Characterization and regulation of Schizosaccharomyces pombe gene encoding thioredoxin

A cDNA coding thioredoxin (TRX) was isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The 438 bp EcoRI fragment, which was detected by Southern hybridization, reveals an open reading frame which encodes a protein of 103 amino acids. The genomic DNA encoding TRX was also isolated from S. pombe chromosomal DNA using PCR. The cloned sequence contains 1795 bp and encodes a protein of 103 amino acids. However, the C-terminal region obtained from the cDNA clone is -Val-Arg-Leu-Asn-Arg-Ser-Leu, whereas the C-terminal region deduced from the genomic DNA appears to contain -Ala-Ser-Ile-Lys-Ala-Asn-Leu. This indicates that S. pombe cells contain two kinds of TRX genes which have dissimilar amino acid sequences only at the C-terminal regions. The heterologous TRX 1C produced from the cDNA clone could be used as a subunit of T7 DNA polymerase, while the TRX 1G from the genomic DNA could not. The upstream sequence and the region encoding the N-terminal 18 amino acids of the genomic DNA were fused into the promoterless beta-galactosidase gene of the shuttle vector YEp357 to generate the fusion plasmid pYKT24. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by hydrogen peroxide, menadione and aluminum chloride. It indicates that the expression of the cloned TRX gene is induced by oxidative stress.     

Genome size rather than content might affect call properties in toads of three ploidy levels (Anura: Bufonidae: Bufo viridis subgroup)

In vertebrates, genome size has been shown to correlate with nuclear and cell sizes, and influences phenotypic features, such as brain complexity. In three different anuran families, advertisement calls of polyploids exhibit longer notes and intervals than diploids, and difference in cellular dimensions have been hypothesized to cause these modifications. We investigated this phenomenon in green toads (Bufo viridis subgroup) of three ploidy levels, in a different call type (release calls) that may evolve independently from advertisement calls, examining 1205 calls, from ten species, subspecies, and hybrid forms. Significant differences between pulse rates of six diploid and four polyploid (3n, 4n) green toad forms across a range of temperatures from 7 to 27 °C were found. Laboratory data supported differences in pulse rates of triploids vs. tetraploids, but failed to reach significance when including field recordings. This study supports the idea that genome size, irrespective of call type, phylogenetic context, and geographical background, might affect call properties in anurans and suggests a common principle governing this relationship. The nuclear‐cell size ratio, affected by genome size, seems the most plausible explanation. However, we cannot rule out hypotheses under which call‐influencing genes from an unexamined diploid ancestral species might also affect call properties in the hybrid‐origin polyploids. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105 , 584–590.     

Effects of 12‐Hour Rotating Shifts on Menstrual Cycles of Photoelectronic Workers in Taiwan

12 h rotating shifts are common in high‐tech industries in Taiwan. The aim of this longitudinal study was to evaluate the effect of the disruption of circadian rhythms by the shift schedule on menstrual cycle length (MCL) and regularity of female workers at an optoelectronic company in Taiwan. We recruited females who worked rotating shifts in a clean room environment as the shift‐work group and female office workers who worked normal business hours as the comparison group. Every participant recorded their MCL for each menstruation cycle up to eight consecutive months prospectively and provided demographic characteristics, reproductive history, and menstrual characteristics. We collected data on 1,135 and 117 menstruation cycles in the shift‐work (n=280) and comparison groups (n=49). Whereas the two groups had similar group means for MCL and number of menstrual bleeding days, the prevalence of menstrual cycle irregularity (cycles<25 or>35 days) was higher in the shift‐work group (p=0.04). Univariate and multivariate logistic regression analyses demonstrated that rotating shift work was an independent predictor of menstrual cycle irregularity (odds ratio=1.71, 95% confidence interval: 1.03–2.88) after adjusting for shift‐work history, employment duration, coffee consumption, and pre‐employment menstrual cycle irregularity. Although further study is required to confirm our findings plus to explore prevention and control measures, our data indicate rotating shift work can increase the risk of MCL irregularity.     

Targeting Photoreceptors via Intravitreal Delivery Using Novel,Capsid-Mutated AAV Vectors

Development of viral vectors capable of transducing photoreceptors by less invasive methods than subretinal injection would provide a major advancement in retinal gene therapy. We sought to develop novel AAV vectors optimized for photoreceptor transduction following intravitreal delivery and to develop methodology for quantifying this transduction in vivo. Surface exposed tyrosine (Y) and threonine (T) residues on the capsids of AAV2, AAV5 and AAV8 were changed to phenylalanine (F) and valine (V), respectively. Transduction efficiencies of self-complimentary, capsid-mutant and unmodified AAV vectors containing the smCBA promoter and mCherry cDNA were initially scored in vitro using a cone photoreceptor cell line. Capsid mutants exhibiting the highest transduction efficiencies relative to unmodified vectors were then injected intravitreally into transgenic mice constitutively expressing a Rhodopsin-GFP fusion protein in rod photoreceptors (Rho-GFP mice). Photoreceptor transduction was quantified by fluorescent activated cell sorting (FACS) by counting cells positive for both GFP and mCherry. To explore the utility of the capsid mutants, standard, (non-self-complementary) AAV vectors containing the human rhodopsin kinase promoter (hGRK1) were made. Vectors were intravitreally injected in wildtype mice to assess whether efficient expression exclusive to photoreceptors was achievable. To restrict off-target expression in cells of the inner and middle retina, subsequent vectors incorporated multiple target sequences for miR181, an miRNA endogenously expressed in the inner and middle retina. Results showed that AAV2 containing four Y to F mutations combined with a single T to V mutation (quadY−F+T−V) transduced photoreceptors most efficiently. Robust photoreceptor expression was mediated by AAV2(quadY−F+T−V) −hGRK1−GFP. Observed off-target expression was reduced by incorporating target sequence for a miRNA highly expressed in inner/middle retina, miR181c. Thus we have identified a novel AAV vector capable of transducing photoreceptors following intravitreal delivery to mouse. Furthermore, we describe a robust methodology for quantifying photoreceptor transduction from intravitreally delivered AAV vectors.     

Cigarette Smoking Increases Abdominal and Visceral Obesity but Not Overall Fatness: An Observational Study

Background

Cigarette smoking and obesity are leading public health concerns. Both increase the risk for cardiovascular disease, cancer, and metabolic abnormalities. This study was conducted to assess the association between cigarette smoking and different types of obesity.

Methodology/Principal Findings

Two hundred eighty-three visitors to university hospitals located in four main provinces of South Korea were participated. All participants were classified as either current/past or never smokers and were divided into quartiles according to the total pack-years. Body mass index, waist circumference, total body fat percentage, and area of visceral and abdominal subcutaneous fat were measured. These results of each groups were compared. Waist circumference, and visceral fat area showed a J- or U-shaped association with total smoking amount during a lifetime. After restricting the analyses to past/current smokers, we found significant dose-dependent associations of smoking pack-years with abdominal and visceral obesity. Overall obesity measured by body mass index and total body fat percentage did not show such associations. Although current smokers clearly showed significant associations, we could not demonstrate these in past smokers, possibly because of the limited sample size.

Conclusions/Significance

Although smokers did not show significant difference in mean body mass index than those who never smoked, they showed more metabolically adverse fat distributions with increasing smoking amounts. This finding suggests that smoking is not beneficial for weight control. Therefore, smoking cessation and avoidance of smoking commencement should be addressed as important public health issues in preventing obesity and related complications.     

Important marine habitat off east Antarctica revealed by two decades of multi‐species predator tracking

Satellite telemetry data are a key source of animal distribution information for marine ecosystem management and conservation activities. We used two decades of telemetry data from the East Antarctic sector of the Southern Ocean. Habitat utilization models for the spring/summer period were developed for six highly abundant, wide‐ranging meso‐ and top‐predator species: Adélie Pygoscelis adeliae and emperor Aptenodytes forsteri penguins, light‐mantled albatross Phoebetria palpebrata, Antarctic fur seals Arctocephalus gazella, southern elephant seals Mirounga leonina, and Weddell seals Leptonychotes weddellii. The regional predictions from these models were combined to identify areas utilized by multiple species, and therefore likely to be of particular ecological significance. These areas were distributed across the longitudinal breadth of the East Antarctic sector, and were characterized by proximity to breeding colonies, both on the Antarctic continent and on subantarctic islands to the north, and by sea‐ice dynamics, particularly locations of winter polynyas. These areas of important habitat were also congruent with many of the areas reported to be showing the strongest regional trends in sea ice seasonality. The results emphasize the importance of on‐shore and sea‐ice processes to Antarctic marine ecosystems. Our study provides ocean‐basin‐scale predictions of predator habitat utilization, an assessment of contemporary habitat use against which future changes can be assessed, and is of direct relevance to current conservation planning and spatial management efforts.     

LabCaS: Labeling calpain substrate cleavage sites from amino acid sequence using conditional random fields

The calpain family of Ca²⁺‐dependent cysteine proteases plays a vital role in many important biological processes which is closely related with a variety of pathological states. Activated calpains selectively cleave relevant substrates at specific cleavage sites, yielding multiple fragments that can have different functions from the intact substrate protein. Until now, our knowledge about the calpain functions and their substrate cleavage mechanisms are limited because the experimental determination and validation on calpain binding are usually laborious and expensive. In this work, we aim to develop a new computational approach (LabCaS) for accurate prediction of the calpain substrate cleavage sites from amino acid sequences. To overcome the imbalance of negative and positive samples in the machine‐learning training which have been suffered by most of the former approaches when splitting sequences into short peptides, we designed a conditional random field algorithm that can label the potential cleavage sites directly from the entire sequences. By integrating the multiple amino acid features and those derived from sequences, LabCaS achieves an accurate recognition of the cleave sites for most calpain proteins. In a jackknife test on a set of 129 benchmark proteins, LabCaS generates an AUC score 0.862. The LabCaS program is freely available at: http://www.csbio.sjtu.edu.cn/bioinf/LabCaS . Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Lack of promotion of mammary, lung and skin tumorigenesis by 20 kHz triangular magnetic fields

In order to evaluate possible tumorigenic effects of a 20 kHz intermediate frequency triangular magnetic field (IF), a frequency emitted from TV and PC monitors at 6.25 microT rms, which is the regulated exposure limit of magnetic field for the public in Korea, mammary tumors were produced in female Sprague-Dawley rats by oral intubation of dimethylbenz(a)anthracene (DMBA), lung tumors in ICR mice by scapular region injection of benzo(a)pyrene (BP), and skin tumors in female ICR mice by topical application of DMBA and tetradecanoylphorbol ester (TPA). IF was applied 8 h/day for 14 weeks beginning the day after DMBA treatment for mammary tumor experiment, for 6 weeks after weaning for lung tumor, and for 20 weeks beginning 1 week after DMBA application for skin tumor experiment. For skin tumors, TPA was applied once a week for 19 weeks. Results showed no significant differences in tumor incidence, mean tumor number and volume, and histological patterns between IF magnetic-field exposed and sham control rats in the above three tumor models. Therefore, we conclude that within the limitation or number of animals and the experimental conditions, 20 kHz IF triangular magnetic field exposure of 6.25 microT does not appear to be a strong co-tumorigenic agent in the chosen murine mammary, lung and skin models.