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A free-space optical interconnection scheme is described for massively parallel processors based on the interconnection-cached network architecture. The optical network operates in a circuit-switching mode. Combined with a packet-switching operation among the circuit-switched optical channels, a high-bandwidth, low-latency network for massively parallel processing results. The design and assembly of a 64-channel experimental prototype is discussed, and operational results are presented.

Abstract Background Euglenophytes are a group of photosynthetic flagellates possessing a plastid derived from a green algal endosymbiont, which was incorporated into an ancestral host cell via secondary endosymbiosis. However, the impact of endosymbiosis on the euglenophyte nuclear genome is not fully understood due to its complex nature as a 'hybrid' of a non-photosynthetic host cell and a secondary endosymbiont. Results We analyzed an EST dataset of the model euglenophyte Euglena gracilis using a gene mining program designed to detect laterally transferred genes. We found E. gracilis genes showing affinity not only with green algae, from which the secondary plastid in euglenophytes evolved, but also red algae and/or secondary algae containing red algal-derived plastids. Phylogenetic analyses of these 'red lineage' genes suggest that E. gracilis acquired at least 14 genes via eukaryote-to-eukaryote lateral gene transfer from algal sources other than the green algal endosymbiont that gave rise to its current plastid. We constructed an EST library of the aplastidic euglenid Peranema trichophorum, which is a eukaryovorous relative of euglenophytes, and also identified 'red lineage' genes in its genome. Conclusions Our data show genome mosaicism in E. gracilis and P. trichophorum. One possible explanation for the presence of these genes in these organisms is that some or all of them were independently acquired by lateral gene transfer and contributed to the successful integration and functioning of the green algal endosymbiont as a secondary plastid. Alternative hypotheses include the presence of a phagocytosed alga as the single source of those genes, or a cryptic tertiary endosymbiont harboring secondary plastid of red algal origin, which the eukaryovorous ancestor of euglenophytes had acquired prior to the secondary endosymbiosis of a green alga.

This study investigated the prognostic impact of vitamin D deficiency and reduced skeletal muscle mass in diffuse large B-cell lymphoma (DLBCL) patients. A retrospective analysis of 186 newly diagnosed DLBCL patients from 2012 to 2022 was conducted, measuring serum 25-hydroxyvitamin D [25(OH)D] levels and the skeletal muscle index (SMI). Decreased vitamin D levels were linked to more severe DLBCL disease, with a median 25(OH)D concentration of 13 (4.0-27) ng/mL. Males in the group with a low SMI had a considerably lower 25(OH)D concentration. The optimal threshold of 25(OH)D levels for overall survival (OS) was 9.6 ng/mL, with lower values associated with a higher likelihood of recurrence and mortality. Multivariable analysis showed hazard ratios for OS of 1.4 [95% CI 0.77-2.5] for a low SMI and 3.2 [95% CI 1.8-5.8] for low 25(OH)D concentration. The combination of a low SMI and low vitamin D concentration resulted in the worst prognosis. Thus, low levels of vitamin D associated with disease progression significantly impact DLBCL prognosis, which can be further stratified by the SMI, providing valuable insights for patient management and potential therapeutic interventions.

Legumes control root nodule symbiosis (RNS) in response to environmental nitrogen availability. Despite the recent understanding of the molecular basis of external nitrate-mediated control of RNS, it remains mostly elusive how plants regulate physiological processes depending on internal nitrogen status. In addition, iron (Fe) acts as an essential element that enables symbiotic nitrogen fixation; however, the mechanism of Fe accumulation in nodules is poorly understood. Here, we focus on the transcriptome in response to internal nitrogen status during RNS in Lotus japonicus and identify that IRON MAN (IMA) peptide genes are expressed during symbiotic nitrogen fixation. We show that LjIMA1 and LjIMA2 expressed in the shoot and root play systemic and local roles in concentrating internal Fe to the nodule. Furthermore, IMA peptides have conserved roles in regulating nitrogen homeostasis by adjusting nitrogen-Fe balance in L. japonicus and Arabidopsis thaliana. These findings indicate that IMA-mediated Fe provision plays an essential role in regulating nitrogen-related physiological processes., (© 2024. The Author(s).)

Precise detection of the transcriptional start site (TSS) is a key for characterizing transcriptional regulation of genes and for annotation of newly sequenced genomes. Here, we describe the development of an improved method, designated 'TSS-seq2.' This method is an iterative improvement of TSS-seq, a previously published enzymatic cap-structure conversion method to detect TSSs in base sequences. By modifying the original procedure, including by introducing split ligation at the key cap-selection step, the yield and the accuracy of the reaction has been substantially improved. For example, TSS-seq2 can be conducted using as little as 5 ng of total RNA with an overall accuracy of 96%; this yield a less-biased and more precise detection of TSS. We then applied TSS-seq2 for TSS analysis of four plant species that had not yet been analyzed by any previous TSS method., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Importance: Omics analyses suggested a mutually indispensable tripartite association among the host D. citri and organelle-like bacteriome associates, Carsonella and Profftella , which are vertically transmitted through host generations. This relationship is based on the metabolic complementarity among these organisms, which is partly enabled by horizontal gene transfer between partners. However, little was known about the fine morphology of the symbionts and the bacteriome, the interface among these organisms. As a first step to address this issue, the present study performed transmission electron microscopy, which revealed previously unrecognized ultrastructures, including aggregations of ribosomes in Carsonella , numerous tubes and occasional protrusions of Profftella , apparently degrading Profftella , and host organelles with different abundance and morphology in distinct cell types. These findings provide insights into the behaviors of the symbionts and host cells to maintain the symbiotic relationship in D. citri., Competing Interests: The authors declare no conflict of interest.

Kleptoplasts (kP) are distinct among photosynthetic organelles in eukaryotes (i.e., plastids) because they are routinely sequestered from prey algal cells and function only temporarily in the new host cell. Therefore, the hosts of kleptoplasts benefit from photosynthesis without constitutive photoendosymbiosis. Here, we report that the euglenozoan Rapaza viridis has only kleptoplasts derived from a specific strain of green alga, Tetraselmis sp., but no canonical plastids like those found in its sister group, the Euglenophyceae. R. viridis showed a dynamic change in the accumulation of cytosolic polysaccharides in response to light-dark cycles, and 13 C isotopic labeling of ambient bicarbonate demonstrated that these polysaccharides originate in situ via photosynthesis; these data indicate that the kleptoplasts of R. viridis are functionally active. We also identified 276 sequences encoding putative plastid-targeting proteins and 35 sequences of presumed kleptoplast transporters in the transcriptome of R. viridis . These genes originated in a wide range of algae other than Tetraselmis sp., the source of the kleptoplasts, suggesting a long history of repeated horizontal gene transfer events from different algal prey cells. Many of the kleptoplast proteins, as well as the protein-targeting system, in R. viridis were shared with members of the Euglenophyceae, providing evidence that the early evolutionary stages in the green alga-derived secondary plastids of euglenophytes also involved kleptoplasty.

During the course of plant evolution, leguminous and a few plants species have established root nodule symbiosis (RNS), one of the nitrogen nutrient acquisition strategies based on mutual interaction between plants and nitrogen-fixing bacteria. In addition to its useful agronomic trait, RNS comprises a unique form of plant lateral organogenesis; dedifferentiation and activation of cortical cells in the root are induced upon bacterial infection during nodule development. In the past few years, the elucidations of the significance of NODULE INCEPTION transcription factor as a potentially key innovative factor of RNS, the details of its function, and the successive discoveries of its target genes have advanced our understanding underlying molecular mechanisms of nodule organogenesis. In addition, a recent elucidation of the role of legume SHORTROOT-SCARECROW module has provided the insights into the unique properties of legume cortical cells. Here, I summarize such latest findings on the neofunctionalized key players of nodule organogenesis, which may provide clue to understand an evolutionary basis of RNS., (Copyright © 2023 by JAPANESE SOCIETY OF BREEDING.)

Root nodule symbiosis is promoted in nitrogen-deficient environments, whereas host plants cease the symbiosis if they can obtain enough nitrogen from their surrounding soil. In Lotus japonicus, recent reports indicate that two NODULE INCEPTION (NIN)-LIKE PROTEIN (NLP) transcription factors, LjNLP1 and LjNLP4, play important roles in the regulation of gene expression and nodulation in response to nitrate. To characterize the redundant and unique roles of LjNLP1 and LjNLP4 in more detail, we reanalyzed our previous transcriptome data using Ljnlp1 and Ljnlp4 mutants. Although downstream genes of LjNLP1 and LjNLP4 mostly overlapped, we found that nitrate-induced expression of NITRATE TRANSPORTER 2 (LjNRT2) family genes was specifically regulated by LjNLP1. In contrast, LjNRT1 gene family expression was regulated by both LjNLP1 and LjNLP4. Therefore, it is likely that the two NLPs play distinct roles in the regulation of nitrate transport.

Drought stress is a severe environmental issue that threatens agriculture at a large scale. PHYTOCHROMES (PHYs) are important photoreceptors in plants that control plant growth and development and are involved in plant stress response. The aim of this study was to identify the role of PHYs in the tomato cv. 'Moneymaker' under drought conditions. The tomato genome contains five PHYs , among which mutant lines in tomato PHYA and PHYB ( B1 and B2 ) were used. Compared to the WT, phyA and phyB1B2 mutants exhibited drought tolerance and showed inhibition of electrolyte leakage and malondialdehyde accumulation, indicating decreased membrane damage in the leaves. Both phy mutants also inhibited oxidative damage by enhancing the expression of reactive oxygen species (ROS) scavenger genes, inhibiting hydrogen peroxide (H 2 O 2 ) accumulation, and enhancing the percentage of antioxidant activities via DPPH test. Moreover, expression levels of several aquaporins were significantly higher in phyA and phyB1B2 , and the relative water content (RWC) in leaves was higher than the RWC in the WT under drought stress, suggesting the enhancement of hydration status in the phy mutants. Therefore, inhibition of oxidative damage in phyA and phyB1B2 mutants may mitigate the harmful effects of drought by preventing membrane damage and conserving the plant hydrostatus.

Legumes have adaptive mechanisms that regulate nodulation in response to the amount of nitrogen in the soil. In Lotus japonicus, two NODULE INCEPTION (NIN)-LIKE PROTEIN (NLP) transcription factors, LjNLP4 and LjNLP1, play pivotal roles in the negative regulation of nodulation by controlling the expression of symbiotic genes in high nitrate conditions. Despite an improved understanding of the molecular basis for regulating nodulation, how nitrate plays a role in the signaling pathway to negatively regulate this process is largely unknown. Here, we show that nitrate transport via NITRATE TRANSPORTER 2.1 (LjNRT2.1) is a key step in the NLP signaling pathway to control nodulation. A mutation in the LjNRT2.1 gene attenuates the nitrate-induced control of nodulation. LjNLP1 is necessary and sufficient to induce LjNRT2.1 expression, thereby regulating nitrate uptake/transport. Our data suggest that LjNRT2.1-mediated nitrate uptake/transport is required for LjNLP4 nuclear localization and induction/repression of symbiotic genes. We further show that LjNIN, a positive regulator of nodulation, counteracts the LjNLP1-dependent induction of LjNRT2.1 expression, which is linked to a reduction in nitrate uptake. These findings suggest a plant strategy in which nitrogen acquisition switches from obtaining nitrogen from the soil to symbiotic nitrogen fixation., (© The Author(s) 2022. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Ochrophyta is an algal group belonging to the Stramenopiles and comprises diverse lineages of algae which contribute significantly to the oceanic ecosystems as primary producers. However, early evolution of the plastid organelle in Ochrophyta is not fully understood. In this study, we provide a well-supported tree of the Stramenopiles inferred by the large-scale phylogenomic analysis that unveils the eukaryvorous (nonphotosynthetic) protist Actinophrys sol (Actinophryidae) is closely related to Ochrophyta. We used genomic and transcriptomic data generated from A. sol to detect molecular traits of its plastid and we found no evidence of plastid genome and plastid-mediated biosynthesis, consistent with previous ultrastructural studies that did not identify any plastids in Actinophryidae. Moreover, our phylogenetic analyses of particular biosynthetic pathways provide no evidence of a current and past plastid in A. sol. However, we found more than a dozen organellar aminoacyl-tRNA synthases (aaRSs) that are of algal origin. Close relationships between aaRS from A. sol and their ochrophyte homologs document gene transfer of algal genes that happened before the divergence of Actinophryidae and Ochrophyta lineages. We further showed experimentally that organellar aaRSs of A. sol are targeted exclusively to mitochondria, although organellar aaRSs in Ochrophyta are dually targeted to mitochondria and plastids. Together, our findings suggested that the last common ancestor of Actinophryidae and Ochrophyta had not yet completed the establishment of host-plastid partnership as seen in the current Ochrophyta species, but acquired at least certain nuclear-encoded genes for the plastid functions., (© The Author(s) 2022. Published by Oxford University Press on behalf of Society for Molecular Biology and Evolution.)

Heat stress (HS) is a prevalent negative factor affecting plant growth and development, as it is predominant worldwide and threatens agriculture on a large scale. PHYTOCHROMES (PHYs) are photoreceptors that control plant growth and development, and the stress signaling response partially interferes with their activity. PHYA, B1, and B2 are the most well-known PHY types in tomatoes. Our study aimed to identify the role of tomato 'Money Maker' phyA and phyB1B2 mutants in stable and fluctuating high temperatures at different growth stages. In the seed germination and vegetative growth stages, the phy mutants were HS tolerant, while during the flowering stage the phy mutants revealed two opposing roles depending on the HS exposure period. The response of the phy mutants to HS during the fruiting stage showed similarity to WT. The most obvious stage that demonstrated phy mutants' tolerance was the vegetative growth stage, in which a high degree of membrane stability and enhanced water preservation were achieved by the regulation of stomatal closure. In addition, both mutants upregulated the expression of heat-responsive genes related to heat tolerance. In addition to lower malondialdehyde accumulation, the phyA mutant enhanced proline levels. These results clarified the response of tomato phyA and phyB1B2 mutants to HS.

Leguminous plants produce nodules for nitrogen fixation; however, nodule production incurs an energy cost. Therefore, as an adaptive strategy, leguminous plants halt root nodule development when sufficient amounts of nitrogen nutrients, such as nitrate, are present in the environment. Although legume NODULE INCEPTION (NIN)-LIKE PROTEIN (NLP) transcription factors have recently been identified, understanding how nodulation is controlled by nitrate, a fundamental question for nitrate-mediated transcriptional regulation of symbiotic genes, remains elusive. Here, we show that two Lotus japonicus NLPs, NITRATE UNRESPONSIVE SYMBIOSIS 1 (NRSYM1)/LjNLP4 and NRSYM2/LjNLP1, have overlapping functions in the nitrate-induced control of nodulation and act as master regulators for nitrate-dependent gene expression. We further identify candidate target genes of LjNLP4 by combining transcriptome analysis with a DNA affinity purification-seq approach. We then demonstrate that LjNLP4 and LjNIN, a key nodulation-specific regulator and paralog of LjNLP4, have different DNA-binding specificities. Moreover, LjNLP4-LjNIN dimerization underlies LjNLP4-mediated bifunctional transcriptional regulation. These data provide a basic principle for how nitrate controls nodulation through positive and negative regulation of symbiotic genes., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

The genus Stentor is a relatively well-known ciliate owing to its lucid trumpet shape. Stentor pyriformis represents a green, short, and fat Stentor, but it is a little-known species. We investigated 124 ponds and wetlands in Japan and confirmed the presence of S. pyriformis at 23 locations. All these ponds were noticeably oligotrophic. With the improvement of oligotrophic culture conditions, we succeeded in long-term cultivation of three strains of S. pyriformis. The cytoplasm of S. piriformis contains a large number of 1-3 μm refractive granules that turn brown by Lugol's staining. The granules also show a typical Maltese-cross pattern by polarization microscopy, strongly suggesting that the granules are made of amylopectin-rich starch. By analyzing the algal rDNA, it was found that all S. pyriformis symbionts investigated in this study were Chlorella variabilis. This species is known as the symbiont of Paramecium bursaria and is physiologically specialized for endosymbiosis. Genetic discrepancies between C. variabilis of S. pyriformis and P. bursaria may indicate that algal sharing was an old incident. Having symbiotic algae and storing carbohydrate granules in the cytoplasm is considered a powerful strategy for this ciliate to withstand oligotrophic and cold winter environments in highland bogs.

Legumes utilize a shoot-mediated signaling system to maintain a mutualistic relationship with nitrogen-fixing bacteria in root nodules. In Lotus japonicus, shoot-to-root transfer of microRNA miR2111 that targets TOO MUCH LOVE, a nodulation suppressor in roots, has been proposed to explain the mechanism underlying nodulation control from shoots. However, the role of shoot-accumulating miR2111s for the systemic regulation of nodulation was not clearly shown. Here, we find L. japonicus has seven miR2111 loci, including those mapped through RNA-seq. MIR2111-5 expression in leaves is the highest among miR2111 loci and repressed after rhizobial infection depending on a shoot-acting HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) receptor. MIR2111-5 knockout mutants show significantly decreased nodule numbers and miR2111 levels. Furthermore, grafting experiments using transformants demonstrate scions with altered miR2111 levels influence nodule numbers in rootstocks in a dose-dependent manner. Therefore, miR2111 accumulation in leaves through MIR2111-5 expression is required for HAR1-dependent systemic optimization of nodule number.

Legumes possess the autoregulation of nodulation (AON) pathway which is responsible for maintaining optimal root nodule number. In Lotus japonicus , AON comprises the CLE-HAR1-TML module, which plays an essential role in transmitting signals via root-to-shoot-to-root long-distance signaling. In addition to AON's principal role of negatively regulating nodule number, a recent study revealed another in the systemic control of rhizobial infection. Nitrate also negatively regulates the pleiotropic phases of legume- Rhizobium symbioses, including rhizobial infection and nodule number. Nitrate signaling has recently been shown to use AON components such as CLE-RS2 and HAR1 to control nodule number. Here we consider the role of a loss-of-function mutation in CLE-RS1, -RS2 and TML in rhizobial infection in relation to nitrate. Our results agree with previous findings and support the hypothesis that AON is required for the control of rhizobial infection but not for its nitrate-induced control. Furthermore, we confirm that the tml mutants exhibit nitrate sensitivity that differs from that of cle-rs2 and har1 . Hence, while the nitrate-induced control mechanism of nodule number uses AON components, an unknown pathway specific to nitrate may exist downstream of HAR1, acting in parallel with the HAR1> TML pathway.

Legume-rhizobium symbiosis leads to the formation of nitrogen-fixing root nodules. However, externally applied chemical nitrogen fertilizers (nitrate and ammonia) strongly inhibit nodule formation and nitrogen fixation. Here, we isolated several rhizobial strains exhibiting a superior nodulation and nitrogen fixation with soybean at high nitrate concentrations. The nodulation of soybean symbiont Bradyrhizobium diazoefficiens USDA110 was significantly inhibited at 12.5 mM nitrate; however, three isolates (NKS4, NKM2 and NKTG2) were capable of forming nitrogen-fixing nodules, even at 20 mM nitrate. These isolates exhibited higher nodulation competitiveness and induced larger nodules with higher nitrogen-fixation activity than USDA110 at 5 mM nitrate. Furthermore, these isolates induced more nodules than USDA110 even in nitrate-free conditions. These isolates had a distant lineage within the Bradyrhizobium genus; though they were relatively phylogenetically close to Bradyrhizobium japonicum, their morphological and growth characteristics were significantly different. Notably, in the presence of nitrate, expression of the soybean symbiosis-related genes (GmENOD40 and GmNIN) was significantly higher and expression of GmNIC1 that is involved in nitrate-dependent nodulation inhibition was lower in the roots inoculated with these isolates in contrast with inoculation of USDA110. These novel rhizobia serve as promising inoculants for soybeans cultivated in diverse agroecosystems, particularly on nitrate-applied soils., (© FEMS 2019.)

Legumes survive in nitrogen-limited soil by forming a symbiosis with rhizobial bacteria. During root nodule symbiosis, legumes strictly control the development of their symbiotic organs, the nodules, in a process known as autoregulation of nodulation (AON). The study of hypernodulation mutants has elucidated the molecular basis of AON. Some hypernodulation mutants show an increase in rhizobial infection in addition to developmental alteration. However, the relationship between the AON and the regulation of rhizobial infection has not been clarified. We previously isolated daphne , a nodule inception ( nin ) allelic mutant, in Lotus japonicus . This mutant displayed dramatically increased rhizobial infection, suggesting the existence of NIN-mediated negative regulation of rhizobial infection. Here, we investigated whether the previously isolated components of AON, especially CLAVATA3/ESR (CLE)-RELATED-ROOT SIGNAL1 (CLE-RS1), CLE-RS2, and their putative receptor HYPERNODULATION AND ABERRANT ROOT FORMATION1 (HAR1), were able to suppress increased infection in the daphne mutant. The constitutive expression of LjCLE-RS1/2 strongly reduced the infection in the daphne mutant in a HAR1-dependent manner. Moreover, reciprocal grafting analysis showed that strong reduction of infection in daphne rootstock constitutively expressing LjCLE-RS1 was canceled by a scion of the har1 or klavier mutant, the genes responsible for encoding putative LjCLE-RS1 receptors. These data indicate that rhizobial infection is also systemically regulated by CLE-HAR1 signaling, a component of AON. In addition, the constitutive expression of NIN in daphne har1 double-mutant roots only partially reduced the rhizobial infection. Our findings indicate that the previously identified NIN-mediated negative regulation of infection involves unknown local signaling, as well as CLE-HAR1 long-distance signaling.

Freshwater protists often harbor unicellular green algae within their cells. In ciliates, possibly because of large host cell sizes and the small size of algal coccoids, a single host cell typically contains more than a hundred algal cells. While surveying such algae-bearing protists on Minami Daito Jima Island in Japan, we found a green Loxodes ciliate (Loxodida, Karyorelictea) that contained one or two dozens of very large coccoid algae. We isolated one of these algae and analyzed its characteristics in detail. A small subunit (SSU) rDNA phylogeny indicated Pseudodidymocystis species (Scenedesmaceae, Chlorophyceae) to be the taxon closest to the alga, although it was clearly separated from this by 39 or more different sites (inclusive of gaps). SSU rRNA structure analyses indicated that these displacements included eight compensatory base changes (CBCs) and seven hemi-CBCs. We therefore concluded that this alga belongs to a separate genus, and described it as Pediludiella daitoensis gen. et sp. nov. The shape of the isolated and cultured P. daitoensis was nearly spherical and reached up to 30 µm in diameter. Chloroplasts were arranged peripherally and often split and elongated. Cells were often vacuolated and possessed a net-like cytoplasm that resembled a football (soccer ball) in appearance, which was reflected in the genus name.

Arabidopsis SIZ1 encodes a SUMO E3 ligase to regulate abiotic and biotic stress responses. Among SIZ1 or mammalian PIAS orthologs, plant SIZ1 proteins contain the plant homeodomain (PHD) finger, a C 4 HC 3 zinc finger. Here, we investigated the importance of PHD of Arabidopsis SIZ1. The Pro SIZ1 ::SIZ1(ΔPHD):GFP was unable to complement growth retardation, ABA hypersensitivity, and the cold-sensitive phenotype of the siz1 mutant, but Pro SIZ1 ::SIZ1:GFP could. Substitution of C162S in the PHD finger was unable to complement the siz1 mutation. Tri-methylated histone H3K4 (H3K4me3) was recognized by PHD, not by PHD(C162S). WRKY70 was up-regulated in the siz1-2 mutant and H3K4me3 accumulated at high levels in the WRKY70 promoter. PHD interacts with ATX, which mediates methylation of histone, probably leading to suppression of ATX's function. These results suggest that the PHD finger of SIZ1 is important for recognition of the histone code and is required for SIZ1 function and transcriptional suppression.

Extant eukaryote ecology is primarily sustained by oxygenic photosynthesis, in which chlorophylls play essential roles. The exceptional photosensitivity of chlorophylls allows them to harvest solar energy for photosynthesis, but on the other hand, they also generate cytotoxic reactive oxygen species. A risk of such phototoxicity of the chlorophyll must become particularly prominent upon dynamic cellular interactions that potentially disrupt the mechanisms that are designed to quench photoexcited chlorophylls in the phototrophic cells. Extensive examination of a wide variety of phagotrophic, parasitic, and phototrophic microeukaryotes demonstrates that a catabolic process that converts chlorophylls into nonphotosensitive 13 2 ,17 3 -cyclopheophorbide enols (CPEs) is phylogenetically ubiquitous among extant eukaryotes. The accumulation of CPEs is identified in phagotrophic algivores belonging to virtually all major eukaryotic assemblages with the exception of Archaeplastida, in which no algivorous species have been reported. In addition, accumulation of CPEs is revealed to be common among phototrophic microeukaryotes (i.e., microalgae) along with dismantling of their secondary chloroplasts. Thus, we infer that CPE-accumulating chlorophyll catabolism (CACC) primarily evolved among algivorous microeukaryotes to detoxify chlorophylls in an early stage of their evolution. Subsequently, it also underpinned photosynthetic endosymbiosis by securing close interactions with photosynthetic machinery containing abundant chlorophylls, which led to the acquisition of secondary chloroplasts. Our results strongly suggest that CACC, which allowed the consumption of oxygenic primary producers, ultimately permitted the successful radiation of the eukaryotes throughout and after the late Proterozoic global oxygenation.

[This corrects the article DOI: 10.1371/journal.pgen.1007865.].

Legumes can survive in nitrogen-deficient environments by forming root-nodule symbioses with rhizobial bacteria; however, forming nodules consumes energy, and nodule numbers must thus be strictly controlled. Previous studies identified major negative regulators of nodulation in Lotus japonicus, including the small peptides CLAVATA3/ESR (CLE)-RELATED-ROOT SIGNAL1 (CLE-RS1), CLE-RS2, and CLE-RS3, and their putative major receptor HYPERNODULATION AND ABERRANT ROOT FORMATION1 (HAR1). CLE-RS2 is known to be expressed in rhizobia-inoculated roots, and is predicted to be post-translationally arabinosylated, a modification essential for its activity. Moreover, all three CLE-RSs suppress nodulation in a HAR1-dependent manner. Here, we identified PLENTY as a gene responsible for the previously isolated hypernodulation mutant plenty. PLENTY encoded a hydroxyproline O-arabinosyltransferase orthologous to ROOT DETERMINED NODULATION1 in Medicago truncatula. PLENTY was localized to the Golgi, and an in vitro analysis of the recombinant protein demonstrated its arabinosylation activity, indicating that CLE-RS1/2/3 may be substrates for PLENTY. The constitutive expression experiments showed that CLE-RS3 was the major candidate substrate for PLENTY, suggesting the substrate preference of PLENTY for individual CLE-RS peptides. Furthermore, a genetic analysis of the plenty har1 double mutant indicated the existence of another PLENTY-dependent and HAR1-independent pathway negatively regulating nodulation.

Nitrogen-fixing rhizobia and arbuscular mycorrhizal fungi (AMF) form symbioses with plant roots and these are established by precise regulation of symbiont accommodation within host plant cells. In model legumes such as Lotus japonicus and Medicago truncatula, rhizobia enter into roots through an intracellular invasion system that depends on the formation of a root-hair infection thread (IT). While IT-mediated intracellular rhizobia invasion is thought to be the most evolutionarily derived invasion system, some studies have indicated that a basal intercellular invasion system can replace it when some nodulation-related factors are genetically modified. In addition, intracellular rhizobia accommodation is suggested to have a similar mechanism as AMF accommodation. Nevertheless, our understanding of the underlying genetic mechanisms is incomplete. Here we identify a L. japonicus nodulation-deficient mutant, with a mutation in the LACK OF SYMBIONT ACCOMMODATION (LAN) gene, in which root-hair IT formation is strongly reduced, but intercellular rhizobial invasion eventually results in functional nodule formation. LjLAN encodes a protein that is homologous to Arabidopsis MEDIATOR 2/29/32 possibly acting as a subunit of a Mediator complex, a multiprotein complex required for gene transcription. We also show that LjLAN acts in parallel with a signaling pathway including LjCYCLOPS. In addition, the lan mutation drastically reduces the colonization levels of AMF. Taken together, our data provide a new factor that has a common role in symbiont accommodation process during root nodule and AM symbiosis., Competing Interests: The authors have declared that no competing interests exist.

Transient protein expression is an effective tool to rapidly unravel novel gene functions, such as transcriptional activity of promoters and sub-cellular localization of proteins. However, transient expression is not applicable to some species and varieties because of insufficient expression levels. We recently developed one of the strongest agroinfiltration-based transient protein expression systems for plant cells, termed 'Tsukuba system.' About 4 mg/g fresh weight of protein expression in Nicotiana benthamiana was obtained using this system. The vector pBYR2HS, which contains a geminiviral replication system and a double terminator, can be used in various plant species and varieties, including lettuces, eggplants, tomatoes, hot peppers, and orchids. In this study, we assessed the applicability of the Tsukuba system to several species of legumes, including Lotus japonicus , soybean Glycine max , and common bean Phaseolus vulgaris . The GFP protein was transiently expressed in the seedpods of all examined legume species, however, protein expression in leaves was observed only in P. vulgaris . Taken together, our system is an effective tool to examine gene function rapidly in several legume species based on transient protein expression., (© 2019 The Japanese Society for Plant Cell and Molecular Biology.)

Regulation of intracellular pH is critically important for many cellular functions. The quantification of proton extrusion in different types of cells and physiological conditions is pivotal to fully elucidate the mechanisms of pH homeostasis. Here we show the use of gold nanoparticles (AuNP) to create a high spatial resolution sensor for measuring extracellular pH in proximity of the cell membrane. We test the sensor on HepG2 liver cancer cells and MKN28 gastric cancer cells before and after inhibition of Na + /H + exchanger. The gold surface conjugation strategy is conceived with a twofold purpose: i) to anchor the AuNP to the membrane proteins and ii) to quantify the local pH from AuNP using surface enhanced Raman spectroscopy (SERS). The nanometer size of the cell membrane anchored sensor and the use of SERS enable us to visualize highly localized variation of pH induced by H + extrusion, which is particularly upregulated in cancer cells.

Ants are known to use a colony-specific blend of cuticular hydrocarbons (CHCs) as a pheromone to discriminate between nestmates and non-nestmates and the CHCs were sensed in the basiconic type of antennal sensilla ( S. basiconica ). To investigate the functional design of this type of antennal sensilla, we observed the ultra-structures at 2D and 3D in the Japanese carpenter ant, Camponotus japonicus , using a serial block-face scanning electron microscope (SBF-SEM), and conventional and high-voltage transmission electron microscopes. Based on the serial images of 352 cross sections of SBF-SEM, we reconstructed a 3D model of the sensillum revealing that each S. basiconica houses > 100 unbranched dendritic processes, which extend from the same number of olfactory receptor neurons (ORNs). The dendritic processes had characteristic beaded-structures and formed a twisted bundle within the sensillum. At the "beads," the cell membranes of the processes were closely adjacent in the interdigitated profiles, suggesting functional interactions via gap junctions (GJs). Immunohistochemistry with anti-innexin (invertebrate GJ protein) antisera revealed positive labeling in the antennae of C. japonicus . Innexin 3, one of the five antennal innexin subtypes, was detected as a dotted signal within the S. basiconica as a sensory organ for nestmate recognition. These morphological results suggest that ORNs form an electrical network via GJs between dendritic processes. We were unable to functionally certify the electric connections in an olfactory sensory unit comprising such multiple ORNs; however, with the aid of simulation of a mathematical model, we examined the putative function of this novel chemosensory information network, which possibly contributes to the distinct discrimination of colony-specific blends of CHCs or other odor detection.

Root nodule symbiosis is one of the best characterized mutualistic relationships of plant-microbe symbiosis, where mainly leguminous species can obtain nitrogen sources fixed by nitrogen-fixing rhizobia through the formation of symbiotic organ root nodules. In order to drive this symbiotic process, plants need to provide carbon sources that should be used for their growth. Therefore, a balance between the benefits of obtaining nitrogen sources and the costs of losing carbon sources needs to be maintained during root nodule symbiosis. Plants have developed at least two negative regulatory systems of root nodule symbiosis. One strategy involves the regulation of nodule number in response to rhizobial infection. For this regulation, a systemic long-range signaling between roots and shoots called autoregulation of nodulation has a pivotal role. Another strategy involves the regulation of root nodule symbiosis in response to nitrate, the most abundant form of nitrogen nutrients in the soil. Recent studies indicate that long-distance signaling is shared between the two strategies, where NIN and NRSYM1, two paralogous RWP-RK transcription factors, can activate the production of nodulation-related CLE peptides in response to different inputs. Here, we provide an overview of such progress in our understanding of molecular mechanisms relevant to the control of the symbiotic balance, including their biological significance., (� The Author(s) 2018. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The expression of hundreds of genes is induced by low temperatures via a cold signaling pathway. ICE1, a MYC-type transcription factor, plays an important role in the induction of CBF3/DREB1A to control cold-responsive genes and cold tolerance. To elucidate other molecular factors, a yeast 2-hybrid screening was performed. Two MYC-type transcription factors, MYC67 and MYC70, were identified as ICE1-interacting proteins. The myc mutants were more tolerant to freezing temperatures than wild type. CBF3/DREB1A and other cold-responsive genes were up-regulated in the myc mutants. Overexpression of the MYC genes increased the cold sensitivity and down-regulated the expression of cold-responsive genes. The MYC proteins interacted with the cis-elements in the CBF3/DREB1A promoter, probably to interfere interaction between ICE1 and the cis-elements. Taken together, these results demonstrate that MYC67 and MYC70, ICE1 interactors, negatively regulate cold-responsive genes and cold tolerance.

Legumes and rhizobia establish symbiosis in root nodules. To balance the gains and costs associated with the symbiosis, plants have developed two strategies for adapting to nitrogen availability in the soil: plants can regulate nodule number and/or stop the development or function of nodules. Although the former is accounted for by autoregulation of nodulation, a form of systemic long-range signaling, the latter strategy remains largely enigmatic. Here, we show that the Lotus japonicus NITRATE UNRESPONSIVE SYMBIOSIS 1 (NRSYM1) gene encoding a NIN-LIKE PROTEIN transcription factor acts as a key regulator in the nitrate-induced pleiotropic control of root nodule symbiosis. NRSYM1 accumulates in the nucleus in response to nitrate and directly regulates the production of CLE-RS2, a root-derived mobile peptide that acts as a negative regulator of nodule number. Our data provide the genetic basis for how plants respond to the nitrogen environment and control symbiosis to achieve proper plant growth.

Cold shock triggers an immediate rise in the cytosolic free calcium concentration ([Ca 2+ ] cyt ) in Arabidopsis thaliana and this cold-induced elevation of [Ca 2+ ] cyt is inhibited by lanthanum or EGTA. It is suggested that intracellular calcium mainly contributes to the cold-induced [Ca 2+ ] cyt response by entering into the cytosol. Two calcium-permeable mechanosensitive channels, MCA1 and MCA2 (mid1-complementing activity), have been identified in Arabidopsis. Here, we demonstrate that MCA1 and MCA2 are involved in a cold-induced increase in [Ca 2+ ] cyt . The cold-induced [Ca 2+ ] cyt increase in mca1 and mca2 mutants was markedly lower than that in wild types. The mca1 mca2 double mutant exhibited chilling and freezing sensitivity, compared to wild-type plants. Expression of At5g61820, At3g51660, and At4g15490, which are not regulated by the CBF/DREB1s transcription factor, was down-regulated in mca1 mca2. These results suggest that MCA1 and MCA2 are involved in the cold-induced elevation of [Ca 2+ ] cyt , cold tolerance, and CBF/DREB1-independent cold signaling.