Plant gene responses to frequency-specific sound signals

We identified a set of sound-responsive genes in plants using a sound-treated subtractive library and demonstrated sound regulation through mRNA expression analyses. Under both light and dark conditions, sound up-regulated expression of rbcS and ald. These are also light-responsive genes and these results suggest that sound could represent an alternative to light as a gene regulator. Ald mRNA expression increased significantly with treatment at 125 and 250 Hz, whereas levels decreased significantly with treatment at 50 Hz, indicating a frequency-specific response. To investigate whether the ald promoter responds to sound, we generated transgenic rice plants harboring a chimeric gene comprising a fusion of the ald promoter and GUS reporter. In three independent transgenic lines treated with 50 or 250 Hz for 4 h, GUS mRNA expression was up-regulated at 250 Hz, but down-regulated at 50 Hz. Thus, the sound-responsive mRNA expression pattern observed for the ald promoter correlated closely with that of ald, suggesting that the 1,506 bp ald promoter is sound-responsive. Therefore, we propose that in transgenic plants, specific frequencies of sound treatment could be used to regulate the expression of any gene fused to the ald promoter.     

Genus delimitation,biogeography and diversification of Choristoneura Lederer (Lepidoptera: Tortricidae) based on molecular evidence

Widely known for pest species that include major modulators of temperate forests, the genus Choristoneura is part of the species‐rich tribe Archipini of leafroller moths (Tortricidae). Delimitation of the genus has remained unresolved because no phylogeny has included species endemic to Africa and studies have often omitted the type species of the genus. Further taxonomic confusion has been generated by the transfer of Archips occidentalis (Walsingham) to Choristoneura, creating a homonym with Choristoneura occidentalis Freeman, an important defoliator of North American forests. To define the limits of the genus, we reconstructed a phylogeny using DNA sequences for mitochondrial cytochrome oxidase subunit I and nuclear ribosomal 28S genes. Our ingroup included 23 Choristoneura species‐level taxa, complemented by a large sample of outgroups comprising 82 species of Archipini and other Tortricidae. We generated a time‐calibrated tree using fossil and secondary calibrations and we inferred biogeographic and diversification processes in Choristoneura. Our analysis recovered the genus as polyphyletic, with Archips occidentalis, Choristoneura simonyi and Choristoneura evanidana excluded from the main clade. Based on the recovered phylogenies and a redefinition, we restrict Choristoneura primarily to species with a northern hemisphere distribution. Our analysis supports A. occidentalis as the sister group of Cacoecimorpha pronubana, C. simonyi as the sister of ‘Xenotemna’ pallorana, and C. evanidana as the sister of Archips purpurana. A new combination is proposed: Archips evanidana comb.n. ; the availability of ‘Xenotemna’ as a valid name is discussed and A. occidentalis is considered as an orphaned name within the Archipini. We found support for a Holarctic origin of Choristoneura about 23 Ma, followed by early divergence in the Palearctic region. The main divergence occurred at 16 Ma, with one clade in the Nearctic and another in the Palearctic. Subsequent cladogenetic events were synchronous and related to herbivorous specialization, with each clade divided into coniferophagous and polyphagous lineages. Their specialization as conifer feeders temporally matched the expansion of boreal forest during the Miocene.     

Cloning of a ribonucleotide reductase gene of the herpes simplex virus type 2 strain G

The ribonucleotide reductase (RR) 2 gene of the HSV-2 strain G was cloned, sequenced, and expressed in an E. coli cell. The RR2 gene was located on the PstI 2.4 kb fragment, which was cloned and sequenced. The ORF of the gene was 1,011 bp and its termination codon was TAG; also, the CATATAA sequence was present in the promoter of the RR2 gene. A Poly A signal sequence (AATAAA) was found in the 3'-noncoding region. The RR2 proteins that were produced in the E. coli and Vero cells were confirmed using a Western blot analysis. SDS-PAGE revealed that the molecular weights of the fusion-RR2 that was produced in the E. coli cells were approximately 24 kDa and 38 kDa in the Vero cells. The RR2 proteins were soluble. The differences in the molecular weights might be due to modifications in the Vero cells.     

The involvement of histone methylation in osteoblastic differentiation of human periosteum‐derived cells cultured in vitro under hypoxic conditions

Although oxygen concentrations affect the growth and function of mesenchymal stem cells (MSCs), the impact of hypoxia on osteoblastic differentiation is not understood. Likewise, the effect of hypoxia‐induced epigenetic changes on osteoblastic differentiation of MSCs is unknown. The aim of this study was to examine the in vitro hypoxic response of human periosteum‐derived cells (hPDCs). Hypoxia resulted in greater proliferation of hPDCs as compared with those cultured in normoxia. Further, hypoxic conditions yielded decreased expression of apoptosis‐ and senescence‐associated genes by hPDCs. Osteoblast phenotypes of hPDCS were suppressed by hypoxia, as suggested by alkaline phosphatase activity, alizarin red‐S‐positive mineralization, and mRNA expression of osteoblast‐related genes. Chromatin immunoprecipitation assays showed an increased presence of H3K27me3, trimethylation of lysine 27 on histone H3, on the promoter region of bone morphogenetic protein‐2. In addition, mRNA expression of histone lysine demethylase 6B (KDM6B) by hPDCs was significantly decreased in hypoxic conditions. Our results suggest that an increased level of H3K27me3 on the promoter region of bone morphogenetic protein‐2, in combination with downregulation of KDM6B activity, is involved in the suppression of osteogenic phenotypes of hPDCs cultured in hypoxic conditions. Although oxygen tension plays an important role in the viability and maintenance of MSCs in an undifferentiated state, the effect of hypoxia on osteoblastic differentiation of MSCs remains controversial. In addition, evidence regarding the importance of epigenetics in regulating MSCs has been limited. This study was to examine the role hypoxia on osteoblastic differentiation of hPDCs, and we examined whether histone methylation is involved in the observed effect of hypoxia on osteogenic differentiation of hPDCs.     

The Use of Exome Genotyping to Predict Pathological Gleason Score Upgrade after Radical Prostatectomy in Low-Risk Prostate Cancer Patients

Background

Active surveillance (AS) is a promising option for patients with low-risk prostate cancer (PCa), however current criteria could not select the patients correctly, many patients who fulfilled recent AS criteria experienced pathological Gleason score upgrade (PGU) after radical prostatectomy (RP). In this study, we aimed to develop an accurate model for predicting PGU among low-risk PCa patients by using exome genotyping.

Methods

We genotyped 242,221 single nucleotide polymorphisms (SNP)s on a custom HumanExome BeadChip v1.0 (Illuminam Inc.) in blood DNA from 257 low risk PCa patients (PSA <10 ng/ml, biopsy Gleason score (GS) ≤6 and clinical stage ≤T2a) who underwent radical prostatectomy. Genetic data were analyzed using an unconditional logistic regression to calculate an odds ratio as an estimate of relative risk of PGU, which defined pathologic GS above 7. Among them, we selected persistent SNPs after multiple testing using FDR method, and we compared accuracies from the multivariate logistic model incorporating clinical factors between included and excluded selected SNP information.

Results

After analysis of exome genotyping, 15 SNPs were significant to predict PGU in low risk PCa patients. Among them, one SNP – rs33999879 remained significant after multiple testing. When a multivariate model incorporating factors in Epstein definition – PSA density, biopsy GS, positive core number, tumor per core ratio and age was devised for the prediction of PGU, the predictive accuracy of the multivariate model was 78.4% (95%CI: 0.726–0.834). By addition the factor of rs33999879 in aforementioned multivariate model, the predictive accuracy was 82.9%, which was significantly increased (p = 0.0196).

Conclusion

The rs33999879 SNP is a predictor for PGU. The addition of genetic information from the exome sequencing effectively enhanced the predictive accuracy of the multivariate model to establish suitable active surveillance criteria.     

Controlling plant growth via the gibberellin biosynthesis system – I. Growth parameter alterations in apple seedlings

'York Imperial' apple seedlings ( Malus domestica Borkh.) were continuously supplied via the roots with paclobutrazol [(2RS, 3RS)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)pentan-3-ol)], a triazole GA biosynthesis inhibitor, at 0.68 μ M in a nutrient solution. In comparison to controls, seedlings treated with paclobutrazol for 66 days showed a 91% reduction in shoot length, a 66% reduction in leaf area but only a 17% reduction in leaf number. This effect could be reversed by GA₃ applied to the foliage at 71.4 μ M 0, 19 or 35 days after paclobutrazol was initially supplied and leaf area values for paclobutrazol-treated seedlings given both treatments did not differ significantly from controls. Plots of growth data indicate linearity of shoot longitudinal growth of GA₃-treated seedlings. Leaf area increase was non-linear after GA₃ treatment up to approximately 30 days, when the rate dropped. On a per shoot basis, leaf weight closely followed leaf area but on a per unit area basis, paclobutrazol-treated leaves were heavier than controls; GA₃ applications temporarily reversed this trend.     

Carba analogs of cyclic phosphatidic acid are selective inhibitors of autotaxin and cancer cell invasion and metastasis

Autotaxin (ATX, nucleotide pyrophosphate/phosphodiesterase-2) is an autocrine motility factor initially characterized from A2058 melanoma cell-conditioned medium. ATX is known to contribute to cancer cell survival, growth, and invasion. Recently ATX was shown to be responsible for the lysophospholipase D activity that generates lysophosphatidic acid (LPA). Production of LPA is sufficient to explain the effects of ATX on tumor cells. Cyclic phosphatidic acid (cPA) is a naturally occurring analog of LPA in which the sn-2 hydroxy group forms a 5-membered ring with the sn-3 phosphate. Cellular responses to cPA generally oppose those of LPA despite activation of apparently overlapping receptor populations, suggesting that cPA also activates cellular targets distinct from LPA receptors. cPA has previously been shown to inhibit tumor cell invasion in vitro and cancer cell metastasis in vivo. However, the mechanism governing this effect remains unresolved. Here we show that 3-carba analogs of cPA lack significant agonist activity at LPA receptors yet are potent inhibitors of ATX activity, LPA production, and A2058 melanoma cell invasion in vitro and B16F10 melanoma cell metastasis in vivo.