Multidrug Sensitive Yeast Strains, Useful Tools for Chemical Genetics

The budding yeast Saccharomyces cerevisiae is a useful eukaryote model organism for application to chemical biology studies, for example, drug screening, drug evaluation, and target identification. To use yeast for chemical biology research, however, it has been necessary to construct yeast strains suitable for various compounds because of their high drug resistance. Hence, the deletion of all multidrug resistance genes except for those that are important for viability and for genetic experiments/manipulation could increase the drug sensitivity without influencing the transformation, mating, or sporulation efficiency. There are two major factors conferring multidrug resistance in S. cerevisiae: one is the drug efflux system and the other is the permeability barrier. We therefore constructed a strain which shows high sensitivity to multiple drugs by disrupting the drug efflux system using ATP-binding cassette transporters and suppressing the membrane barrier system by introducing an ERG6-inducible system. In this review, we discuss the construction of our multidrug-sensitive yeast strains and their application in chemical biology

Photometry and Polarimetry of 2010 XC15_{15}: Observational Confirmation of E-type Near-Earth Asteroid Pair

Asteroid systems such as binaries and pairs are indicative of physical properties and dynamical histories of the Small Solar System Bodies. Although numerous observational and theoretical studies have been carried out, the formation mechanism of asteroid pairs is still unclear, especially for near-Earth asteroid (NEA) pairs. We conducted a series of optical photometric and polarimetric observations of a small NEA 2010 XC$_{15}$ in 2022 December to investigate its surface properties. The rotation period of 2010 XC$_{15}$ is possibly a few to several dozen hours and color indices of 2010 XC$_{15}$ are derived as $g-r=0.435\pm0.008$, $r-i=0.158\pm0.017$, and $r-z=0.186\pm0.009$ in the Pan-STARRS system. The linear polarization degrees of 2010 XC$_{15}$ are a few percent at the phase angle range of 58$^{\circ}$ to 114$^{\circ}$. We found that 2010 XC$_{15}$ is a rare E-type NEA on the basis of its photometric and polarimetric properties. Taking the similarity of not only physical properties but also dynamical integrals and the rarity of E-type NEAs into account, we suppose that 2010 XC$_{15}$ and 1998 WT$_{24}$ are of common origin (i.e., asteroid pair). These two NEAs are the sixth NEA pair and first E-type NEA pair ever confirmed, possibly formed by rotational fission. We conjecture that the parent body of 2010 XC$_{15}$ and 1998 WT$_{24}$ was transported from the main-belt through the $\nu_6$ resonance or Hungaria region.Comment: Resubmitted to AAS Journals. Any comments are welcom

Multicolor Photometry of Tiny Near-Earth Asteroid 2015 RN35 across a Wide Range of Phase Angles: Possible Mission-accessible A-type Asteroid

peer reviewedStudying small near-Earth asteroids is important in order to understand their dynamical histories and origins as well as to mitigate the damage caused by asteroid impacts on Earth. We report the results of multicolor photometry of the tiny near-Earth asteroid 2015 RN35 using the 3.8 m Seimei telescope in Japan and the TRAPPIST-South telescope in Chile over 17 nights in 2022 December and 2023 January. We observed 2015 RN35 across a wide range of phase angles from 2° to 30° in the g, r, i, and z bands in the Pan-STARRS system. These lightcurves show that 2015 RN35 is in a nonprincipal axis spin state with two characteristic periods of 1149.7 ± 0.3 s and 896.01 ± 0.01 s. We found that the slope of the visible spectrum of 2015 RN35 is as red as asteroid (269) Justitia, one of the very red objects in the main belt, which indicates that 2015 RN35 can be classified as an A- or Z-type asteroid. In conjunction with the shallow slope of the phase curve, we suppose that 2015 RN35 is a high-albedo A-type asteroid. We demonstrated that surface properties of tiny asteroids could be well constrained by intensive observations across a wide range of phase angles. 2015 RN35 is a possible mission-accessible A-type near-Earth asteroid with a small Δv of 11.801 km s-1 in the launch window between 2030 and 2035

Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness

Phytochrome regulates lettuce (Lactuca sativa L. cv. Grand Rapids) seed germination via the control of the endogenous level of bioactive gibberellin (GA). In addition to the previously identified LsGA20ox1, LsGA20ox2, LsGA3ox1, LsGA3ox2, LsGA2ox1, and LsGA2ox2, five cDNAs were isolated from lettuce seeds: LsCPS, LsKS, LsKO1, LsKO2, and LsKAO. Using an Escherichia coli expression system and functional assays, it is shown that LsCPS and LsKS encode ent-copalyl diphosphate synthase and ent-kaurene synthase, respectively. Using a Pichia pastoris system, it was found that LsKO1 and LsKO2 encode ent-kaurene oxidases and LsKAO encodes ent-kaurenoic acid oxidase. A comprehensive expression analysis of GA metabolism genes using the quantitative reverse transcription polymerase chain reaction suggested that transcripts of LsGA3ox1 and LsGA3ox2, both of which encode GA 3-oxidase for GA activation, were primarily expressed in the hypocotyl end of lettuce seeds, were expressed at much lower levels than the other genes tested, and were potently up-regulated by phytochrome. Furthermore, LsDELLA1 and LsDELLA2 cDNAs that encode DELLA proteins, which act as negative regulators in the GA signalling pathway, were isolated from lettuce seeds. The transcript levels of these two genes were little affected by light. Lettuce seeds in which de novo GA biosynthesis was suppressed responded almost identically to exogenously applied GA, irrespective of the light conditions, suggesting that GA responsiveness is not significantly affected by light in lettuce seeds. It is proposed that lettuce seed germination is regulated mainly via the control of the endogenous content of bioactive GA, rather than the control of GA responsiveness

Investigation of biopharmaceutical and physicochemical drug properties suitable for orally disintegrating tablets

The purpose of this study was to evaluate the biopharmaceutical drug properties suitable for orally disintegrating tablets (ODTs). The net charge at pH 7.4, log D6.5, the highest dose strength, solubility in water, dose number, and elimination t1/2 of 57 ODT drugs and 113 drugs of immediate-release (IR) formulations were compared. These drugs were classified according to the Biopharmaceutical Classification System (BCS). A lower dose strength and a longer elimination t1/2 have been observed as characteristic properties of ODTs. The proportion of basic drugs was higher in the ODTs than in the IR. A significant difference was not observed between the ODT and the IR formulation for log D6.5, solubility in water, and dose number. The distributions of the ODT and IR formulations among each BCS class were similar, suggesting that an ODT can be developed regardless of the BCS class of a drug

Phenotypic effects of the U-genome variation in nascent synthetic hexaploids derived from interspecific crosses between durum wheat and its diploid relative Aegilops umbellulata

Aegilops umbellulata is a wild diploid wheat species with the UU genome that is an important genetic resource for wheat breeding. To exploit new synthetic allohexaploid lines available as bridges for wheat breeding, a total of 26 synthetic hexaploid lines were generated through crossing between the durum wheat cultivar Langdon and 26 accessions of Ae. umbellulata. In nascent synthetic hexaploids with the AABBUU genome, the presence of the set of seven U-genome chromosomes was confirmed with U-genome chromosome-specific markers developed based on RNA-seq-derived data from Ae. umbellulata. The AABBUU synthetic hexaploids showed large variations in flowering- and morphology-related traits, and these large variations transmitted well from the parental Ae. umbellulata accessions. However, the variation ranges in most traits examined were reduced under the AABBUU hexaploid background compared with under the diploid parents. The AABBUU and AABBDD synthetic hexaploids were clearly discriminated by several morphological traits, and an increase of plant height and in the number of spikes and a decrease of spike length were commonly observed in the AABBUU synthetics. Thus, interspecific differences in several morphological traits between Ae. umbellulata and A. tauschii largely affected the basic plant architecture of the synthetic hexaploids. In conclusion, the AABBUU synthetic hexaploid lines produced in the present study are useful resources for the introgression of desirable genes from Ae. umbellulata to common wheat

Phenotypic effects of Am genomes in nascent synthetic hexaploids derived from interspecific crosses between durum and wild einkorn wheat.

Allopolyploid speciation is a major evolutionary process in wheat (Triticum spp.) and the related Aegilops species. The generation of synthetic polyploids by interspecific crosses artificially reproduces the allopolyploidization of wheat and its relatives. These synthetic polyploids allow breeders to introduce agriculturally important traits into durum and common wheat cultivars. This study aimed to evaluate the genetic and phenotypic diversity in wild einkorn Triticum monococcum ssp. aegilopoides (Link) Thell., to generate a set of synthetic hexaploid lines containing the various Am genomes from wild einkorn, and to reveal their trait characteristics. We examined the genetic diversity of 43 wild einkorn accessions using simple sequence repeat markers covering all the chromosomes and revealed two genetically divergent lineages, L1 and L2. The genetic divergence between these lineages was linked to their phenotypic divergence and their habitats. L1 accessions were characterized by early flowering, fewer spikelets, and large spikelets compared to L2 accessions. These trait differences could have resulted from adaptation to their different habitats. We then developed 42 synthetic hexaploids containing the AABBAmAm genome through interspecific crosses between T. turgidum cv. Langdon (AABB genome) as the female parent and the wild einkorn accessions (AmAm genome) as the male parents. Two of the 42 AABBAmAm synthetic hexaploids exhibited hybrid dwarfness. The phenotypic divergence between L1 and L2 accessions of wild einkorn, especially for days to flowering and spikelet-related traits, significantly reflected phenotypic differences in the synthetic hexaploids. The differences in plant height and internodes between the lineages were more distinct in the hexaploid backgrounds. Furthermore, the AABBAmAm synthetic hexaploids had longer spikelets and grains, long awns, high plant heights, soft grains, and late flowering, which are distinct from other synthetic hexaploid wheat lines such as AABBDD. Utilization of various Am genomes of wild einkorn resulted in wide phenotypic diversity in the AABBAmAm synthetic hexaploids and provides promising new breeding materials for wheat

RNA-seq analysis reveals considerable genetic diversity and provides genetic markers saturating all chromosomes in the diploid wild wheat relative Aegilops umbellulata

Abstract Background Aegilops umbellulata Zhuk. (2n = 14), a wild diploid wheat relative, has been the source of trait improvement in wheat breeding. Intraspecific genetic variation of Ae. umbellulata, however, has not been well studied and the genomic information in this species is limited. Results To develop novel genetic markers distributed over all chromosomes of Ae. umbellulata and to evaluate its genetic diversity, we performed RNA sequencing of 12 representative accessions and reconstructed transcripts by de novo assembly of reads for each accession. A large number of single nucleotide polymorphisms (SNPs) and insertions/deletions (indels) were obtained and anchored to the pseudomolecules of Ae. tauschii and barley (Hordeum vulgare L.), which were regarded as virtual chromosomes of Ae. umbellulata. Interestingly, genetic diversity in Ae. umbellulata was higher than in Ae. tauschii, despite the narrow habitat of Ae. umbellulata. Comparative analyses of nucleotide polymorphisms between Ae. umbellulata and Ae. tauschii revealed no clear lineage differentiation and existence of alleles with rarer frequencies predominantly in Ae. umbellulata, with patterns clearly distinct from those in Ae. tauschii. Conclusions The anchored SNPs, covering all chromosomes, provide sufficient genetic markers between Ae. umbellulata accessions. The alleles with rarer frequencies might be the main source of the high genetic diversity in Ae. umbellulata

Kaempherol and Luteolin Decrease Claudin-2 Expression Mediated by Inhibition of STAT3 in Lung Adenocarcinoma A549 Cells

Claudin-2 is highly expressed in human lung adenocarcinoma tissues and may be a novel target for cancer chemotherapy because knockdown of claudin-2 decreases cell proliferation. We found that flavonoids including kaempferol, chrysin, and luteolin concentration-dependently decrease claudin-2 expression in lung adenocarcinoma A549 cells. Claudin-2 expression is up-regulated by mitogen-activated protein kinase kinase (MEK)/ extracellular signal-regulated kinase (ERK)/c-Fos and phosphoinositide 3-kinase (PI3K)/Akt/nuclear factor-κB (NF-κB) pathways, but these activities were not inhibited by kaempferol, chrysin, and luteolin. Promoter deletion assay using luciferase reporter vector showed that kaempferol and luteolin inhibit the function of transcriptional factor that binds to the region between −395 and −144 of claudin-2 promoter. The decrease in promoter activity was suppressed by mutation in signal transducers and activators of transcription (STAT)-binding site, which is located between −395 and −144. The phosphorylation level of STAT3 was not decreased, but the binding of STAT3 on the promoter region is suppressed by kaempferol and luteolin in chromatin immunoprecipitation assay. The inhibition of cell proliferation caused by kaempferol and luteolin was partially recovered by ectopic claudin-2 expression. Taken together, kaempferol and luteolin decreased claudin-2 expression and proliferation in A549 cells mediated by the inhibition of binding of STAT3 on the promoter region of claudin-2. The intake of foods and nutrients rich in these flavonoids may prevent lung adenocarcinoma developmen

Efficacy and Long-Term Safety of Ibuprofen Gargle for Oral Lichen Planus: A Study Protocol of Randomized Crossover and Long-Term Extension Trials

Oral lichen planus (OLP) is a type of chronic and refractory stomatitis characterized by abnormal keratinization, which is often painful. There is no consensus regarding treatment options for OLP, particularly in the presence of pain. The current study protocol focuses on the short-term efficacy and long-term safety of an ibuprofen gargle for pain management in patients with OLP. Patients (n = 24) with painful OLP will be enrolled. During a crossover study period, patients in the ibuprofen–placebo (IP) group will receive an ibuprofen gargle (0.6%) on day 1, a placebo gargle on day 2, and an ibuprofen gargle on days 3–5 at least once daily. Patients in the placebo–ibuprofen (PI) group will receive a placebo gargle on day 1, an ibuprofen gargle on day 2, and an ibuprofen gargle on days 3–5 at least once daily. The primary endpoint of the crossover study period is the change in pain level as measured by a visual analogue scale score from before gargle administration to 5 min after gargle administration on days 1 and 2. The primary endpoint of the long-term extension study is assessment of long-term safety. The results of this study may support existing evidence regarding the effectiveness of ibuprofen rinses in treating OLP