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The revision of the sub-order Microchiroptera is one of the most intriguing outcomes in recent mammalian molecular phylogeny. The unexpected sister–taxon relationship between rhinolophoid microbats and megabats, with the exclusion of other microbats, suggests that megabats arose in a relatively short period of time from a microbat-like ancestor. In order to understand the genetic mechanism underlying adaptive evolution in megabats, we determined the whole-genome sequences of two rousette megabats, Leschenault’s rousette (Rousettus leschenaultia) and the Egyptian fruit bat (R. aegyptiacus). The sequences were compared with those of 22 other mammals, including nine bats, available in the database. We identified that megabat genomes are distinct in that they have extremely low activity of SINE retrotranspositions, expansion of two chemosensory gene families, including the trace amine receptor (TAAR) and olfactory receptor (OR), and elevation of the dN/dS ratio in genes for immunity and protein catabolism. The adaptive signatures discovered in the genomes of megabats may provide crucial insight into their distinct evolution, including key processes such as virus resistance, loss of echolocation, and frugivorous feeding.

As a new method to determine the resonance frequency, Rabi-oscillation spectroscopy has been developed. In contrast to the conventional spectroscopy which draws the resonance curve, Rabi-oscillation spectroscopy fits the time evolution of the Rabi oscillation. By selecting the optimized frequency, it is shown that the precision is twice as good as the conventional spectroscopy with a frequency sweep. Furthermore, the data under different conditions can be treated in a unified manner, allowing more efficient measurements for systems consisting of a limited number of short-lived particles produced by accelerators such as muons. We have developed a fitting function that takes into account the spatial distribution of muonium and the spatial distribution of the microwave intensity to apply the new method to ground-state muonium hyperfine structure measurements at zero field. This was applied to the actual measurement data and the resonance frequencies were determined under various conditions. The result of our analysis gives $\nu_{\rm HFS}=4\ 463\ 301.61 \pm 0.71\ {\rm kHz}$, which is the world's highest precision under zero field conditions., Comment: 6 pages, 5 figures

We recently achieved targeted disruptions of cytoplasmic male sterility (CMS)-associated genes in the mitochondrial genomes of rice and rapeseed by using mitochondria-targeted transcription activator-like effector nucleases (mitoTALENs). It was the first report of stable and heritable targeted gene modification of plant mitochondrial genomes. Here, we attempted to use mitoTALENs to disrupt two mitochondrial genes in the model plant Arabidopsis thaliana(Arabidopsis) using three different promoters and two types of TALENs. The targets were the two isoforms of the ATP synthase subunit 6 gene, atp6-1 and atp6-2. Each of these genes was successfully deleted and the mitochondrial genomes were recovered in a homoplasmic state. The nuclear genome also has a copy of atp6-1, and we were able to confirm that it was the mitochondrial gene and not the nuclear pseudogene that was knocked out. Among the three mitoTALEN promoters tried, the RPS5A promoter was the most effective. Conventional mitoTALENs were more effective than single-molecule mito-compactTALENs. Targeted mitochondrial gene deletion was achieved by crossing as well as by floral-dip transformation to introduce the mitoTALEN constructs into the nucleus. The gene disruptions were caused by large (kb-size) deletions. The ends of the remaining sequences were connected to distant loci, mostly by illegitimate homologous recombinations between repeats.

Restriction–modification (RM) systems are typically regarded as “primitive immune systems” in bacteria. The roles of methylation in gene regulation, segregation, and mismatch repair are increasingly recognized. To analyze methyltransferase (MTase) diversity in Streptococcus pyogenes, we compared the RM system distribution in eight new complete genome sequences obtained here and in the database-deposited complete genome sequences of 51 strains. The MTase gene distribution showed that type I MTases often change DNA sequence specificity via switching target recognition domains between strains. The type II MTases in the included strains fell into two groups: a prophage-dominant one and a CRISPR-dominant one. Some highly variable type II MTases were found in the prophage region, suggesting that MTases acquired from phage DNA can generate methylome diversity. Additionally, to investigate the possible contribution of DNA methylation to phenotype, we compared the methylomes and transcriptomes from the four most closely related strains, the results of which suggest that phage-derived methylases possibly regulate the methylome, and, hence, regulate expression levels in S. pyogenes. Our findings will benefit further experimental work on the relationship between virulence genes and pathogenicity in S. pyogenes.

Background Periodontopathic bacteria such as Porphyromonas gingivalis produce several metabolites, including lipopolysaccharide (LPS) and n-butyric acid (BA). Past work suggested that periodontal infection may cause cognitive impairment in mice. Aims To elucidate the mechanisms by which metabolites such as LPS and BA, resulting from Porphyromonas gingivalis activity, induce immunological and physiological abnormalities in mice. Methods In the present work, 28 male ICR mice were placed in an open-field arena and the total distance (cm/600 s) they covered was recorded. Based on their moving distances, mice were divided into 4 groups (n = 7) and injected the following substances into their gingival tissues for 32 consecutive days: saline (C), 5 mmol/L of BA (B), 1 μg/mouse of LPS (L), and BA-LPS (BL) solutions. Distances covered by mice were also measured on days 14 and 21, with their habituation scores considered as “(moving distance on day 14 or 21)/(moving distance on day 0)”. Afterwards, mice were dissected, and hippocampal gene expression and the concentrations of short-chain fatty acids, neurotransmitters and cytokines in their blood plasma and brains were analyzed. In addition, mouse brain and liver tissues were fixed and visually assessed for histopathological abnormalities. Results Group BL had significantly higher habituation scores than C and B on day 14. LPS induced higher habituation scores on day 21. LPS induced significant decreases in the mRNA levels of interleukin (IL)-6 and brain-derived neurotrophic factors, and an increase in neurotrophic tyrosine kinase receptor type 2. In both plasma and brain, LPS induced a significant acetate increase. Moreover, LPS significantly increased acetylcholine in brain. In plasma alone, LPS and BA significantly decreased monocyte chemoattractant protein 1 (MCP-1). However, while LPS significantly decreased tyrosine, BA significantly increased it. Lastly, LPS significantly decreased IL-6 and tumor necrosis factor in plasma. No histopathological abnormalities were detected in liver or brain tissues of mice. Conclusion We showed that injections of LPS and/or BA induced mice to move seemingly tireless and that both LPS and BA injections strongly induced a reduction of MCP-1 in blood plasma. We concluded that LPS and BA may have been crucial to induce and/or aggravate abnormal behavior in mice.

Some sea slugs sequester chloroplasts from algal food in their intestinal cells and photosynthesize for months. This phenomenon, kleptoplasty, poses a question of how the chloroplast retains its activity without the algal nucleus. There have been debates on the horizontal transfer of algal genes to the animal nucleus. To settle the arguments, this study reported the genome of a kleptoplastic sea slugPlakobranchus ocellatusand found no evidence of photosynthetic genes encoded on the nucleus. Nevertheless, it was confirmed that light illumination prolongs the life of mollusk under starvation. These data presented a paradigm that a complex adaptive trait, as typified by photosynthesis, can be transferred between eukaryotic kingdoms by a unique organelle transmission without nuclear gene transfer. Our phylogenomic analysis showed that genes for proteolysis and immunity undergo gene expansion and are up-regulated in chloroplast-enriched tissue, suggesting that these molluskan genes are involved in the DNA-independent phenotype acquisition.

Kojima, M.; Otabi, H.; Kumanogoh, H.; Toyoda, A.; Ikawa, M.; Okabe, M.; Mizui, T. Reduction in BDNF from Inefficient Precursor Conversion Influences Nest Building and Promotes Depressive-Like Behavior in Mice. Int. J. Mol. Sci. 2020, 21, 3984. https://doi.org/10.3390/ijms21113984, We generated a knock-in mouse line in which the gene encoding brain-derived neurotrophic factor (Bdnf) was replaced with a sequence for proBDNF containing human single nucleotide polymorphisms encoding arginines proximal to the cleavage site (R125M and R127L). The ratio of the mature form of BDNF (mBDNF) to precursor BDNF (proBDNF) in hippocampal tissue lysates was decreased in a manner dependent on the number of copies of the mutant gene, indicating that the mutations inhibited proteolytic conversion of proBDNF into mBDNF. Although homozygous mice had a proBDNF/mBDNF ratio of ~9:1, they survived until adulthood. The levels of mBDNF were reduced by 57% in heterozygous mutant mice, which exhibited a depressive-like behavior in the tail suspension test and weight gain when housed in social isolation, showing that impaired proBDNF cleavage contributes to stress-induced depressive-like phenotypes. Furthermore, socially isolated heterozygous mice displayed a pronounced deficit in daily nestbuilding behaviors. These findings suggest that the decreased production of mBDNF by impaired proBDNF cleavage disturbs daily activities in mice.

Citrus fruits have several potential benefits for maintaining our health. In this study, we investigated the anti-obesity effects of immature Citrus tumida hort. ex Tanaka (C. tumida) peels using an acute obesity mice model. C57BL/6J male mice were divided into 2 groups; HFD-LL subjected to a high fat diet (HFD) and constant light exposure (LL), and HFDC-LL subjected to a HFD containing immature peel powder of C. tumida (5% w/w) and LL. Dietary ingestion of immature C. tumida peels significantly suppressed body weight gain following decreased epidydimal, perirenal, and subcutaneous fat weights. Blood levels of triglyceride and total cholesterol in the HFDC-LL were significantly lower than those in the HFD-LL group; however, there was no significant difference in food or calorie intake between the 2 groups. These results suggested that immature C. tumida peels have a beneficial effect on the prevention of obesity and metabolic syndrome via its biochemical activities of lipid metabolism.

Like animals, plants accommodate a rich diversity of microbes, typically without discernible disease symptoms. How their pathogenesis is prevented in the host remains obscure. Here, we show that the root-infecting fungus Colletotrichum fructicola of the C. gloeosporioides clade (CgE), isolated from field-grown healthy Brassicaceae plants, inhibits growth of pathogenic fungi in Arabidopsis thaliana, in a phosphate status-dependent manner. Loss of host ethylene signaling or phytoalexins, camalexin or indole glucosinolates, however, allows CgE to display pathogenesis, suggesting host contributions to endophytic CgE colonization and benefit. Compared to a closely-related C. gloeosporioides pathogen (CgP), CgE is characterized by genome expansion and >700 fungal genes (4.34%) specifically induced in the host roots when co-inoculated with CgP, including genes related to fungal secondary metabolism. This may underlie antimicrobial tolerance of CgE and its dominance over pathogenic fungi within the host, pointing to a role for fungus-fungus competition in asymptomatic fungal colonization in plants.

BackgroundThe cryptic parasite Sparganum proliferum proliferates in humans and invades tissues and organs. Only scattered cases have been reported, but S. proliferum infection is always fatal. However, the S. proliferum phylogeny and lifecycle are still an enigma.ResultsTo investigate the phylogenetic relationships between S. proliferum and other cestode species, and to examine the underlying mechanisms of pathogenicity, we sequenced the entire S. proliferum genome. Additionally, S. proliferum plerocercoid larvae transcriptome analyses were performed to identify genes involved in asexual reproduction in the host. The genome sequences confirmed that the S. proliferum genetic sequence is distinct from that of the closely related Spirometra erinaceieuropaei. Moreover, nonordinal extracellular matrix coordination allows for asexual reproduction in the host and loss of sexual maturity in S. proliferum is related to its fatal pathogenicity in humans.ConclusionsThe high-quality reference genome sequences generated should prove valuable for future studies of pseudophyllidean tapeworm biology and parasitism.

To investigate the effects of environmental enrichment on laboratory monkeys, we studied behavioral and physiological differences following changes in housing conditions. Ten male and female juvenile cynomolgus monkeys were first housed in pairs for 8 weeks after quarantine/acclimatization (singly housed) and subsequently housed alone for the next 8 weeks. Monkeys were subjected to evaluations of body weight gain, stereotypic or affiliative behaviors, cortisol, 4-ethylphenyl sulfate (4EPS) and catecholamine concentrations in biological samples, and blood chemistry tests under both housing conditions. Under paired housing, the stereotypic behavioral score decreased in both sexes, and the affiliative behavioral score increased in males and showed an increasing trend in females. Under single housing, the stereotypic score increased in both sexes, and the affiliative score decreased in males. Paired housing decreased serum calcium and urine cortisol concentrations in both sexes and decreased plasma cortisol in males and plasma 4EPS concentrations in females. The stereotypic score was positively correlated with serum calcium, plasma and urine cortisol, and plasma 4EPS concentration and negatively correlated with the affiliative score. The feces painting score, affiliative score, and plasma cortisol and serum calcium concentrations showed sex differences, suggesting differences in responsiveness to environmental changes between males and females. In conclusion, paired housing improved behavioral abnormalities in juvenile cynomolgus monkeys, suggesting that it may be an effective environmental enrichment paradigm. Calcium, cortisol, and 4EPS concentrations in biological samples may be useful indices for evaluating the effects of environmental enrichment.

The intestinal ecosystem is involved in the pathogenesis of mood disorders such as depression. Intestinal microbes can affect the central nervous system through the gut-brain axis, which raises the possibility of using probiotics for preventing depression. In this study, we examined the effect of heat-inactivated Lactobacillus gasseri CP2305 (CP2305) in a subchronic and mild social defeat stress (sCSDS) mouse model. sCSDS suppressed food intake. However, dietary CP2305 intake rescued it, suggesting that CP2305 improved the decreased appetite in sCSDS mice. sCSDS did not alter the gene expression of brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3 in the hippocampus. However, dietary CP2305 provided following sCSDS increased the gene expression of these neurotrophins in the hippocampus. These findings suggest that CP2305 supplementation would aid in preventing psychosocial stress-induced disorders.

Chromatin architecture governs cell lineages by regulating the specific gene expression; however, its role in the diversity of cancer development remains unknown. Among pancreatic cancers, pancreatic ductal adenocarcinoma (PDAC) and intraductal papillary mucinous neoplasms (IPMN) with an associated invasive carcinoma (IPMNinv) arise from 2 distinct precursors, and their fundamental differences remain obscure. Here, we aimed to assess the difference of chromatin architecture regulating the transcriptional signatures or biological features in pancreatic cancers.We established 28 human organoids from distinct subtypes of pancreatic tumors, including IPMN, IPMNinv, and PDAC. We performed exome sequencing (seq), RNA-seq, assay for transposase-accessible chromatin-seq, chromatin immunoprecipitation-seq, high-throughput chromosome conformation capture, and phenotypic analyses with short hairpin RNA or clustered regularly interspaced short palindromic repeats interference.Established organoids successfully reproduced the histology of primary tumors. IPMN and IPMNinv organoids harbored GNAS, RNF43, or KLF4 mutations and showed the distinct expression profiles compared with PDAC. Chromatin accessibility profiles revealed the gain of stomach-specific open regions in IPMN and the pattern of diverse gastrointestinal tissues in IPMNinv. In contrast, PDAC presented an impressive loss of accessible regions compared with normal pancreatic ducts. Transcription factor footprint analysis and functional assays identified that MNX1 and HNF1B were biologically indispensable for IPMN lineages. The upregulation of MNX1 was specifically marked in the human IPMN lineage tissues. The MNX1-HNF1B axis governed a set of genes, including MYC, SOX9, and OLFM4, which are known to be essential for gastrointestinal stem cells. High-throughput chromosome conformation capture analysis suggested the HNF1B target genes to be 3-dimensionally connected in the genome of IPMNinv.Our organoid analyses identified the MNX1-HNF1B axis to be biologically significant in IPMN lineages.

Among Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H7 strains, those producing Stx2a cause more severe diseases. Atypical STEC O157:H7 strains showing a β-glucuronidase-positive phenotype (GP STEC O157:H7) have rarely been isolated from humans, mostly from persons with asymptomatic or mild infections; Stx2a-producing strains have not been reported. We isolated, from a patient with bloody diarrhea, a GP STEC O157:H7 strain (PV15-279) that produces Stx2a in addition to Stx1a and Stx2c. Genomic comparison with other STEC O157 strains revealed that PV15-279 recently emerged from the stx1a/stx2c-positive GP STEC O157:H7 clone circulating in Japan. Major virulence genes are shared between typical (β-glucuronidase-negative) and GP STEC O157:H7 strains, and the Stx2-producing ability of PV15-279 is comparable to that of typical STEC O157:H7 strains; therefore, PV15-279 presents a virulence potential similar to that of typical STEC O157:H7. This study reveals the importance of GP O157:H7 as a source of highly pathogenic STEC clones.

Genome editing is a powerful tool as a new breeding technology including for aquaculture because of the high efficiency of gene targeting without the requirement for exogenous gene integration. CRISPR/Cas9 system, a genome editing tool, has been widely used in various species due to its efficiency and flexibility. We demonstrate the establishment of a new breed of myostatin (Pm-mstn) complete knockout red sea bream (Pagrus major) using CRISPR/Cas9. This is the first report of the establishment of a new breed in aquaculture marine fish using genome editing. The mutations were formed by deletions in the first exon of the Pm-mstn, which cause disruption of the C-terminal active domain of MSTN. The breed exhibited a 16% increase of skeletal muscle, that is, an increase of edible parts. The breed showed the phenotype of short body length and small centrum, which is not observed in mice and other teleost fish. We established the homozygous gene disrupted breed in 2 years, which is far shorter than the conventional breeding method. Our study indicates that genome editing can accelerate the speed of aquaculture fish breeding.

How genetic information is modified to generate phenotypic variation within a species is one of the central questions in evolutionary biology. Here we focus on the striking intraspecific diversity of >200 aposematic elytral (forewing) colour patterns of the multicoloured Asian ladybird beetle, Harmonia axyridis, which is regulated by a tightly linked genetic locus h. Our loss-of-function analyses, genetic association studies, de novo genome assemblies, and gene expression data reveal that the GATA transcription factor gene pannier is the major regulatory gene located at the h locus, and suggest that repeated inversions and cis-regulatory modifications at pannier led to the expansion of colour pattern variation in H. axyridis. Moreover, we show that the colour-patterning function of pannier is conserved in the seven-spotted ladybird beetle, Coccinella septempunctata, suggesting that H. axyridis’ extraordinary intraspecific variation may have arisen from ancient modifications in conserved elytral colour-patterning mechanisms in ladybird beetles.

Significance Environmental temperature acclimation is essential to animal survival, yet thermoregulation mechanisms remain poorly understood. In this study, we describe Ca 2+ -dependent endoribonuclease (EndoU) ENDU-2 located in ADL chemosensory neurons and specific muscle cells as a regulator of multiple pleiotropic phenomena including cold tolerance, life span, and brood size through cell-autonomous and cell-nonautonomous pathways in nematode Caenorhabditis elegans . Ca 2+ imaging revealed ADL temperature response to be the result of transient receptor potential (TRP) channel activity and regulated by ENDU-2 via cell-autonomous and cell-nonautonomous pathways. Transcriptome analysis revealed that ENDU-2 influences expression of the caspase gene ced-3 . Moreover, ENDU-2 downregulates cold tolerance and synaptic remodeling in the dorsal nerve cord through caspase signaling. We therefore propose a model for cold tolerance regulation that occurs via EndoU action.

Plastid genomes are not normally celebrated for being large. But researchers are steadily uncovering algal lineages with big and, in rare cases, enormous plastid DNAs (ptDNAs), such as volvocine green algae. Plastome sequencing of five different volvocine species has revealed some of the largest, most repeat-dense plastomes on record, including that of Volvox carteri (∼525 kb). Volvocine algae have also been used as models for testing leading hypotheses on organelle genome evolution (e.g., the mutational hazard hypothesis), and it has been suggested that ptDNA inflation within this group might be a consequence of low mutation rates and/or the transition from a unicellular to multicellular existence. Here, we further our understanding of plastome size variation in the volvocine line by examining the ptDNA sequences of the colonial species Yamagishiella unicocca and Eudorina sp. NIES-3984 and the multicellular Volvox africanus, which are phylogenetically situated between species with known ptDNA sizes. Although V. africanus is closely related and similar in multicellular organization to V. carteri, its ptDNA was much less inflated than that of V. carteri. Synonymous- and noncoding-site nucleotide substitution rate analyses of these two Volvox ptDNAs suggest that there are drastically different plastid mutation rates operating in the coding versus intergenic regions, supporting the idea that error-prone DNA repair in repeat-rich intergenic spacers is contributing to genome expansion. Our results reinforce the idea that the volvocine line harbors extremes in plastome size but ultimately shed doubt on some of the previously proposed hypotheses for ptDNA inflation within the lineage.

Diverse commercially available feeds are used in animal studies according to the purpose of the studies. We sought to understand the relationship between feed ingredients and their effects on animal physiology and behaviors. Here, we investigated how male laboratory mice (C57BL/6J ("B6") mice) were affected by chronic feeding with two commercially available diets, a non-purified diet (MF) and a semi-purified diet (AIN-93G). In B6 mice, both diets similarly induced spontaneous activities in the home cage and the open field box, anxiety in the elevated plus maze test, and depressive-like behaviors in tail-suspension and forced-swimming tests, and with both diets, similar data were obtained on calorie intake, water intake, body weight gain, and plasma corticosterone levels. By contrast, liver weight was significantly higher in MF-fed B6 mice than in AIN-93G-fed B6 mice. Furthermore, the cecum microbiome was drastically affected by the diets, and, specifically, Allobaculum was the major genus (43.4%) in the cecum microbiota of AIN-93G-fed mice but its abundance was reduced (to 3.8%) in the case of MF-fed mice. Future studies should address whether the differences in diet purity and cecum microbiota influence brain functions and behaviors in B6 mice.

Three strains, JCM 5343T, JCM 5344 and JCM 1130, currently identified as Lactobacillus gasseri , were investigated using a polyphasic taxonomic approach. Although these strains shared high 16S rRNA gene sequence similarities with L. gasseri ATCC 33323T (99.9 %), they formed a clade clearly distinct from ATCC 33323T based on whole-genome relatedness. The average nucleotide identity and in silico DNA–DNA hybridization values of these three strains compared to L. gasseri ATCC 33323T were 93.4–93.7 and 53.1–54.1 %, respectively, and both were less than the widely accepted threshold to distinguish two species (95 and 70 %, respectively). The three strains were Gram-stain positive, non-motile, non-spore-forming, catalase-negative and rod-shaped bacteria. They grew at 25–45 °C and in the presence of 2.0 % (w/v) NaCl. The major fatty acids of the three strains were C16 : 0 and C18 : 1 ω9c. Based on phylogenetic analyses of the phenylalanyl-tRNA synthase alpha subunit and RNA polymerase alpha subunit genes, and on phenotypic and chemotaxonomic characteristics, strains JCM 5343T, JCM 5344 and JCM 1130 represent a novel species distinct from L. gasseri , for which we propose the name Lactobacillus paragasseri sp. nov. In addition, a large portion of genomes currently labelled as L. gasseri in the public sequence database should be reclassified as L. paragasseri based on whole-genome relatedness.

Human intestinal absorption is estimated using a human colon carcinoma cell line (Caco-2) cells from human colorectal adenocarcinoma, intestinal perfusion, or a mammalian model. These current evaluation systems are limited in their ability to estimate human intestinal absorption. In addition, in vivo evaluation systems using laboratory animals such as mice and rats entail animal ethics problems, and it is difficult to screen compounds on a large scale at the drug discovery stage. Thus, we propose the use of Bombyx mori larvae for evaluation of intestinal absorption of compounds as an alternative system in this study. First, to compare the characteristics among Caco-2 cells, human intestine, and B. mori larval midgut, we analyzed their RNA-seq data, and we found 26 drug transporters common to humans and B. mori. Next, we quantitatively developed an oral administration technique in B. mori and established a method using silkworm B. mori larvae that can easily estimate the intestinal permeability of compounds. Consequently, we could determine the dose and technique for oral administration in B. mori larvae. We also developed a B. mori model to evaluate the intestinal permeability of orally administered. Our constructed evaluation system will be useful for evaluating intestinal permeability in medical drug development.

Black Aspergillus luchuensis and its white albino mutant are essential fungi for making alcoholic beverages in Japan. A large number of industrial strains have been created using novel isolation or gene/genome mutation techniques. Such mutations influence metabolic and phenotypic characteristics in industrial strains, but few comparative studies of inter-strain mutation have been conducted. We carried out comparative genome analyses of 8 industrial strains of A. luchuensis and A. kawachii IFO 4308, the latter being the first albino strain to be isolated. Phylogenetic analysis based on 8938 concatenated genes exposed the diversity of black koji strains and uniformity among albino industrial strains, suggesting that passaged industrial albino strains have more genetic mutations compared with strain IFO 4308 and black koji strains. Comparative analysis showed that the albino strains had mutations in genes not only for conidial pigmentation but also in those that encode N-terminal acetyltransferase A and annexin XIV-like protein. The results also suggest that some mutations may have emerged through subculturing of albino strains. For example, mutations in the genes for isocitrate lyase and sugar transporters were observed only in industrial albino strains. This implies that selective pressure for increasing enzyme activity or secondary metabolites may have influenced the mutation of genes associated with environmental stress responses in A. luchuensis albino strains. Our study clarifies hitherto unknown genetic and metabolic characteristics of A. luchuensis industrial strains and provides potential applications for comparative genome analysis for breeding koji strains.

Lactate is known to have diverse roles in the brain at the molecular and behavioral levels under both physiological and pathophysiological conditions, such as learning and memory and regulation of mood. Recently, a novel post-translational modification called lysine lactylation has been found in histone H3 of mouse macrophages, and the lactylation levels paralleled the intracellular lactate levels. However, it was unknown whether lysine lactylation occurs in brain cells, and if it does, whether lactylation is induced by the stimuli that accompany changes in lactate levels. Herein, for the first time, we reveal that lysine lactylation in brain cells is regulated by systemic changes in lactate levels, neural excitation, and behaviorally relevant stimuli. The expression level of pan-lysine lactylation was increased by lactate treatment and by high-potassium-induced depolarization in cultured primary neurons; these increases were attenuated by pharmacological inhibition of monocarboxylate transporter 2 and lactate dehydrogenase, respectively, suggesting that both cell-autonomous and non-cell-autonomous neuronal mechanisms are involved in overall lysine lactylation. In vivo, electroconvulsive stimulation increased lysine lactylation levels in the prefrontal cortices of mice, and its levels were positively correlated with the expression levels of the neuronal activity marker c-Fos on an individual cell basis. To investigate the behavior of lysine lactylation in the brain in response to physiologically relevant stimuli, we adopted the social defeat stress model of depression in which lactate levels increase in the brains of mice. Lactylation levels were increased in the prefrontal cortices of the defeated mice, which was accompanied by increased c-Fos expression, decreased social behaviors, and increased anxiety-like behaviors, suggesting that stress-induced neuronal excitation may induce lysine lactylation, thereby affecting mood-related behaviors. Further, we identified 53 candidate lysine-lactylated proteins in the mouse cortex and found that lactylation levels in histone H1 increased in response to defeat stress. The present study may open up an avenue for exploration of a novel role of lactate mediated by protein lactylation in the brain.

Acquisition of new cis -regulatory elements (CREs) can cause alteration of developmental gene regulation and may introduce morphological novelty in evolution. Although structural variation in the genome generated by chromosomal rearrangement is one possible source of new CREs, only a few examples are known, except for cases of retrotransposition. In this study, we show the acquisition of novel regulatory sequences as a result of large genomic insertion in the spontaneous mouse mutation Hammer toe ( Hm ). Hm mice exhibit syndactyly with webbing, due to suppression of interdigital cell death in limb development. We reveal that, in the Hm genome, a 150-kb noncoding DNA fragment from chromosome 14 is inserted into the region upstream of the Sonic hedgehog ( Shh ) promoter in chromosome 5. Phenotyping of mouse embryos with a series of CRISPR/Cas9-aided partial deletion of the 150-kb insert clearly indicated that two different regions are necessary for the syndactyly phenotype of Hm . We found that each of the two regions contains at least one enhancer for interdigital regulation. These results show that a set of enhancers brought by the large genomic insertion elicits the interdigital Shh expression and the Hm phenotype. Transcriptome analysis indicates that ectopic expression of Shh up-regulates Chordin ( Chrd ) that antagonizes bone morphogenetic protein signaling in the interdigital region. Indeed, Chrd- overexpressing transgenic mice recapitulated syndactyly with webbing. Thus, the Hm mutation provides an insight into enhancer acquisition as a source of creation of novel gene regulation.

The arms race between parasitic sequences and their hosts is a major driving force for evolution of gene control systems. Since transposable elements (TEs) are potentially deleterious, eukaryotes silence them by epigenetic mechanisms such as DNA methylation. Little is known about how TEs counteract silencing to propagate during evolution. Here, we report behavior of sequence-specific anti-silencing proteins used by Arabidopsis TEs and evolution of those proteins and their target sequences. We show that VANC, a TE-encoded anti-silencing protein, induces extensive DNA methylation loss throughout TEs. Related VANC proteins have evolved to hypomethylate TEs of completely different spectra. Targets for VANC proteins often form tandem repeats, which vary considerably between related TEs. We propose that evolution of VANC proteins and their targets allow propagation of TEs while causing minimal host damage. Our findings provide insight into the evolutionary dynamics of these apparently “selfish” sequences. They also provide potential tools to edit epigenomes in a sequence-specific manner., Eukaryotes often silence transposable elements (TEs) via DNA methylation. Here, the authors show that evolution of VANC, an Arabidopsis anti-silencing factor, and its target motifs allows sequence-specific demethylation, suggesting a way TEs can proliferate while minimizing damage to the host genome.

Land plants produce specialized low molecular weight metabolites to adapt to various environmental stressors, such as UV radiation, pathogen infection, wounding and animal feeding damage. Due to the large variety of stresses, plants produce various chemicals, particularly plant species-specific alkaloids, through specialized biosynthetic pathways. In this study, using a draft genome sequence and querying known biosynthetic cytochrome P450 (P450) enzyme-encoding genes, we characterized the P450 genes involved in benzylisoquinoline alkaloid (BIA) biosynthesis in California poppy (Eschscholzia californica), as P450s are key enzymes involved in the diversification of specialized metabolism. Our in silico studies showed that all identified enzyme-encoding genes involved in BIA biosynthesis were found in the draft genome sequence of approximately 489 Mb, which covered approximately 97% of the whole genome (502 Mb). Further analyses showed that some P450 families involved in BIA biosynthesis, i.e. the CYP80, CYP82 and CYP719 families, were more enriched in the genome of E. californica than in the genome of Arabidopsis thaliana, a plant that does not produce BIAs. CYP82 family genes were highly abundant, so we measured the expression of CYP82 genes with respect to alkaloid accumulation in different plant tissues and two cell lines whose BIA production differs to estimate the functions of the genes. Further characterization revealed two highly homologous P450s (CYP82P2 and CYP82P3) that exhibited 10-hydroxylase activities with different substrate specificities. Here, we discuss the evolution of the P450 genes and the potential for further genome mining of the genes encoding the enzymes involved in BIA biosynthesis., ハナビシソウゲノムの解読から見えてきたイソキノリンアルカロイド生合成酵素遺伝子の進化 --植物が有用物質生産系を進化させる仕組みの解読--. 京都大学プレスリリース. 2018-01-26.

Nitrification, the microbial oxidation of ammonia to nitrate via nitrite, occurs in a wide range of acidic soils. However, the ammonia-oxidizing bacteria (AOB) that have been isolated from soil to date are acid-sensitive. Here we report the isolation and characterization of an acid-adapted AOB from an acidic agricultural soil. The isolated AOB, strain TAO100, is classified within the Gammaproteobacteria based on phylogenetic characteristics. TAO100 can grow in the pH range of 5-7.5 and survive in highly acidic conditions until pH 2 by forming cell aggregates. Whereas all known gammaproteobacterial AOB (γ-AOB) species, which have been isolated from marine and saline aquatic environments, are halophiles, TAO100 is not phenotypically halophilic. Thus, TAO100 represents the first soil-originated and non-halophilic γ-AOB. The TAO100 genome is considerably smaller than those of other γ-AOB and lacks several genes associated with salt tolerance which are unnecessary for survival in soil. The ammonia monooxygenase subunit A gene of TAO100 and its transcript are higher in abundance than those of ammonia-oxidizing archaea and betaproteobacterial AOB in the strongly acidic soil. These results indicate that TAO100 plays an important role in the nitrification of acidic soils. Based on these results, we propose TAO100 as a novel species of a new genus, Candidatus Nitrosoglobus terrae.

Chlamydomonadalean green algae are no stranger to linear mitochondrial genomes, particularly members of the Reinhardtinia clade. At least nine different Reinhardtinia species are known to have linear mitochondrial DNAs (mtDNAs), including the model species Chlamydomonas reinhardtii. Thus, it is no surprise that some have suggested that the most recent common ancestor of the Reinhardtinia clade had a linear mtDNA. But the recent uncovering of circular-mapping mtDNAs in a range of Reinhardtinia algae, such as Volvox carteri and Tetrabaena socialis, has shed doubt on this hypothesis. Here, we explore mtDNA sequence and structure within the colonial Reinhardtinia algae Yamagishiella unicocca and Eudorina sp. NIES-3984, which occupy phylogenetically intermediate positions between species with opposing mtDNA mapping structures. Sequencing and gel electrophoresis data indicate that Y. unicocca has a linear monomeric mitochondrial genome with long (3 kb) palindromic telomeres. Conversely, the mtDNA of Eudorina sp., despite having an identical gene order to that of Y. unicocca, assembled as a circular-mapping molecule. Restriction digests of Eudorina sp. mtDNA supported its circular map, but also revealed a linear monomeric form with a matching architecture and gene order to the Y. unicocca mtDNA. Based on these data, we suggest that there have been at least three separate shifts in mtDNA conformation in the Reinhardtinia, and that the common ancestor of this clade had a linear monomeric mitochondrial genome with palindromic telomeres.

We thank Renner (1) for summarizing and comparing our study (2) with that of Cui et al. (3). Renner questions the logic of our domesticated/wild classification. As already mentioned (2), without a genetic tool, plant collectors assigned the cultivar/wild status purely based on whether a plant was cultivated. However, many cultures, such as the Amis people in Taiwan, frequently consume wild vegetables and sometimes transplant them into gardens. A plant collector would record such plant as a “cultivar” despite it genetically being a wild accession (e.g., THMC170). All of our analyses followed the grouping from solid genomic evidence. We agree that a broader sampling would be beneficial, if funding permits. Cui … [↵][1]1To whom correspondence may be addressed. Email: chengrueilee{at}ntu.edu.tw. [1]: #xref-corresp-1-1

The Oryza officinalis complex is the largest species group in Oryza, with more than nine species from four continents, and is a tertiary gene pool that can be exploited in breeding programs for the improvement of cultivated rice. Most diploid and tetraploid members of this group have a C genome. Using a new reference C genome for the diploid species O. officinalis, and draft genomes for two other C genome diploid species Oryza eichingeri and Oryza rhizomatis, we examine the influence of transposable elements on genome structure and provide a detailed phylogeny and evolutionary history of the Oryza C genomes. The O. officinalis genome is 1.6 times larger than the A genome of cultivated Oryza sativa, mostly due to proliferation of Gypsy type long-terminal repeat transposable elements, but overall syntenic relationships are maintained with other Oryza genomes (A, B, and F). Draft genome assemblies of the two other C genome diploid species, Oryza eichingeri and Oryza rhizomatis, and short-read resequencing of a series of other C genome species and accessions reveal that after the divergence of the C genome progenitor, there was still a substantial degree of variation within the C genome species through proliferation and loss of both DNA and long-terminal repeat transposable elements. We provide a detailed phylogeny and evolutionary history of the Oryza C genomes and a genomic resource for the exploitation of the Oryza tertiary gene pool., Genome Biology and Evolution, 12 (4), ISSN:1759-6653

BackgroundPopulation outbreaks of the crown-of-thorns starfish (Acanthaster plancisensu lato; COTS), a primary predator of reef-building corals in the Indo-Pacific Ocean, are major concerns in coral reef management. While biological and ecological knowledge of COTS has been accumulating since the 1960s, little is known about its associated bacteria. The aim of this study was to provide fundamental information on dominant COTS-associated bacteria through a multifaceted molecular approach.MethodsA total of 205 COTS individuals from 17 locations throughout the Indo-Pacific Ocean were examined for the presence of COTS-associated bacteria. We conducted 16S rRNA metabarcoding of COTS to determine the bacterial profiles of different parts of the body, and generated a full-length 16S rRNA gene sequence from a single dominant bacterium, which we designated COTS27. We performed phylogenetic analysis to determine the taxonomy, screening of COTS27 across the Indo-Pacific, FISH to visualize it within the COTS tissues, and reconstruction of the chromosome from the hologenome sequence data.ResultsWe discovered that a single bacterium exists at high densities in the subcuticular space in COTS forming a biofilm-like structure between the cuticle and the epidermis. COTS27 belongs to a clade that presumably represents a distinct order (so-called marine spirochetes) in the phylumSpirochaetesand is universally present in COTS throughout the Indo-Pacific Ocean. The reconstructed genome of COTS27 includes some genetic traits that are probably linked to adaptation to marine environments and evolution as an extracellular endosymbiont in subcuticular spaces.ConclusionsCOTS27 can be found in three allopatrically speciated COTS species, ranging from northern Red Sea to the Pacific, implying that symbiotic relationship arose before the speciation (approximately 2 million years ago). The universal association of COTS27 with COTS and nearly mono-specific association at least with the Indo-Pacific COTS potentially provides a useful model system for studying symbiont-host interactions in marine invertebrates.

Nest building is an instinctive behavior toward protection from predators, body temperature regulation, and courtship. Previously, we discovered that acute and chronic social defeat stress suppresses the onset of nest-building behavior in male mice (C57BL/6J). Here, we analyzed nest building and other behavioral deficits induced by acute social defeat stress (ASDS). We utilized a customized cage and specifically developed observational programs for nest building, social avoidance, and other behaviors using an infrared depth camera to acquire three-dimensional (3D) data of animal behavior (Negura system). We determined the volume of nesting materials from these 3D depth images. Mice exposed to ASDS showed increased spontaneous activities, decreased rearing, and delayed nest building; however, nest-building activity was gradually recovered during the dark period of the 24 hr observation interval. At the endpoint following 24 hr, the ASDS and control groups showed no differences in nest volumes. Furthermore, we observed the time courses of both nest building and social avoidance behaviors and their relationship using the Negura system. Our data demonstrated a weak positive correlation between nest-building delay and social avoidance in ASDS mice. The Negura system can observe various behaviors that reflect the effects of social defeat stress.

Feeding behavior is closely related to hypothalamic malonyl-CoA level in the brain and diet-induced obesity affects total CoA pools in liver. Herein, we performed a comprehensive analysis of the CoA pools formed in thirteen tissues of Zucker and Zucker diabetic fatty (ZDF) rats. Hypothalamic malonyl-CoA levels in obese rats remained low and were almost the same as those of lean rats, despite obese rats having much higher content of leptin, insulin, and glucose in their sera. Regardless of the fa-genotypes, larger total CoA pools were formed in the livers of ZDF rats and the size of hepatic total CoA pools in Zucker rats showed almost one tenth of the size of ZDF rats. The decreased total CoA pool sizes in Zucker rats was observed in the brown adipose tissues, while ZDF-fatty rats possessed 6% of total CoA pool in the lean rats in response to fa deficiency. This substantially lower CoA content in the obese rats would be disadvantageous to non-shivering thermogenesis. Thus, comparing the intracellular CoA behaviors between Zucker and ZDF rats, as well as the lean and fatty rats of each strain would help to elucidate features of obesity and type 2 diabetes in combination with result (s) of differential gene expression analysis and/or comparative genomics.

Caenorhabditis elegans mechanoreceptors located in ASG sensory neurons have been found to sense ambient temperature, which is a key trait for animal survival. Here, we show that experimental loss of xanthine dehydrogenase (XDH‐1) function in AIN and AVJ interneurons results in reduced cold tolerance and atypical neuronal response to changes in temperature. These interneurons connect with upstream neurons such as the mechanoreceptor‐expressing ASG. Ca2+ imaging revealed that ASG neurons respond to warm temperature via the mechanoreceptor DEG‐1, a degenerin/epithelial Na+ channel (DEG/ENaC), which in turn affects downstream AIN and AVJ circuits. Ectopic expression of DEG‐1 in the ASE gustatory neuron results in the acquisition of warm sensitivity, while electrophysiological analysis revealed that DEG‐1 and human MDEG1 were involved in warm sensation. Taken together, these results suggest that cold tolerance is regulated by mechanoreceptor‐mediated circuit calculation., A DEG/ENaC‐type mechanoreceptor‐mediated neural circuit regulates cold tolerance in Caenorhabditis elegans.

Phages and plasmids play important roles in bacterial evolution and diversification. Although many draft genomes have been generated, phage and plasmid genomes are usually fragmented, limiting our understanding of their dynamics. Here, we performed a systematic analysis of 239 draft genomes and 7 complete genomes of Shiga toxin (Stx)-producing Escherichia coli O145:H28, the major virulence factors of which are encoded by prophages (PPs) or plasmids. The results indicated that PPs are more stably maintained than plasmids. A set of ancestrally acquired PPs was well conserved, while various PPs, including Stx phages, were acquired by multiple sublineages. In contrast, gains and losses of a wide range of plasmids have frequently occurred across the O145:H28 lineage, and only the virulence plasmid was well conserved. The different dynamics of PPs and plasmids have differentially impacted the pangenome of O145:H28, with high proportions of PP- and plasmid-associated genes in the variably present and rare gene fractions, respectively. The dynamics of PPs and plasmids have also strongly impacted virulence gene repertoires, such as the highly variable distribution of stx genes and the high conservation of a set of type III secretion effectors, which probably represents the core effectors of O145:H28 and the genes on the virulence plasmid in the entire O145:H28 population. These results provide detailed insights into the dynamics of PPs and plasmids, and show the application of genomic analyses using a large set of draft genomes and appropriately selected complete genomes.

How evolutionary novelties have arisen is one of the central questions in evolutionary biology. Pre-existing gene regulatory networks or signaling pathways have been shown to be co-opted for building novel traits in several organisms. However, the structure of entire gene regulatory networks and evolutionary events of gene co-option for emergence of a novel trait are poorly understood. In this study, we used a novel wing pigmentation pattern of the polka-dotted fruit fly, and identified the complete set of genes for pigmentation pattern formation byde novogenome sequencing and transcriptome analyses. In pigmentation areas of wings, 151 genes were positively or negatively regulated bywingless, a master regulator of wing pigmentation. Genes for neural development, Wnt signaling, Dpp signaling, Zinc finger transcription factors, and effectors (such as enzymes) for melanin pigmentation were included among these 151 genes. None of the known regulatory genes that regulate pigmentation pattern formation in other fruit fly species were included. Our results suggest that the novel pigmentation pattern of the polka-dotted fruit fly emerged through multi-step co-options of multiple gene regulatory networks, signaling pathways, and effector genes, rather than recruitment of one large gene circuit.

The exposure of germ cells to radiation introduces mutations in the genomes of offspring, and a previous whole-genome sequencing study indicated that the irradiation of mouse sperm induces insertions/deletions (indels) and multisite mutations (clustered single nucleotide variants and indels). However, the current knowledge on the mutation spectra is limited, and the effects of radiation exposure on germ cells at stages other than the sperm stage remain unknown. Here, we performed whole-genome sequencing experiments to investigate the exposure of spermatogonia and mature oocytes. We compared de novo mutations in a total of 24 F1 mice conceived before and after the irradiation of their parents. The results indicated that radiation exposure, 4 Gy of gamma rays, induced 9.6 indels and 2.5 multisite mutations in spermatogonia and 4.7 indels and 3.1 multisite mutations in mature oocytes in the autosomal regions of each F1 individual. Notably, we found two types of deletions, namely, small deletions (mainly 1~12 nucleotides) in non-repeat sequences, many of which showed microhomology at the breakpoint junction, and single-nucleotide deletions in mononucleotide repeat sequences. The results suggest that these deletions and multisite mutations could be a typical signature of mutations induced by parental irradiation in mammals.

Phages and plasmids play important roles in bacterial evolution and diversification. Although many draft genomes have been generated, phage and plasmid genomes are usually fragmented, limiting our understanding of their dynamics. Here, we performed a systematic analysis of 239 draft genomes and 7 complete genomes of Shiga toxin (Stx)-producing Escherichia coli O145:H28, the major virulence factors of which are encoded by prophages (PPs) or plasmids. The results indicated that PPs are more stably maintained than plasmids. A set of ancestrally acquired PPs was well conserved, while various PPs, including Stx phages, were acquired by multiple sublineages. In contrast, gains and losses of a wide range of plasmids have frequently occurred across the O145:H28 lineage, and only the virulence plasmid was well conserved. The different dynamics of PPs and plasmids have differentially impacted the pangenome of O145:H28, with high proportions of PP- and plasmid-associated genes in the variably present and rare gene fractions, respectively. The dynamics of PPs and plasmids have also strongly impacted virulence gene repertoires, such as the highly variable distribution of stx genes and the high conservation of a set of type III secretion effectors, which probably represents the core effectors of O145:H28 and the genes on the virulence plasmid in the entire O145:H28 population. These results provide detailed insights into the dynamics of PPs and plasmids, and show the application of genomic analyses using a large set of draft genomes and appropriately selected complete genomes.

How evolutionary novelties have arisen is one of the central questions in evolutionary biology. Preexisting gene regulatory networks or signaling pathways have been shown to be co-opted for building novel traits in several organisms. However, the structure of entire gene regulatory networks and evolutionary events of gene co-option for emergence of a novel trait are poorly understood. In this study, to explore the genetic and molecular bases of the novel wing pigmentation pattern of a polka-dotted fruit fly (Drosophila guttifera), we performed de novo genome sequencing and transcriptome analyses. As a result, we comprehensively identified the genes associated with the pigmentation pattern. Furthermore, we revealed that 151 of these associated genes were positively or negatively regulated by wingless, a master regulator of wing pigmentation. Genes for neural development, Wnt signaling, Dpp signaling, and effectors (such as enzymes) for melanin pigmentation were included among these 151 genes. None of the known regulatory genes that regulate pigmentation pattern formation in other fruit fly species were included. Our results suggest that the novel pigmentation pattern of a polka-dotted fruit fly might have emerged through multistep co-options of multiple gene regulatory networks, signaling pathways, and effector genes, rather than recruitment of one large gene circuit.