PU.1, a novel capase-3 substrate, partially contributes to chemotherapeutic agents-induced apoptosis in leukemic cells

PU.1 is one of key regulators of hematopoietic cell development, a tightly-regulated lineage-specific process. Here we provide the first evidence that PU.1 protein is cleaved into two fragments of 24 kDa and 16 kDa during apoptosis progression in leukemic cell lines and primary leukemic cells. Further experiments with specific capase-3 inhibitor Z-DEVD-fmk and the in vitro proteolytic system confirmed that PU.1 is a direct target of caspase-3. Using site-directed mutagenesis analyses, the aspartic acid residues at positions 97 and 151 of PU.1 protein were identified as capsase-3 target sites. More intriguingly, the suppression of PU.1 expression by small interfering RNAs (siRNAs) significantly inhibits DNA-damaging agents NSC606985 and etoposide-induced apoptosis in leukemic cells, together with the up-regulated expression of anti-apoptotic bcl-2 gene. These results would provide new insights for understanding the mechanism of PU.1 protein in hematopoiesis and leukemogenesis.     

Probing the structural basis for differential KCNQ1 modulation by KCNE1 and KCNE2

KCNE1 associates with KCNQ1 to increase its current amplitude and slow the activation gating process, creating the slow delayed rectifier channel that functions as a “repolarization reserve” in human heart. The transmembrane domain (TMD) of KCNE1 plays a key role in modulating KCNQ1 pore conductance and gating kinetics, and the extracellular juxtamembrane (EJM) region plays a modulatory role by interacting with the extracellular surface of KCNQ1. KCNE2 is also expressed in human heart and can associate with KCNQ1 to suppress its current amplitude and slow the deactivation gating process. KCNE1 and KCNE2 share the transmembrane topology and a high degree of sequence homology in TMD and surrounding regions. The structural basis for their distinctly different effects on KCNQ1 is not clear. To address this question, we apply cysteine (Cys) scanning mutagenesis to TMDs and EJMs of KCNE1 and KCNE2. We analyze the patterns of functional perturbation to identify high impact positions, and probe disulfide formation between engineered Cys side chains on KCNE subunits and native Cys on KCNQ1. We also use methanethiosulfonate reagents to probe the relationship between EJMs of KCNE subunits and KCNQ1. Our data suggest that the TMDs of both KCNE subunits are at about the same location but interact differently with KCNQ1. In particular, the much closer contact of KCNE2 TMD with KCNQ1, relative to that of KCNE1, is expected to impact the allosteric modulation of KCNQ1 pore conductance and may explain their differential effects on the KCNQ1 current amplitude. KCNE1 and KCNE2 also differ in the relationship between their EJMs and KCNQ1. Although the EJM of KCNE1 makes intimate contacts with KCNQ1, there appears to be a crevice between KCNQ1 and KCNE2. This putative crevice may perturb the electrical field around the voltage-sensing domain of KCNQ1, contributing to the differential effects of KCNE2 versus KCNE1 on KCNQ1 gating kinetics.     

Complete Genome Sequence of Hepatitis E Virus from Rabbits in the United States

Hepatitis E virus (HEV) is a single-strand positive-sense RNA virus in the family Hepeviridae. The disease caused by HEV, hepatitis E, is an important public health problem in developing countries of Asia and Africa and is also endemic in many industrialized countries, including the United States. HEV has been identified from several other animal species in addition to humans, including the pig, chicken, mongoose, deer, rabbit, ferret, bat, and fish. Here we report the complete genome sequence of the first strain of HEV from rabbits in the United States. Sequence and phylogenetic analyses revealed that the U.S. rabbit HEV is a distant member of the zoonotic genotype 3 HEV, thus raising a concern for potential zoonotic human infection. A unique 90-nucleotide insertion within the X domain of the ORF1 was identified in the rabbit HEV, and this insertion may play a role in the species tropism of HEV.     

Dynamics of Yersinia pestis and Its Antibody Response in Great Gerbils (Rhombomys opimus) by Subcutaneous Infection

Background

Rhombomys opimus (great gerbil) is a reservoir of Yersinia pestis in the natural plague foci of Central Asia. Great gerbils are highly resistant to Y. pestis infection. The coevolution of great gerbils and Y. pestis is believed to play an important role in the plague epidemics in Central Asia plague foci. However, the dynamics of Y. pestis infection and the corresponding antibody response in great gerbils have not been evaluated. In this report, animal experiments were employed to investigate the bacterial load in both the liver and spleen of infected great gerbils. The dynamics of the antibody response to the F1 capsule antigen of Y. pestis was also determined.

Methodology

Captured great gerbils that tested negative for both anti-F1 antibodies and bacterial isolation were infected subcutaneously with different doses (10⁵ to 10¹¹ CFU) of a Y. pestis strain isolated from a live great gerbil during routine plague surveillance in the Junggar Basin, Xinjiang, China. The clinical manifestations, changes in body weight, anal temperature, and gross anatomy of the infected animals were observed. The blood cell count, bacterial load, and anti-F1 antibody titers were determined at different time points after infection using a blood analyzer, plate counts, and an indirect hemagglutination assay, respectively.

Conclusions/Significance

The dynamics of bacterial load and the anti-F1 antibody concentration in great gerbils are highly variable among individuals. The Y. pestis infection in great gerbils could persist as long as 15 days. They act as an appropriate reservoir for plague in the Junggar Basin, which is part of the natural plague foci in Central Asia. The dynamics of the Y. pestis susceptibility of great gerbil will improve the understanding of its variable resistance, which would facilitate the development of more effective countermeasures for controlling plague epidemics in this focus.     

Endurance exercise training inhibits neointimal formation via enhancement of FOXOs expression in balloon-induced atherosclerosis rat model

[Purpose]

This study investigated the effect of endurance exercise on neointimal formation, endothelial-dependant relaxation and FOXO expression in balloon-induced carotid arteries of rats.

[Methods]

Male SD(Sprague-Dawley) rats of 8 weeks ages were randomly divided into 3 groups; Sham-operated control (SO, n=10), Balloon-induced control (BIC, n=10), and Balloon-induced exercise (BIE, n=10). Endurance exercise training was performed on treadmill (18 m/min, 0% grade, 60 min/day, 5 days/week, 4 weeks).

[Results]

Body weight is significantly reduced in BIE compared with BIC. Neointiaml formation in BIC was significantly higher than SO, but it was significantly recovered in BIE compared with BIC. Endothelial-dependent relaxation in BIC was significantly lower than SO, but it was significantly recovered in BIE compared with BIC and expression of FOXO1 and FOXO3a also were significantly increased BIE compared with BIC.

[Conclusion]

These data suggest that endurance exercise inhibits neointimal formation and endothelial-dependent relaxation via FOXO expression in balloon-induce atherosclerosis rat model.     

Accelerated regeneration of the skeletal muscle in RNF13-knockout mice is mediated by macrophage-secreted IL-4/IL-6

RING finger protein 13 (RNF13) is a newly identified E3 ligase reported to be functionally significant in the regulation of cancer development, muscle cell growth, and neuronal development. In this study, the function of RNF13 in cardiotoxin-induced skeletal muscle regeneration was investigated using RNF13-knockout mice. RNF13^-/- mice exhibited enhanced muscle regeneration —characterized by accelerated satellite cell proliferation —compared with wild-type mice. The expression of RNF13 was remarkably induced in macrophages rather than in the satellite cells of wild-type mice at the very early stage of muscle damage. This result indicated that inflammatory cells are important in RNF13-mediated satellite cell functions. The cytokine levels in skeletal muscles were further analyzed and showed that RNF13^-/- mice produced greater amounts of various cytokines than wild-type mice. Among these, IL-4 and IL-6 levels significantly increased in RNF13^-/- mice. The accelerated muscle regeneration phenotype was abrogated by inhibiting IL-4/IL-6 action in RNF13^-/- mice with blocking antibodies. These results indicate that RNF13 deficiency promotes skeletal muscle regeneration via the effects on satellite cell niche mediated by IL-4 and IL-6.     

益气活血方对肝星形细胞（HSC）Ⅰ型胶原的表达影响的研究

研究益气活血方剂抑制HSC合成Ⅰ型胶原的作用和生物学机制。采用免疫性肝纤维化大鼠模型,制备不同造模时间的病理血清、正常血清和益气活血方药物血清培养肝星形细胞HSC（Hepatic stellate cells）激光共 焦显微镜定量分析Ⅰ型胶原的表达。结果显示：（1）正常大鼠血清培养继代HSC表达较多的Ⅰ型胶原。纤维化模型大鼠血清诱导HSC表达的Ⅰ型胶原低于正常大鼠,益气活血药物血清可抑制纤维化模型血清诱导的HSCⅠ型胶原的表达,P＜0.05。（2）3周模型血清培养的HSC中分别加入10^-6、10^-7和10^-8ngγ-干扰素表明;10^-6、10^-7ngγ-干扰素和益气活血方具有相同的抑制HSC表达Ⅰ型胶原的作用,10^-7ngγ－干扰素抑制作用次之,P＜0.01。（3）1％胎牛血清可使原代HSC表达低水平Ⅰ型胶原。结果表明:益气活血方剂能够抑制HSC合成Ⅰ型胶原,其作用机制是改变了HSC的生活状态。     

Tools for integrated sequence-structure analysis with UCSF Chimera

Background  

Comparing related structures and viewing the structures in the context of sequence alignments are important tasks in protein structure-function research. While many programs exist for individual aspects of such work, there is a need for interactive visualization tools that: (a) provide a deep integration of sequence and structure, far beyond mapping where a sequence region falls in the structure and vice versa; (b) facilitate changing data of one type based on the other (for example, using only sequence-conserved residues to match structures, or adjusting a sequence alignment based on spatial fit); (c) can be used with a researcher's own data, including arbitrary sequence alignments and annotations, closely or distantly related sets of proteins, etc.; and (d) interoperate with each other and with a full complement of molecular graphics features. We describe enhancements to UCSF Chimera to achieve these goals.     

Response of physiologic metabolism and cell structures in mango fruit to exogenous methyl salicylate under low-temperature stress

We explored the effects of exogenous methyl salicylate (MeSA) on the development of chilling injury symptom, and the structure and composition of the pericarp, in mango ( Mangifera indica L. cv. 'Red 6') fruit under low-temperature stress using histochemical analysis and scanning electron microscopy together with Fourier transform infrared (FTIR) analysis. The results indicated that chilling injury symptom was remarkably limited in the fruit treated with MeSA at 0.1 m M as compared to the 5°C control, demonstrating the positive effects of MeSA in reducing chilling injury of mango fruit in low-temperature storage. In MeSA-treated fruit, the pericarp wax surface showed many cracks, and exocarp cells exhibited normal separation. The cell wall of exocarp contained lower amounts of pectic substances, aliphatics and phenolics in MeSA-treated fruit. In addition, MeSA-treated fruit contained more esterified substances and less carboxylate and carboxyl substances. Our work revealed the importance of MeSA in enhancing fruit tolerance to low-temperature stress and suggested a contribution of cellular structure and composition to this effect, which has not been reported previously.