IDENTIFICATION OF THE HIGH‐TEMPERATURE RESPONSE GENES FROM PORPHYRA SERIATA (RHODOPHYTA) EXPRESSION SEQUENCE TAGS AND ENHANCEMENT OF HEAT TOLERANCE OF CHLAMYDOMONAS (CHLOROPHYTA) BY EXPRESSION OF THE PORPHYRA HTR2 GENE1

Temperature is one of the major environmental factors that affect the distribution, growth rate, and life cycle of intertidal organisms, including red algae. In an effort to identify the genes involved in the high‐temperature tolerance of Porphyra, we generated 3,979 expression sequence tags (ESTs) from gametophyte thalli of P. seriata Kjellm. under normal growth conditions and high‐temperature conditions. A comparison of the ESTs from two cDNA libraries allowed us to identify the high temperature response (HTR) genes, which are induced or up‐regulated as the result of high‐temperature treatment. Among the HTRs, HTR2 encodes for a small polypeptide consisting of 144 amino acids, which is a noble nuclear protein. Chlamydomonas expressing the Porphyra HTR2 gene shows higher survival and growth rates than the wild‐type strain after high‐temperature treatment. These results suggest that HTR2 may be relevant to the tolerance of high‐temperature stress conditions, and this Porphyra EST data set will provide important genetic information for studies of the molecular basis of high‐temperature tolerance in marine algae, as well as in Porphyra.     

A hybrid peptide derived from cecropin-A and magainin-2 inhibits osteoclast differentiation

The adult skeleton is in a dynamic state, being continually broken down and reformed by the coordinated actions of osteoclasts and osteoblasts. Increased osteoclast activity may contribute to the development of osteoporosis. Therefore, the intervention of osteoclast-mediated bone resorption is considered as an effective therapeutic approach in the treatment of osteoporosis. In the course of searching for agents that inhibit osteoclast differentiation and activation, we found that a novel hybrid peptide P1 derived from cecropin-A and magainin-2 reduced osteoclast differentiation in various osteoclast culture systems. As this peptide had no cytotoxicity on various cultures of primary cells and established cell lines, its inhibitory effect on osteoclastogenesis was not due to general cytotoxicity. The effects of P1 on osteoclasts appear to be mediated through the inhibition of NF-kappaB and JNK activation induced by the osteoclastogenic cytokine, receptor activator of NF-kappaB ligand (RANKL). These results provide an evidence for the potential usefulness of P1 for the treatment of bone-resorbing diseases.     

A novel mechanism for the antibacterial effect of silver nanoparticles on Escherichia coli

Silver nanoparticles are known to have antimicrobial properties and have been used extensively in medicine, although the mechanism(s) of action have not yet been clearly established. In the present study, the findings suggest a novel mechanism for the antibacterial effect of silver nanoparticles on Escherichia coli, namely, the induction of a bacterial apoptosis-like response. We propose a possible mechanism for the bacterial apoptosis-like response that includes the following: accumulation of reactive oxygen species (ROS) (detected with H₂DCFDA staining), increased intracellular calcium levels (detected with Fura-2 AM), phosphatidylserine exposure in the outer membrane (detected with Annexin V) which is the hallmarks of early apoptosis, disruption of the membrane potential [detected with DiBAC₄(3)], activation of a bacterial caspase-like protein (detected by FITC-VAD-FMK staining) and DNA degradation (detected with TUNEL assay) which is the hallmarks of late apoptosis in bacterial cells treated with silver nanoparticles. We also performed RecA expression assay with western blotting and observed activation of SOS response to repair the damaged DNA. To summarize, silver nanoparticles are involved in the apoptosis-like response in E. coli and the novel mechanisms which were identified in this study, suggest that silver nanoparticles may be an effective antimicrobial agent with far lower propensity for inducing microbial resistance than antibiotics.     

Life history differences of Psilotreta locumtenens (Trichoptera: Odontoceridae) in two reaches of a mountain stream in Korea

The life history traits of the caddisfly, Psilotreta locumtenens Botosaneanu (Odontoceridae), were studied in two stream reaches with different thermal ranges (main and branch streams) of the Gapyeong Stream, a typical mountain stream located in the central Korean Peninsula. Psilotreta locumtenens larvae were quantitatively sampled monthly from November 2008 to July 2010, and biweekly during the emergence period (late April to early July), using a Surber sampler (30 × 30 cm). Adults were quantitatively sampled with a sweep net. Larval density in the main stream (324.21 ± 38.59 m^?2) was higher than that in the branch stream (60.48 ± 10.86 m^?2). The larvae hatched in the early summer and overwintered as 5th and 3rd instars in the main and branch streams, respectively. The emergence peak at the main stream was approximately 2 weeks earlier. The sex ratio at both sites was approximately 0.3. The life history in both streams was univoltine. Secondary production in the main stream was much higher than in the branch stream, owing to high larval densities, and the P/B ratios at the two sites were similar. This study demonstrated remarkable differences in larval growth patterns and emergence peaks in P. locumtenens between the two stream reaches due to differences in accumulated degree days (230.30 DD) and other phenological cues such as daily mean threshold water temperature (9°C) during the ascending phase, despite their relatively small mean annual water temperature difference of 0.58°C.     

Validation of a chloroquine-induced cell death mechanism for clinical use against malaria

An alternative antimalarial pathway of an ‘outdated'' drug, chloroquine (CQ), may facilitate its return to the shrinking list of effective antimalarials. Conventionally, CQ is believed to interfere with hemozoin formation at nanomolar concentrations, but resistant parasites are able to efflux this drug from the digestive vacuole (DV). However, we show that the DV membrane of both resistant and sensitive laboratory and field parasites is compromised after exposure to micromolar concentrations of CQ, leading to an extrusion of DV proteases. Furthermore, only a short period of exposure is required to compromise the viability of late-stage parasites. To study the feasibility of this strategy, mice malaria models were used to demonstrate that high doses of CQ also triggered DV permeabilization in vivo and reduced reinvasion efficiency. We suggest that a time-release oral formulation of CQ may sustain elevated blood CQ levels sufficiently to clear even CQ-resistant parasites.Along with improvements in vector control, surveillance/diagnosis and treatment accessibility, the development of new drugs to counteract the problem of drug resistance remains integral to the eradication agenda.¹ Efforts to develop novel antimalarials have been promising,^{2, 3} and drugs designed specifically to reverse drug resistance are also being uncovered.⁴ However, novel chemical entities are expensive to test and take considerable time before they can be deployed. In comparison, alternative strategies to fully exploit the existing arsenal of antimalarials (largely already affordable and accessible) are likely to be relatively expedient and cost-effective.We had previously demonstrated the existence of a novel parasite programmed cell death (PCD) mechanism that was induced by high concentrations of chloroquine (CQ) and shown that clan CA cysteine proteases were key mediators of the pathway.⁵ We had also observed that the permeabilization of the parasite digestive vacuole (DV) was an important upstream trigger of this pathway and that other lysosomotropic compounds that are not parasite-specific could similarly destabilize the DV to initiate parasite PCD.⁶ We hypothesize that by altering the dosing regimen or formulation of CQ, it might be possible to reinstate CQ into antimalarial chemotherapy by making use of this novel mechanism.⁷In this present study, we begin by showing evidence that CQ treatment is able to result in the extrusion of DV proteases into the parasite cytoplasm. Second, we validate the existence of this PCD pathway in multiple laboratory strains and field isolates to suggest its clinical relevance and universality. Third, we investigate the minimum concentration and duration required for CQ to trigger PCD to determine if the pharmacokinetics of the current CQ regimen might be suitable for initiating PCD. Finally, we make use of two murine malaria models to demonstrate that a short exposure to high levels of CQ is able to induce parasite DV permeabilization in vivo and that this procedure reduces parasite viability.     

Effects of elevated carbon dioxide, ozone and water availability on spring wheat growth and yield

Spring wheat (Triticum aestivum L. cv. Dragon) was exposed to elevated carbon dioxide (CO₂), alone (1995) or in combination with two levels of increased ozone (O₃) (1994) or increased irrigation (1996) during three successive growing seasons as part of the EU ESPACE‐wheat programme and conducted in open‐top chambers (OTCs) and ambient air (AA) plots at Östad, 50 km north‐east of Göteborg, Sweden. Doubling the CO₂ concentration had a positive effect on grain yield in all 3 years (+21, +7 and +11%, respectively), although only statistically significant in 1994. That year was characterised by a warm and dry summer in comparison with 1995 and 1996, in which the summers were more humid and typical for south‐west Sweden. In 1994, the CO₂‐induced increase in grain yield was associated with an increase in the duration of the green leaf area, a positive effect on straw yield and on the number of ears per square metre and a negative effect (?13%) on grain protein concentration. Harvest index was unaffected by the elevated CO₂ concentration. The only statistically significant effect of elevated CO₂ in 1995 was a decrease in the grain protein concentration (?11% in both CO₂ concentrations), and in 1996 an increase (+21%) in the straw yield. In 1996 the soil water potential was less negative in elevated CO₂, which is likely to reflect a lower water consumption of these plants. Addition of extra O₃ significantly affected the grain yield (?6 and ?10%, respectively) and the 1 000‐grain weight negatively (?3 and ?6%). Statistically significant interactions between CO₂ and O₃ were obtained for the number of ears per unit area and for the 1 000‐grain weight. The 1 000‐grain weight was negatively affected by O₃ in low CO₂, but remained unaffected in the high CO₂ treatment. There was a significant decrease (?6%) in the grain protein concentration induced by elevated irrigation. The chambers, compared with AA plots, had a positive effect on plant development and on grain yield in all 3 years.     

A bacterial effector counteracts host autophagy by promoting degradation of an autophagy component

Beyond its role in cellular homeostasis, autophagy plays anti‐ and promicrobial roles in host–microbe interactions, both in animals and plants. One prominent role of antimicrobial autophagy is to degrade intracellular pathogens or microbial molecules, in a process termed xenophagy. Consequently, microbes evolved mechanisms to hijack or modulate autophagy to escape elimination. Although well‐described in animals, the extent to which xenophagy contributes to plant–bacteria interactions remains unknown. Here, we provide evidence that Xanthomonas campestris pv. vesicatoria (Xcv) suppresses host autophagy by utilizing type‐III effector XopL. XopL interacts with and degrades the autophagy component SH3P2 via its E3 ligase activity to promote infection. Intriguingly, XopL is targeted for degradation by defense‐related selective autophagy mediated by NBR1/Joka2, revealing a complex antagonistic interplay between XopL and the host autophagy machinery. Our results implicate plant antimicrobial autophagy in the depletion of a bacterial virulence factor and unravel an unprecedented pathogen strategy to counteract defense‐related autophagy in plant–bacteria interactions.