Characterization of a rice mutant having an increased susceptibility to light stress at high temperature

The effect of light irradiation at high temperature was investigated in a rice mutant, spl-2 , which is sensitive to solar radiation. Dead spots appeared on the mutant leaves when cultured at a high temperature (40°C) under strong white-light illumination (15 W m⁻²). A similar damage was also observed in the wild-type leaves under the same conditions when the plants were preincubated in the dark for one day. Preillumination with weak light (6 W m⁻²) lessened the irradiation-induced damage in the wild-type. These observations suggest that in rice plants the acclimatization to weak light has a protective effect against strong irradiation at high temperature, and the spl-2 locus participates in the mechanisms of the acclimatization. The action spectrum for the irradiation-induced damage at the high temperature (40°C) in the spl-2 leaves indicated that the maximum damage occurred at around 480 nm and, in a lesser extent, at around 680 nm. Activity of the O⁻₂₋ and H₂O₂₋scavenging enzymes in the spl−2 leaves were almost the same or somewhat higher than those in the wild-type after irradiation with strong white light (15 W m⁻²) at 40°C, whereas the content of ascorbic acid in the spl−2 decreased significantly compared with the wild type.     

Characterization of the interaction of Rhodobacter capsulatus cytochrome c peroxidase with charge reversal mutants of cytochrome c(2)

Steady-state kinetics for the reaction of Rhodobacter capsulatus bacterial cytochrome c peroxidase (BCCP) with its substrate cytochrome c(2) were investigated. The Rb. capsulatus BCCP is dependent on calcium for activation as previously shown for the Pseudomonas aeruginosa BCCP and Paracoccus denitrificans enzymes. Furthermore, the activity shows a bell-shaped pH dependence with optimum at pH 7.0. Enzyme activity is greatest at low ionic strength and drops off steeply as ionic strength increases, resulting in an apparent interaction domain charge product of -13. All cytochromes c(2) show an asymmetric distribution of surface charge, with a concentration of 14 positive charges near the exposed heme edge of Rb. capsulatus c(2) which potentially may interact with approximately 6 negative charges, localized near the edge of the high-potential heme of the Rb. capsulatus BCCP. To test this proposal, we constructed charge reversal mutants of the 14 positively charged residues located on the front face of Rb. capsulatus cytochrome c(2) and examined their effect on steady-state kinetics with BCCP. Mutated residues in Rb. capsulatus cytochrome c(2) that showed the greatest effects on binding and enzyme activity are K12E, K14E, K54E, K84E, K93E, and K99E, which is consistent with the site of electron transfer being located at the heme edge. We conclude that a combination of long-range, nonspecific electrostatic interactions as well as localized salt bridges between, e.g., cytochrome c(2) K12, K14, K54, and K99 with BCCP D194, D241, and D6, account for the observed kinetics.     

Growth and progression of melanoma and non-melanoma skin cancers regulated by ubiquitination

Basal cell carcinomas (BCC), squamous cell carcinoma (SCC), and melanomas are the major types of skin tumors. Despite being skin cancers, the characteristics of each cancer are widely varied. BCCs often do not proliferate rapidly, and rarely metastasize. Squamous cell carcinomas are more malignant and a certain subtype of SCC is highly metastatic. Melanomas are highly proliferative and invasive, and are most frequently metastatic. Ubiquitin and ubiquitin-related proteins post-translationally modify proteins and thereby alter the functions of their target proteins. The ubiquitination process is involved in various physiological responses, including cell growth, cell death, and DNA damage repair. Accumulating evidence suggests that ubiquitin pathways are involved in different types of cancers, including skin cancers. This review describes the major ubiquitin pathways in BCC, SCC, and melanoma. The ubiquitin pathways that are activated among the skin cancers are highly diverse, which might reflect the various characteristics of these three cancer types. Meanwhile, there are also common pathways between BCC, SCC, and melanoma. Therefore, examining the ubiquitin pathways will reveal the mechanisms of these three major skin cancer types and will suggest treatment options.     

Breakdown of distyly in a tetraploid variety of <Emphasis Type="Italic">Ophiorrhiza</Emphasis><Emphasis Type="Italic"> japonica</Emphasis> (Rubiaceae) and its phylogenetic analysis

We examined the floral morph of tetraploid Ophiorrhiza japonica Blume var. amamiana Hatus. and diploid O. japonica var. japonica to elucidate the association of distyly and ploidy levels. Chloroplast DNA phylogeny was reconstructed to determine the number of tetraploidization events and floral morph shifts in O. japonica. All individuals of O. japonica var. amamiana proved to be long-homostylous, whereas O. japonica var. japonica was distylous with typical long- and short-styled flowers. Distyly is related to the ploidy level. The bagging treatment of flowers indicated that O. japonica var. amamiana is self-compatible and potentially automatically self-pollinating. In cpDNA sequencing analysis, no haplotype was shared between the two varieties. The cpDNA haplotype network displayed the monophyly of O. japonica var. amamiana, suggesting a single origin of this variety. Hence, both tetraploidization and the breakdown of distyly to homostyly in O. japonica var. amamiana likely occurred just once. Because O. japonica var. amamiana having the morphological and cytological entity is recognized as a single lineage and clearly separated from O. japonica var. japonica, this variety can be considered to be a distinct species. We therefore propose to raise O. japonica var. amamiana to the rank of species. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Upper temperature limits of tropical marine ectotherms: global warming implications

Animal physiology, ecology and evolution are affected by temperature and it is expected that community structure will be strongly influenced by global warming. This is particularly relevant in the tropics, where organisms are already living close to their upper temperature limits and hence are highly vulnerable to rising temperature. Here we present data on upper temperature limits of 34 tropical marine ectotherm species from seven phyla living in intertidal and subtidal habitats. Short term thermal tolerances and vertical distributions were correlated, i.e., upper shore animals have higher thermal tolerance than lower shore and subtidal animals; however, animals, despite their respective tidal height, were susceptible to the same temperature in the long term. When temperatures were raised by 1°C hour(-1), the upper lethal temperature range of intertidal ectotherms was 41-52°C, but this range was narrower and reduced to 37-41°C in subtidal animals. The rate of temperature change, however, affected intertidal and subtidal animals differently. In chronic heating experiments when temperature was raised weekly or monthly instead of every hour, upper temperature limits of subtidal species decreased from 40°C to 35.4°C, while the decrease was more than 10°C in high shore organisms. Hence in the long term, activity and survival of tropical marine organisms could be compromised just 2-3°C above present seawater temperatures. Differences between animals from environments that experience different levels of temperature variability suggest that the physiological mechanisms underlying thermal sensitivity may vary at different rates of warming.     

Arabidopsis thaliana subcellular responses to compatible Erysiphe cichoracearum infections

Subcellular events of Erysiphe cichoracearum infections of epidermal cells were visualized in living tissues of Arabidopsis plants carrying various green fluorescent protein (GFP)-tagged organelles via laser scanning confocal microscopy. Early in the infection sequence, cytoplasm and organelles moved towards penetration sites and accumulated near penetration pegs. Peroxisomes appeared to accumulate preferentially relative to the cytoplasm at penetration sites. Another early event, which preceded haustorium formation, was the aggregation of some GFP-tagged plasma membrane marker proteins into rings around penetration sites, which extended across cell-wall boundaries into neighboring cells. This feature localized to sites where papillae were deposited. The extrahaustorial membrane (EHM) encases the fungal feeding structure, the haustorium, separating it from the host cytoplasm. Eight plasma membrane markers were excluded from the EHM and remained in a collar-like formation around the haustorial neck. These observations support the suggestions that the EHM is a unique, specialized membrane and is different from the plasma membrane. Our results suggested two possibilities for the origin of the EHM: invagination of the plasma membrane coupled with membrane differentiation; or de novo synthesis of the EHM by targeted vesicle trafficking.     

Is a genetic defect in Fkbp6 a common cause of azoospermia in humans?

FK506-binding protein 6 (Fkbp6) is a member of a gene family containing a prolyl isomerase/FK506-binding domain and tetratricopeptide protein-protein interaction domains. Recently, the targeted inactivation of Fkbp6 in mice has been observed to result in aspermic males and the absence of normal pachytene spermatocytes. The loss of Fkbp6 results in abnormal pairing and a misalignment of the homologous chromosomes, and in non-homologous partner switches and autosynapsis of the X chromosome cores in meiotic spermatocytes. In this study, we analyzed whether human FKBP6 gene defects might be associated with human azoospermia. We performed a mutation analysis in all the coding regions of the human FKBP6 gene in 19 patients with azoospermia resulting from meiotic arrest. The expression of the human FKBP6 gene was specific to the testis, and a novel polymorphism site, 245C → G (Y60X) could be found in exon 3. Our findings suggest that the human FKBP6 gene might be imprinted in the testis based on an analysis using two polymorphism sites. These authors equally contributed to this paper     

p53 inhibits adriamycin-induced down-regulation of cyclin D1 expression in human cancer cells.

The tumor suppressor p53 gene product is an essential component of the cytotoxic pathway triggered by DNA-damaging stimuli such as chemotherapeutic agents and ionizing radiation. We previously demonstrated that adenovirus-mediated wild-type p53 gene transfer could enhance the cytotoxic actions of chemotherapeutic drugs both in vitro and in vivo; however, the molecular mechanism of this chemosensitization is still unclear. Cyclin D1 is a major regulator of the progression of cells into the proliferative stage of the cell cycle. Here we show that infection with an adenovirus vector expressing the wild-type p53 gene (Ad-p53) caused an increase in cyclin D1 protein levels in human colorectal cancer cell lines DLD-1 and SW620; treatment with the anti-cancer drug adriamycin, however, down-regulated their cyclin D1 protein expression in a dose-dependent manner. The suppression of cyclin D1 expression following adriamycin treatment could be blocked by simultaneous Ad-p53 infection. Furthermore, DLD-1 and SW620 cells transfected with the cyclin D1 expression construct displayed increased sensitivity to adriamycin compared to that of the vector-transfected control. Our results suggest that ectopic wild-type p53 gene transfer results in increased cyclin D1 expression and, consequently, sensitizes human colorectal cancer cells to chemotherapeutic agents.     

Characterizing Heterogeneity within Head and Neck Lesions Using Cluster Analysis of Multi-Parametric MRI Data

Purpose

To describe a methodology, based on cluster analysis, to partition multi-parametric functional imaging data into groups (or clusters) of similar functional characteristics, with the aim of characterizing functional heterogeneity within head and neck tumour volumes. To evaluate the performance of the proposed approach on a set of longitudinal MRI data, analysing the evolution of the obtained sub-sets with treatment.

Material and Methods

The cluster analysis workflow was applied to a combination of dynamic contrast-enhanced and diffusion-weighted imaging MRI data from a cohort of squamous cell carcinoma of the head and neck patients. Cumulative distributions of voxels, containing pre and post-treatment data and including both primary tumours and lymph nodes, were partitioned into k clusters (k = 2, 3 or 4). Principal component analysis and cluster validation were employed to investigate data composition and to independently determine the optimal number of clusters. The evolution of the resulting sub-regions with induction chemotherapy treatment was assessed relative to the number of clusters.

Results

The clustering algorithm was able to separate clusters which significantly reduced in voxel number following induction chemotherapy from clusters with a non-significant reduction. Partitioning with the optimal number of clusters (k = 4), determined with cluster validation, produced the best separation between reducing and non-reducing clusters.

Conclusion

The proposed methodology was able to identify tumour sub-regions with distinct functional properties, independently separating clusters which were affected differently by treatment. This work demonstrates that unsupervised cluster analysis, with no prior knowledge of the data, can be employed to provide a multi-parametric characterization of functional heterogeneity within tumour volumes.     

Expression Patterns of miRNA-423-5p in the Serum and Pericardial Fluid in Patients Undergoing Cardiac Surgery

Background

Recently, it has been reported that specific microRNA (miRNA) levels are elevated in serum and can be used as biomarkers in patients with cardiovascular diseases. However, miRNAs expression profiles and their sources in pericardial fluid (PF) are unclear.

Methods and Results

The purpose of this study was to identify the levels of miRNAs in PF in relation to those in the serum in patients undergoing cardiac surgery. Serum (S) and PF from patients undergoing coronary artery bypass graft (CABG) due to stable angina pectoris (sAP) and unstable AP (uAP) and aortic valve replacement due to aortic stenosis (AS) were analyzed for the detection of miRNAs. We named these samples S-sAP, S-uAP, S-AS, PF-sAP, PF-uAP, and PF-AS, respectively. We first measured the levels of miR-423-5p, which was recognized previously as a biomarker for heart failure. miR-423-5p levels were significantly higher in PF than serum. Although there was no difference in miR-423-5p levels among the PF-AS, PF-sAP, and PF-uAP, its levels were significantly elevated in S-uAP compared with those in S-AS and S-sAP. In order to clarify the source of miR-423-5p in PF, we measured the levels of muscle-enriched miR-133a and vascular-enriched miR-126 and miR-92a in the same samples. miR-133a levels were significantly higher in serum than in PF, and it was elevated in S-uAP compared with S-AS. miR-126 level was significantly increased in serum compared with PF, and the level of miR-92a the similar tendency. miR-423-5p is located in the first intron of NSRP1. There is another miRNA, miR-3184, encoded in the opposite direction in the same region. In vitro experiments indicated that the duplex of miR-423-5p and miR-3184-3p was more resistant to RNase than the duplex of miR-423-5p and miR-133-3p, which may help to stabilize miR-423-5p in the PF.

Conclusions

Our results suggested that miR-423-5p is enriched in PF, and serum miR-423-5p may be associate with uAP. Its expression pattern was different to that of muscle- and vascular-enriched miRNAs, miR-133a, miR-126, and miR-92a.