Color patterns in angiosperm flowers are produced by spatially and temporally restricted deposition of pigments. Identifying the mechanisms responsible for restricted pigment deposition is a topic of broad interest. Some dicots species develop bicolor petals, which are often caused by the post-transcriptional gene silencing (PTGS) of chalcone synthase (CHS) genes. An Asiatic hybrid lily (Lilium spp.) cultivar Lollypop develops bicolor tepals with pigmented tips and white bases. Here, we analyzed the global transcription of pigmented and non-pigmented tepal parts from Lollypop, to determine the main transcriptomic differences. De novo assembly of RNA-seq data yielded 49,239 contigs (39,426 unigenes), which included a variety of novel transcripts, such as those involved in flavonoid-glycosylation and sequestration and in regulation of anthocyanin biosynthesis. Additionally, 1258 of the unigenes exhibited significantly differential expression between the tepal parts (false discovery rates 2-fold higher in the pigmented parts. Thus, LhMYB12 should be involved in the transcriptional regulation of the biosynthesis genes in bicolor tepals. Other factors that potentially suppress or enhance the expression of anthocyanin biosynthesis genes, including a WD40 gene, were identified, and their involvement in bicolor development is discussed. Our results indicate that the bicolor trait of Lollypop tepals is caused by the transcriptional regulation of anthocyanin biosynthesis genes and that the transcription profile of LhMYB12 provides a clue for elucidating the mechanisms of the trait. The tepal transcriptome constructed in this study will accelerate investigations of the genetic controls of anthocyanin color patterns, including the bicolor patterns, of Lilium spp.

Circular patterns called “fairy rings” in fields are a natural phenomenon that arises through the interaction between basidiomycete fungi and plants. Acceleration or inhibition of plant vegetative growth and the formation of mushroom fruiting bodies are both commonly observed when fairy rings form. The gene of an enzyme involved in the biosynthesis of these regulators was recently isolated in the fairy ring-forming fungus, Lepista sordida. To identify other genes involved in L. sordida fairy ring formation, we used previously generated sequence data to produce a more complete draft genome sequence for this species. Finally, we predicted the metabolic pathways of the plant growth regulators and 29 candidate enzyme-coding genes involved in fairy-ring formation based on gene annotations. Comparisons of protein coding genes among basidiomycete fungi revealed two nitric oxide synthase gene candidates that were uniquely encoded in genomes of fairy ring-forming fungi. These results provide a basis for the discovery of genes involved in fairy ring formation and for understanding the mechanisms involved in the interaction between fungi and plants. We also constructed a new web database F-RINGS (http://bioinf.mind.meiji.ac.jp/f-rings/) to provide the comprehensive genomic information for L. sordida.

The white-rot fungus Phlebia sp. MG-60 shows valuable properties such as high ethanol yield from several lignocellulosic materials, although white-rot fungi commonly degrade woody components to CO2 and H2O. In order to identify genes involved in ethanol production by Phlebia sp. MG-60, we compared genes differentially expressed by the ethanol producing fungus Phlebia sp. MG-60 and the model white-rot fungus Phanerochaete chrysosporium under ethanol fermenting and non-fermenting conditions using next-generation sequencing technologies. mRNAs from mycelia of Phlebia sp. MG-60 and P. chrysosporium under fermenting and non-fermenting conditions were sequenced using the MiSeq system. To detect differentially expressed genes, expression levels were measured in fragments per kilobase of exon per million mapped reads (FPKM). Differentially expressed genes were annotated using BLAST searches, Gene Ontology classifications, and KEGG pathway analysis. Functional analyses of differentially expressed genes revealed that genes involved in glucose uptake, glycolysis, and ethanol synthesis were widely upregulated in Phlebia sp. MG-60 under fermenting conditions. In this study, we provided novel transcriptomic information on Phlebia sp. MG-60, and these RNA-seq data were useful in targeting genes involved in ethanol production for future genetic engineering.

Poly([2-(methacryloyloxy)ethyl]trimethyl ammonium chloride) (METAC) and the gels were prepared and evaluated for their bactericidal and fungicidal activities. The antimicrobial properties of poly(METAC) were tested against Escherichia coli (E. coli), Bacillus subtilis (B. subtilis), Saccharomyces cerevisiae (Sa. cerevisiae), methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa (P. aeruginosa), and Candida albicans (C. albicans). Moreover, the structural forms of the linear and cross-linked poly(METAC) were investigated for their influences on bacterial aggregation, precipitation, and cell-death. To our knowledge, this is the first report on the comparison of the antimicrobial properties of poly(METAC) and poly(METAC)-gels. The bactericidal and fungicidal activities were evaluated by determining minimum inhibitory concentrations (MICs), UV–Vis spectroscopy, and fluorescence and confocal microscopies. The MICs were found to be 123 (MSSA), 123 (MRSA), 123 (P. aeruginosa), 370 (E. coli), 123 (B. subtilis), 370 (C. albicans), and 370 μg/mL (Sa. cerevisiae), as determined by broth dilution, and 370 (MSSA), 370 (MRSA), 370 (P. aeruginosa), 3300 (E. coli), 370 (B. subtilis), 1100 (C. albicans), and >10,000 μg/mL (Sa. cerevisiae), as determined by paper disc diffusion (on solid medium). The poly(METAC)-gels achieved rapid adsorption/precipitation of bacteria via the cationic surface charge. Thus, these poly(METAC)-based polymers can potentially be used as antibacterial materials.

The antimicrobial peptide (AMP) derived from lactoferricin B, LfcinB (4-9) (RRWQWR), and lissamine rhodamine B red-labeled peptide (Rh-LfcinB (4-9)) exhibit strong antimicrobial activities, and they can enter Escherichia coli cells without damaging the cell membranes. Thus, these peptides are cell-penetrating peptide (CPP) -type AMPs. In this study, to elucidate the effect of the membrane potential (Δφ) on the action of the CPP-type AMP, Rh-LfcinB (4-9), we investigated the interactions of Rh-LfcinB (4-9) with single E. coli cells and spheroplasts containing calcein in the cytosol using confocal laser scanning microscopy. At low peptide concentrations, Rh-LfcinB (4-9) entered the cytosol of single E. coli cells and spheroplasts without damaging the cell membranes, and the H⁺-ionophore carbonyl cyanide m-chlorophenyl-hydrazone (CCCP) suppressed its entry. The studies using the time-kill method indicate that these low concentrations of peptide exhibit antimicrobial activity but CCCP inhibits this activity. Next, we investigated the effect of Δφ on the interaction of Rh-LfcinB (4-9) with single giant unilamellar vesicles (GUVs) comprising E. coli polar lipid extracts and containing a fluorescent probe, Alexa Fluor 647 hydrazide. At low concentrations (0.2−0.5 μM), Rh-LfcinB (4-9) showed significant entry to the single GUV lumen without pore formation in the presence of Δφ. The fraction of entry of peptide increased with increasing negative membrane potential, indicating that the rate of peptide entry into the GUV lumen increased with increasing negative membrane potential. These results indicate that Δφ enhances the entry of Rh-LfcinB (4-9) into single E. coli cells, spheroplasts, and GUVs and its antimicrobial activity. IMPORTANCE: Bacterial cells have a membrane potential (Δφ), but the effect of Δφ on action of cell-penetrating peptide-type antimicrobial peptides (AMPs) is not clear. Here, we investigated the effect of Δφ on the action of fluorescent probe-labeled AMP derived from lactoferricin B, Rh-LfcinB (4-9). At low peptide concentrations, Rh-LfcinB (4-9) enters the cytosol of Escherichia coli cells and spheroplasts without damaging their cell membrane, but a protonophore suppresses this entry and its antimicrobial activity. The rate of entry of Rh-LfcinB (4-9) into the giant unilamellar vesicles (GUVs) comprising E. coli lipids without pore formation increases with increasing Δφ. These results indicate that Δφ enhances the antimicrobial activity of Rh-LfcinB (4-9) and hence LfcinB (4-9) by increasing the rate of their entry into the cytosol.