Structural insights into the enzymatic mechanism of the pathogenic MAPK phosphothreonine lyase

The OspF family of phosphothreonine lyase, including SpvC from Salmonella, irreversibly inactivates the dual-phosphorylated host MAPKs (pT-X-pY) through beta elimination. We determined crystal structures of SpvC and its complex with a phosphopeptide substrate. SpvC adopts a unique fold of alpha/beta type. The disordered N terminus harbors a canonical D motif for MAPK substrate docking. The enzyme-substrate complex structure indicates that recognition of the phosphotyrosine followed by insertion of the threonine phosphate into an arginine pocket places the phosphothreonine into the enzyme active site. This requires the conformational flexibility of pT-X-pY, which suggests that p38 (pT-G-pY) is likely the preferred physiological substrate. Structure-based biochemical and enzymatic analysis allows us to propose a general acid/base mechanism for beta elimination reaction catalyzed by the phosphothreonine lyase. The mechanism described here provides a structural understanding of MAPK inactivation by a family of pathogenic effectors conserved in plant and animal systems and may also open a new route for biological catalysis.     

Reusable evanescent wave DNA biosensor for rapid, highly sensitive, and selective detection of mercury ions

Mercury ions (Hg(2+)) are a highly toxic and ubiquitous pollutants requiring rapid and sensitive on-site detection methods in the environment and foods. Herein, we report an envanescent wave DNA-based biosensor for rapid and very sensitive Hg(2+) detection based on a direct structure-competitive detection mode. In this system, a DNA probe covalently immobilized onto a fiber optic sensor contains a short common oligonucleotide sequences that can hybidize with a fluorescently labeled complementary DNA. The DNA probe also comprises a sequence of T-T mismatch pairs that binds with Hg(2+) to form a T-Hg(2+)-T complex by folding of the DNA segments into a hairpin structure. With a structure-competitive mode, a higher concentration of Hg(2+) leads to less fluorescence-labeled cDNA bound to the sensor surface and thus to lower fluorescence signal. The total analysis time for a single sample, including the measurement and surface regeneration, was under 6 min with a Hg(2+) detection limit of 2.1 nM. The high specificity of the sensor was demonstrated by evaluating its response to a number of potentially interfering metal ions. The sensor's surface can be regenerated with a 0.5% SDS solution (pH 1.9) over 100 times with no significant deterioration of performance. This platform is potentially applicable to detect other heavy metal ions or small-molecule analytes for which DNA/aptamers can be used as specific sensing probes.     

Comparative studies on histological and ultra-structure of the pituitary of different ploidy level fishes

The histological and ultra-structure of the pituitary in diploid red crucian carp(Carassius auratus red var.),triploid crucian carp and allotetraploid hybrids within and after the breeding season were comparatively studied.The result showed that there were six endocrine cell types in the pituitary of these three kinds of fishes,and there was an obvious difference in cell size among different ploidy level fishes.As for the same type of pituitary cells,the cell size was increased gradually with the in- creasing ploidy level.In the breeding season,the allotetraploid hybrids had higher proportion of go- nadotropin cells(GTH)than triploids,and the triploids had higher proportion of GTH than diploids.The results were related to the earlier sexual maturity of allotetraploid hybrids and sterility of triploid cru- cian carp.On the other hand,among the three kinds of fishes,the proportion of somatotropin(STH) cells in triploids crucian carp was the highest,whereas that in allotetraploid hybrids was the lowest. The results might be connected with the faster growth rate of triploids and slower growth rate of al- lotetraploid hybrids.In addition,in GTH cells of meso-adenohypophysis after the breeding season, there were many endocrine particles in triploids,while those endocrine particles were released from the cells in allotetraploids and diploids.This result showed that the sterility of triploid crucian carp might be related to the hormone which was not released from the GTH cells.In a word,the present study indicated that the differences in the structure of pituitary among different ploidy level fishes contributed to their difference in the growth rate and gonadal development.     

慢性肾脏病并发左室肥厚现况及其危险因素

目的：探索中老年慢性肾脏病并发左室肥厚（LVH）的现况及其危险因素。方法：对我院肾内科住院的40-75岁CKD2-5期患者210例的病历资料进行回顾性分析。结果：（1）心脏舒张功能减退发生率高于收缩功能减退（79.1%VS 20.3%P=0.000）;左房扩大检出率高于左室扩大检出率（46.5%VS 19.8%P=0.000）;室间隔增厚检出率（IVSH）也高于左室后壁增厚检出率（LVPWH）（43.0%VS 21.1%P=0.000）;LVH的发生率高于IVSH检出率（47.9%VS 35%P=0.001）,其中女性LVH高于男性（73.2%VS31.0%P=0.000）,然而若采用另外一种诊断标准,两者并无统计学差异（50%VS 34.5%P=0.068）。（2）IVSH组收缩压、脉压、血肌酐均高于无IVSH组。IVSH组除上述因素外血磷尚高于无IVSH组,但在CKD5期的亚组分析中仅收缩压与对照组相比有统计学差异。LVH组收缩压、脉压均高于无LVH组,而血红蛋白、体质指数则低于对照组。进一步Logistic回归分析提示仅性别、体质指数有统计学意义。结论：（1）40-75岁的心血管疾病高危的CKD患者中,采用超声心动图诊断LVH,根据公式计算的LVMI诊断阳性率最高,但诊断切点仍需进一步研究。（2）收缩压升高、脉压增大、贫血、低体质指数、女性均可能是LVH的危险因素,控制血压、纠正贫血和营养不良可能是防治LVH的重要靶点。     

Why Does Rhinopithecus bieti Prefer the Highest Elevation Range in Winter? A Test of the Sunshine Hypothesis

Environmental factors that affect spatiotemporal distribution patterns of animals usually include resource availability, temperature, and the risk of predation. However, they do not explain the counterintuitive preference of high elevation range in winter by the black-and-white snub-nosed monkey (Rhinopithecus bieti). We asked whether variation of sunshine along with elevations is the key driving force. To test this hypothesis, we conducted field surveys to demonstrate that there was a statistically significant pattern of high elevation use during winter. We then asked whether this pattern can be explained by certain environmental factors, namely temperature, sunshine duration and solar radiation. Finally, we concluded with a possible ecological mechanism for this pattern. In this study, we employed GIS technology to quantify solar radiation and sunshine duration across the monkey's range. Our results showed that: 1) R. bieti used the high altitude range between 4100-4400 m in winter although the yearly home range spanned from 3500-4500 m; 2) both solar radiation and sunshine duration increased with elevation while temperature decreased with elevation; 3) within the winter range, the use of range was significantly correlated with solar radiation and sunshine duration; 4) monkeys moved to the areas with high solar radiation and duration following a snowfall, where the snow melts faster and food is exposed earlier. We concluded that sunshine was the main factor that influences selection of high elevation habitat for R. bieti in winter. Since some other endotherms in the area exhibit similar winter distributional patterns, we developed a sunshine hypothesis to explain this phenomenon. In addition, our work also represented a new method of integrating GIS models into traditional field ecology research to study spatiotemporal distribution pattern of wildlife. We suggest that further theoretical and empirical studies are necessary for better understanding of sunshine influence on wildlife range use.     

Genome-scale screen for DNA methylation-based detection markers for ovarian cancer

Background

The identification of sensitive biomarkers for the detection of ovarian cancer is of high clinical relevance for early detection and/or monitoring of disease recurrence. We developed a systematic multi-step biomarker discovery and verification strategy to identify candidate DNA methylation markers for the blood-based detection of ovarian cancer.

Methodology/Principal Findings

We used the Illumina Infinium platform to analyze the DNA methylation status of 27,578 CpG sites in 41 ovarian tumors. We employed a marker selection strategy that emphasized sensitivity by requiring consistency of methylation across tumors, while achieving specificity by excluding markers with methylation in control leukocyte or serum DNA. Our verification strategy involved testing the ability of identified markers to monitor disease burden in serially collected serum samples from ovarian cancer patients who had undergone surgical tumor resection compared to CA-125 levels.We identified one marker, IFFO1 promoter methylation (IFFO1-M), that is frequently methylated in ovarian tumors and that is rarely detected in the blood of normal controls. When tested in 127 serially collected sera from ovarian cancer patients, IFFO1-M showed post-resection kinetics significantly correlated with serum CA-125 measurements in six out of 16 patients.

Conclusions/Significance

We implemented an effective marker screening and verification strategy, leading to the identification of IFFO1-M as a blood-based candidate marker for sensitive detection of ovarian cancer. Serum levels of IFFO1-M displayed post-resection kinetics consistent with a reflection of disease burden. We anticipate that IFFO1-M and other candidate markers emerging from this marker development pipeline may provide disease detection capabilities that complement existing biomarkers.     

Characterization of a fungal strain capable of degrading chlorpyrifos and its use in detoxification of the insecticide on vegetables

A fungal strain capable of utilizing chlorpyrifos as sole carbon and energy sources was isolated from soil by enrichment cultivation approach. The half-lives of degradation (DT₅₀) for chlorpyrifos at concentrations of 1, 10, and 100 mg l⁻¹ by the fungal strain DSP in mineral salt medium were measured to be 2.03, 2.93, and 3.49 days, respectively. Two cell-free extracts [E (1:10) and E (1:20)] from the fungal strain DSP in bran–glucose medium were prepared and used to enhance chlorpyrifos degradation on vegetables. Compared with the controls, the DT₅₀ of chlorpyrifos were reduced by 70.3%, 65.6%, 80.6%, 80.6%, and 86.1%, and by 53.8%, 43.2%, 66.0%, 54.3%, and 67.7% on E (1:20) and E (1:10) treated pakchoi, water spinach, Malabar spinach, haricot beans, and pepper, respectively. The 7-day residual values (R ₇) of chlorpyrifos on E (1:10) treated vegetables were all lower than the corresponding maximum residue levels of European Union (EU MRLs), except that the R ₇ value on haricot beans was slightly higher than the corresponding EU MRLs. The results indicate that cell-free extracts could rapidly degrade chlorpyrifos residues on vegetables.     

慢病毒载体介导IκB激酶-α(IKKα)基因RNA干扰转基因小鼠的制备

IκB激酶-α(IKKα)的功能与淋巴形成和乳腺发育相关,在乳腺肿瘤中观察到IKKα蛋白的过量表达.应用小鼠受精卵卵周隙注射慢病毒载体的方式,建立IKKα基因RNAi转基因小鼠模型,为在体内研究IKKα基因的生物学功能及其与肿瘤发生的关系创造条件.实验构建了针对IKKα基因RNAi(剔降)的慢病毒载体,并将载体导入小鼠受精卵卵周隙,获得携带该载体的转基因小鼠模型,经PCR鉴定首建鼠阳性率为15%.转基因小鼠外周血细胞IKKαmRNA表达量明显降低.初步表型观察分析,IKKα基因RNAi小鼠发育正常,进一步的分析工作正在进行中.