had no negative effects on seed germination and plant growth. The bacterial biomass had to be applied at approximately twice the rate of mineral fertilizer to obtain similar plant growth parameters, roughly corresponding to the predicted 62% rate of mineralization of the bacterial N. Our findings confirm the ability of plants to take up N from the lysed and dried biomass of marine purple photosynthetic bacteria, demonstrating the potential for using R. sulfidophilum as a source of N fertilizer.

In unicellular photosynthetic organisms, circadian rhythm is tightly linked to gating of cell cycle progression, and is entrained by light signal. As several organisms obtain a fitness advantage when the external light/dark cycle matches their endogenous period, and aging alters circadian rhythms, senescence phenotypes of the microalga Euglena gracilis of different culture ages were characterized with respect to the cell division cycle. We report here the effects of prolonged-stationary-phase conditions on the cell division cycles of E. gracilis under non-24-h light/dark cycles (T-cycles). Under T-cycles, cells established from 1-month-old and 2-month-old cultures produced lower cell concentrations after cultivation in the fresh medium than cells from 1-week-old culture. This decrease was not due to higher concentrations of dead cells in the populations, suggesting that cells of different culture ages differ in their capacity for cell division. Cells from 1-week-old cultures had a shorter circadian period of their cell division cycle under shortened T-cycles than aged cells. When algae were transferred to free-running conditions after entrainment to shortened T-cycles, the young cells showed the peak growth rate at a time corresponding to the first subjective night, but the aged cells did not. This suggests that circadian rhythms are more plastic in younger E. gracilis cells.

Gravitaxis is one of the most important issues in the growth of microalgae in the water column; it determines how easily cells receive sunlight with a comfortable intensity that is below the damaging threshold. We quantitatively investigated and analyzed the gravitaxis and cell multiplication of Euglena gracilis using vertically placed microchambers containing a single cell. A temporal change in gravitaxis and cell multiplication was observed after transferring the cells to fresh culture medium for 9 days. We performed 29 individual experiments with 2.5 mm × 2.5 mm × 0.1 mm square microchambers and found that the cells showed positive, negative, and moderate gravitaxis in 8, 7, and 14 cases, respectively, after transferring to fresh culture medium. A common trend was observed for the temporal change in gravitaxis for the eight initially positive gravitaxis cases. The cells with initially positive gravitaxis showed a higher rate of cell multiplication than those with initially negative gravitaxis. We also discussed the gravitaxis mechanism of E. gracilis from the observed trend of gravitaxis change and swimming traces. In addition, bioconvection in a larger and thicker chamber was investigated at a millimeter scale and visualized.

Salt ions promote heterogeneous localization of charged molecules toward the interface of aqueous microdroplets in oil.

Carotenoids are the most universal and most widespread pigments in nature. They have played pivotal roles in the evolution of photosensing mechanisms in microbes and of vision in animals. Several groups of phytoflagellates developed a photoreceptive organelle called the eyespot apparatus (EA) consisting of two separable components: the eyespot, a cluster of carotenoid‐rich globules that acts as a reflector device, and actual photoreceptors for photobehaviors. Unlike other algal eyespots, the eyespot of Euglenophyta lacks reflective properties and is generally considered to act as a shading device for the photoreceptor (paraflagellar body, PFB) for major photomovements. However, the function of the eyespot of Euglenophyta has not yet been fully proven. Here, we report that the blocking carotenoid biosynthesis in Euglena gracilis by suppressing the phytoene synthase gene (crtB) caused a defect in eyespot function resulting in a loss of phototaxis. Raman spectroscopy and transmission electron microscopy suggested that EgcrtB‐suppressed cells formed eyespot globules but had a defect in the accumulation of carotenoids in those packets. Motion analysis revealed the loss of phototaxis in EgcrtB‐suppressed cells: a defect in the initiation of turning movements immediately after a change in light direction, rather than a defect in the termination of cell turning at the appropriate position due to a loss of the shading effect on the PFB. This study revealed that carotenoids are essential for light perception by the EA for the initiation of phototactic movement by E. gracilis, suggesting one possible photosensory role of carotenoids in the EA for the phototaxis.

鞭毛を持ち水中を遊泳する藻類の多くは至適な光環境下で生育するための光走性や光驚動等の光運動反応と呼ばれる応答を示す．微細藻類の中では緑藻とクロムアルベオラータ，そしてエクスカベートに属する光独立栄養性のユーグレナ（Euglenophyta）が三次元方向の光走性を示すことが知られている．多くの場合光走性を示す藻類は「eyespot（眼点）」と呼ばれる器官を有する．「眼点」という名称がついているものの実際の光受容体は眼点の近傍の鞭毛や細胞膜に局在する．眼点はカロテノイドを高濃度に集積した顆粒の集合体であり，鞭毛もしくは鞭毛の運動面の近傍に形成される．この狭義の眼点と実際の光受容体によって藻類の光受容器官である「eyespot apparatus（EA）」が構成される．真核藻類の光走性やEAは分類群ごとに独自に獲得されたと考えられ，眼点の形成位置や形態，特性も系統によって大きく異なる．Euglenophytaは眼点や光走性の研究における主要なモデル生物の一つとして古くから解析されてきたが，実はこの藻類の光走性における眼点の役割についての議論は未だ結論が出ていない．本総説では微細藻類の眼点構造やEuglenophytaの光運動反応に関わる細胞構造について概説すると共に，Euglena gracilis の光走性における眼点の機能の解明に向けた筆者らの取り組みを簡単に紹介する．

Carotenoids participate in photosynthesis and photoprotection in oxygenic phototrophs. Euglena gracilis, a eukaryotic phytoflagellate, synthesizes several carotenoids: β-carotene, neoxanthin, diadinoxanthin and diatoxanthin. Temperature is one of the most striking external stimuli altering carotenoid production. In the present study, to elucidate the regulation of carotenoid synthesis of E. gracilis in response to environmental stimuli, we functionally identified phytoene desaturase genes (crtP1 and crtP2) and the ζ-carotene desaturase gene (crtQ) of this alga and analyzed expression of those genes and the composition of major carotenoids in cells grown under cold (20°C) and high-intensity light (HL; 240 µmol photon m⁻² s⁻¹) conditions. 20°C–HL treatment increased the transcriptional level of the phytoene synthase gene (crtB), and crtP1 and crtP2, whose products catalyze the early steps of carotenoid biosynthesis in this alga. Cultivation at 20°C under illumination at 55 µmol photon m⁻² s⁻¹ (low-intensity light; LL) decreased the cell concentration, Chl and total major carotenoid content by 61, 75 and 50%, respectively, relative to control (25°C–LL) cells. When grown at 20°C–HL, the cells showed a greater decrease in cell concentration and photosynthetic pigment contents than those in 20°C–LL. β-Carotene, neoxanthin and diadinoxanthin contents were decreased by more than half in 20°C–LL and 20°C–HL treatments. On the other hand, when subjected to 20°C–LL and 20°C–HL, the cells retained a diatoxanthin content comparable with control cells. Our findings suggested that diatoxanthin plays crucial roles in the acclimation to cold and intense light condition. To the best of our knowledge, this is the first report on a photosynthetic organism possessing dual crtP genes.

Photosynthetic organisms utilize carotenoids for photoprotection as well as light harvesting. Our previous study revealed that high-intensity light increases the expression of the gene for phytoene synthase (EgcrtB) in Euglena gracilis (a unicellular phytoflagellate), the encoded enzyme catalyzes the first committed step of the carotenoid biosynthesis pathway. To examine carotenoid synthesis of E. gracilis in response to light stress, we analyzed carotenoid species and content in cells grown under various light intensities. In addition, we investigated the effect of suppressing EgcrtB with RNA interference (RNAi) on growth and carotenoid content. After cultivation for 7 days under continuous light at 920 μmol m−2 s−1, β-carotene, diadinoxanthin (Ddx), and diatoxanthin (Dtx) content in cells was significantly increased compared with standard light intensity (55 μmol m−2 s−1). The high-intensity light (920 μmol m−2 s−1) increased the pool size of diadinoxanthin cycle pigments (i.e., Ddx + Dtx) by 1.2-fold and the Dtx/Ddx ratio from 0.05 (control) to 0.09. In contrast, the higher-intensity light treatment caused a 58% decrease in chlorophyll (a + b) content and diminished the number of thylakoid membranes in chloroplasts by approximately half compared with control cells, suggesting that the high-intensity light-induced accumulation of carotenoids is associated with an increase in both the number and size of lipid globules in chloroplasts and the cytoplasm. Transient suppression of EgcrtB in this alga by RNAi resulted in significant decreases in cell number, chlorophyll, and total major carotenoid content by 82, 82 and 86%, respectively, relative to non-electroporated cells. Furthermore, suppression of EgcrtB decreased the number of chloroplasts and thylakoid membranes and increased the Dtx/Ddx ratio by 1.6-fold under continuous illumination even at the standard light intensity, indicating that blocking carotenoid synthesis increased the susceptibility of cells to light stress. Our results indicate that suppression of EgcrtB causes a significant decrease in carotenoid and chlorophyll content in E. gracilis accompanied by changes in intracellular structures, suggesting that Dtx (de-epoxidized form of diadinoxanthin cycle pigments) contributes to photoprotection of this alga during the long-term acclimation to light-induced stress.