Adaptation to the new land or effect of global warming? An age-structured model for rapid voltinism change in an alien lepidopteran pest

1. Hyphantria cunea Drury invaded Japan at Tokyo in 1945 and expanded its distribution gradually into northern and south-western Japan. All populations in Japan were bivoltine until the early 1970s, at which time trivoltine populations appeared in several southern regions. Presently, H. cunea exists as separate bivoltine and trivoltine populations divided around latitude 36 degrees . In the course of this voltinism change, the mean surface temperature in Japan rose by 1.0 degrees C. 2. To determine whether and how this temperature increase might be responsible for the voltinism change, we constructed an age-structured model incorporating growth speed driven by actual daily temperature and detailed mechanisms of diapause induction triggered by both daily photoperiod and temperature. 3. The simulation result suggests that both the acceleration of the growth speed and the prolongation of diapause induction are necessary to cause changes in voltinism, regardless of temperature increase. We concluded that the H. cunea population changed its life-history traits as an adaptation parallel with its invasion into the south-western parts of Japan. 4. Though the temperature increase had little effect on the fitness and heat stress in bivoltine and trivoltine populations, the trivoltine life cycle has become advantageous at least in marginal regions such as Tokyo.     

Improved cellular activity of antisense peptide nucleic acids by conjugation to a cationic peptide-lipid (CatLip) domain

Conjugation to cationic cell penetrating peptides (such as Tat, Penetratin, or oligo arginines) efficiently improves the cellular uptake of large hydrophilic molecules such as oligonucleotides and peptide nucleic acids, but the cellular uptake is predominantly via an unproductive endosomal pathway and therefore mechanisms that promote endosomal escape (or avoid the endosomal route) are required for improving bioavailability. A variety of auxiliary agents (chloroquine, calcium ions, or lipophilic photosensitizers) has this effect, but improved, unaided delivery would be highly advantageous in particular for future in vivo applications. We find that simply conjugating a lipid domain (fatty acid) to the cationic peptide (a CatLip conjugate) increases the biological effect of the corresponding PNA (CatLip) conjugates in a luciferase cellular antisense assay up to 2 orders of magnitude. The effect increases with increasing length of the fatty acid (C8-C16) but in parallel also results in increased cellular toxicity, with decanoic acid being optimal. Furthermore, the relative enhancement is significantly higher for Tat peptide compared to oligoarginine. Confocal microscopy and chloroquine enhancement indicates that the lipophilic domain increases the endosomal uptake as well as promoting significantly endosomal escape. These results provide a novel route for improving the (cellular) bioavailability of larger hydrophilic molecules.     

Differential effects of low- and high-dose X-rays on N-ethyl-N-nitrosourea-induced mutagenesis in thymocytes of B6C3F1 gpt-delta mice

Carcinogenesis in humans is thought to result from exposure to numerous environmental factors. Little is known, however, about how these different factors work in combination to cause cancer. Because thymic lymphoma is a good model of research for combined exposure, we examined the occurrence of mutations in thymic DNA following exposure of B6C3F1 gpt-delta mice to both ionizing radiation and N-ethyl-N-nitrosourea (ENU). Mice were exposed weekly to whole body X-irradiation (0.2 or 1.0 Gy), ENU (200 ppm) in the drinking water, or X-irradiation followed by ENU treatment. Thereafter, genomic DNA was prepared from the thymus and the number and types of mutations in the reporter transgene gpt was determined. ENU exposure alone increased mutant frequency by 10-fold compared to untreated controls and over 80% of mutants had expanded clonally. X-irradiation alone, at either low or high dose, unexpectedly, reduced mutant frequency. Combined exposure to 0.2 Gy X-rays with ENU dramatically decreased mutant frequency, specifically G:C to A:T and A:T to T:A mutations, compared to ENU treatment alone. In contrast, 1.0 Gy X-rays enhanced mutant frequency by about 30-fold and appeared to accelerate clonal expansion of mutated cells. In conclusion, repeated irradiation with 0.2 Gy X-rays not only reduced background mutation levels, but also suppressed ENU-induced mutations and clonal expansion. In contrast, 1.0 Gy irradiation in combination with ENU accelerated clonal expansion of mutated cells. These results indicate that the mode of the combined mutagenic effect is dose dependent.     

Effects of theanine, a unique amino acid in tea leaves, on memory in a rat behavioral test

We identified an effect of theanine on memory functions in a novel object test. Rats were fed theanine for 3 weeks ad libitum, and then they performed the object test. The theanine-fed group performed search behavior for the novel object in the test session. The results suggest that theanine-fed rats showed improved recognition, and that theanine affected learning and memory.     

In situ detection of methylated DNA by histo endonuclease-linked detection of methylated DNA sites: a new principle of analysis of DNA methylation

For a better understanding of epigenetic regulation of cell differentiation, it is important to analyze DNA methylation at a specific site. Although previous studies described methylation of isolated DNA extracted from cells and tissues using a combination of appropriate restriction endonucleases, no application to tissue cell level has been reported. Here, we report a new method, named histo endonuclease-linked detection of methylation sites of DNA (HELMET), designed to detect methylation sites of DNA with a specific sequences in a tissue section. In this study, we examined changes in the methylation level of CCGG sites during spermatogenesis in paraffin-embedded sections of mouse testis. In principle, the 3′-OH ends of DNA strand breaks in a section were firstly labeled with a mixture of dideoxynucleotides by terminal deoxynucleotidyl transferase (TdT), not to be further elongated by TdT. Then the section was digested with Hpa II, resulting in cutting the center portion of non-methylated CCGG. The cutting sites were labeled with biotin-16-dUTP by TdT. Next, the section was treated with Msp I, which can cut the CCGG sequence irrespective of the presence or absence of methylation of the second cytosine, and the cutting sites were labeled with digoxigenin-11-dUTP by TdT. Finally, both biotin and digoxigenin were visualized by enzyme- or fluorescence-immunohistochemistry. Using this method, we found hypermethylation of CCGG sites in most of the germ cells although non-methylated CCGG were colocalized in elongated spermatids. Interestingly, some TUNEL-positive germ cells, which are frequent in mammalian spermatogenesis, became markedly Hpa II-reactive, indicating that the CCGG sites may be demethylated during apoptosis. An erratum to this article can be found at     

Affinity purification and characterization of a key enzyme responsible for circadian rhythmic control of nyctinasty in Lespedeza cuneata L

The synthesis of an affinity gel aimed at leaf-opening factor beta-glucosidase (LOFG) and affinity purification of LOFG is presented. A gluconamidine-based beta-glucosidase inhibitor was used as the ligand of the affinity gel. beta-Glucosidase exhibiting an activity shift throughout the day was selectively purified from Lespedeza cuneata Don by the affinity gel. The resulting LOFG exhibited high substrate specificity toward the leaf-opening factor.     

Regulation of clathrin-dependent endocytosis by diacylglycerol kinase delta: importance of kinase activity and binding to AP2alpha

DGKdelta (diacylglycerol kinase delta), which phosphorylates DAG (diacylglycerol) and converts it into PA (phosphatidic acid), has an important role in signal transduction. In the present study, we have demonstrated the molecular mechanism of DGKdelta-mediated regulation of clathrin-dependent endocytosis that controls the internalization, recycling and degradation of receptors. Involvement of DGKdelta in the regulation of clathrin-dependent endocytosis was previously proposed following genome-wide RNAi (RNA interference) screening. Clathrin-coated pits are mainly formed by clathrin and AP-2 (adaptor protein 2) complex. These proteins assemble a polyhedral lattice at the membrane and gather several endocytic accessory proteins. As the intracellular localization of DGKdelta2 overlapped with clathrin-coated pits, we predicted the possible regulation of clathrin-dependent endocytosis by DGKdelta2 and its interaction with some endocytosis-regulatory proteins. DGKdelta2 contained the DXF-type binding motifs, and DGKdelta2 bound to AP2alpha, a subunit of the AP-2 complex. DGKdelta2 interacted with the platform subdomain in the AP2alpha ear domain via F369DTFRIL and D746PF sequences in the catalytic domain of DGKdelta2. For further insight into the role for DGKdelta2 in clathrin-dependent endocytosis, we measured the transferrin and EGF (epidermal growth factor) uptake-expressing wild-type or mutant DGKdelta2 under knockdown of endogenous DGKdelta. Mutants lacking binding ability to AP2alpha as well as kinase-negative mutants could not compensate for the uptake of transferrin inhibited by siRNA (small interfering RNA) treatment, whereas overexpression of wild-type DGKdelta2 completely recovered the transferrin uptake. These results demonstrate that binding between DGKdelta2 and AP2alpha is involved in the transferrin internalization and that DGK activity is also necessary for the regulation of the endocytic process.