Classification of G proteins and prediction of GPCRs-G proteins coupling specificity using continuous wavelet transform and information theory

The coupling between G protein-coupled receptors (GPCRs) and guanine nucleotide-binding proteins (G proteins) regulates various signal transductions from extracellular space into the cell. However, the coupling mechanism between GPCRs and G proteins is still unknown, and experimental determination of their coupling specificity and function is both expensive and time consuming. Therefore, it is significant to develop a theoretical method to predict the coupling specificity between GPCRs and G proteins as well as their function using their primary sequences. In this study, a novel four-layer predictor (GPCRsG_CWTIT) based on support vector machine (SVM), continuous wavelet transform (CWT) and information theory (IT) is developed to classify G proteins and predict the coupling specificity between GPCRs and G proteins. SVM is used for construction of models. CWT and IT are used to characterize the primary structure of protein. Performance of GPCRsG_CWTIT is evaluated with cross-validation test on various working dataset. The overall accuracy of the G proteins at the levels of class and family is 98.23 and 85.42%, respectively. The accuracy of the coupling specificity prediction varies from 74.60 to 94.30%. These results indicate that the proposed predictor is an effective and feasible tool to predict the coupling specificity between GPCRs and G proteins as well as their functions using only the protein full sequence. The establishment of such an accurate prediction method will facilitate drug discovery by improving the ability to identify and predict protein-protein interactions. GPCRsG_CWTIT and dataset can be acquired freely on request from the authors.     

小鼠胎肝红系细胞在分化过程中形态学观察及时序性表达分析

小鼠胎肝是小鼠发育早期主要的造血器官,红系细胞在胎肝造血过程中形态特征和组成成分等方面发生了明显变化。根据红系细胞体积的变化,利用Countstar细胞计数仪对小鼠E12．5-E17．5胎肝中直径8-14岬细胞进行数量统计,再结合观测到的红系细胞的形态特征和血红蛋白表达量的不同,将E9．5．E17．5胎肝中的细胞分为10类。统计结果显示,随着胎肝造血系统的发育,哺乳类红系细胞在终末分化时出现细胞体积减小、细胞核固缩、排核和血红蛋白表达量增加等时序性变化。红系细胞表面特异标志Terll9和CD71在EryD中高表达而在成体骨髓细胞和外周血细胞中表达较低的结果表明胎肝中红系细胞具有较高的分化能力。这些数据为研究红系分化、克隆红系分化相关基因及探讨红白血病发生的机制提供了理论依据。     

Immobilization of Acetobacter sp. CCTCC M209061 for efficient asymmetric reduction of ketones and biocatalyst recycling

ABSTRACT: BACKGROUND: The bacterium Acetobacter sp. CCTCC M209061 is a promising whole-cell biocatalyst with exclusive anti-Prelog stereoselectivity for the reduction of prochiral ketones that can be used to make valuable chiral alcohols such as (R)-4-(trimethylsilyl)-3-butyn-2-ol. Although it has promising catalytic properties, its stability and reusability are relatively poor compared to other biocatalysts. Hence, we explored various materials for immobilizing the active cells, in order to improve the operational stability of biocatalyst. RESULTS: It was found that Ca-alginate give the best immobilized biocatalyst, which was then coated with chitosan to further improve its mechanical strength and swelling-resistance properties. Conditions were optimized for formation of reusable immobilized beads which can be used for repeated batch asymmetric reduction of 4[prime]-chloroacetophenone. The optimized immobilized biocatalyst was very promising, with a specific activity of 85% that of the free-cell biocatalyst (34.66 mumol/min/g dw of cells for immobilized catalyst vs 40.54 mumol/min/g for free cells in the asymmetric reduction of 4[prime]-chloroacetophenone). The immobilized cells showed better thermal stability, pH stability, solvent tolerance and storability compared with free cells. After 25 cycles reaction, the immobilized beads still retained >50% catalytic activity, which was 3.5 times higher than degree of retention of activity by free cells reused in a similar way. The cells could be recultured in the beads to regain full activity and perform a further 25 cycles of the reduction reaction. The external mass transfer resistances were negligible as deduced from Damkohler modulus Da < <1, and internal mass transfer restriction affected the reduction action but was not the principal rate-controlling step according to effectiveness factors eta < 1 and Thiele modulus 0.3<[empty set] <1. CONCLUSIONS: Ca-alginate coated with chitosan is a highly effective material for immobilization of Acetobacter sp. CCTCC M209061 cells for repeated use in the asymmetric reduction of ketones. Only a small cost in terms of the slightly lower catalytic activity compared to free cells could give highly practicable immobilized biocatalyst.     

Automatic neutrophil nucleus lobe counting based on graph representation of region skeleton

Abnormal neutrophil nucleus lobation (like left shift and right shift) helps to diagnose for some clinical conditions. Currently, quantification of it depends on the manual microscopic inspection of blood smears by clinicians. The quality of the manual inspection is extremely limited by the efficiency of clinicians and their medical background. This article proposed an automatic lobe counting method based on the graph representation of the nucleus region skeletons. Skeletons of the segmented nucleus regions are computed by augmented Fast Marching Method and transformed into plane graphs. Then the nucleus lobes are split based on the extracted graph properties including width distribution along the skeleton and graph structure decomposition. Experiments show that the proposed method could efficiently approaches the real lobe numbers in blood smears and reliably distinguish the stabs from the segmented neutrophils, thus it should be helpful in automatic neutrophil lobe number quantification and abnormal lobation diagnosis. ? 2012 International Society for Advancement of Cytometry.     

Variability and stability of tea weevil‐induced volatile emissions from tea plants with different weevil densities,photoperiod and infestation duration

Abstract After herbivore attack, many plants emit herbivore‐induced plant volatiles (HIPVs). HIPVs can attract carnivores and/or repel herbivores, thereby mediating tritrophic plant–herbivore–carnivore interactions. HIPVs act as chemical information between organisms; hence, their variability and stability are vital. In the present study, variations in the volatile emissions, from the tea plant Camellia sinensis (O. Ktze) damaged by the tea weevil Myllocerinus aurolineatus (Voss) (Coleoptera: Curculionidae), with weevil densities, photoperiod and infestation duration, were investigated. The volatiles induced by high‐density weevils were more abundant in composition and amount than those induced by low‐density weevils, whether at noon, night or after weevil removal. The induced volatile emissions were similar on the second and third day after infestation, and the emissions of the major induced compounds displayed diurnal cycles. Linalool, (E,E)‐α‐farnesene, and benzyl nitrile were emitted mainly at noon, whereas 1,3,8‐p‐menthatriene and (E)‐β‐ocimene were maximally emitted at night. Given the different emission dynamics, significant differences were found between noon‐ and night‐induced volatiles. In summary, tea plants damaged by different weevil densities emitted a relatively stable signal at a particular time. This stability could be attributed to the similarities under the two densities of the main induced volatile compounds, their relative ratios and the emission dynamics of the induced volatiles.     

Metabolic network analysis revealed distinct routes of deletion effects between essential and non-essential genes

Interest in essential genes has arisen recently given their importance in antimicrobial drug development. Although knockouts of essential genes are commonly known to cause lethal phenotypes, there is insufficient understanding on the intermediate changes followed by genetic perturbation and to what extent essential genes correlate to other genes. Here, we characterized the gene knockout effects by using a list of affected genes, termed as 'damage lists'. These damage lists were identified through a refined cascading failure approach that was based on a previous topological flux balance analysis. Using an Escherichia coli metabolic network, we incorporated essentiality information into damage lists and revealed that the knockout of an essential gene mainly affects a large range of other essential genes whereas knockout of a non-essential gene only interrupts other non-essential genes. Also, genes sharing common damage lists tend to have the same essentiality. We extracted 72 core functional modules from the common damage lists of essential genes and demonstrated their ability to halt essential metabolites production. Overall, our network analysis revealed that essential and non-essential genes propagated their deletion effects via distinct routes, conferring mechanistic explanation to the observed lethality phenotypes of essential genes.     

Cyclin-dependent kinase 4 is a novel target in micoRNA-195-mediated cell cycle arrest in bladder cancer cells

miRNAs are a class of small-noncoding RNAs capable of negatively regulating gene expression. Here, we found that miR-195 is down-regulated in human bladder cancer tissue versus normal adjacent tissue. To better characterize the role of miR-195 in bladder cancer, we conducted gain of function analysis by transfecting bladder cancer cell line T24 with chemically synthesized miR-195 mimic. We identified CDK4, an early G1 cell cycle regulator, as a novel target of miR-195. Selective over-expression of miR-195 could induce G1-phase arrest in T24 cells, and subsequently inhibit T24 cell growth. These findings indicate that miR-195 could be a potential tumor suppressor in bladder cancer.     

Phase 1 clinical trial demonstrated that MUC1 positive metastatic seminal vesicle cancer can be effectively eradicated by modified Anti-MUC1 chimeric antigen receptor transduced T cells

Recent progress in chimeric antigen receptor-modified T-cell(CAR-T cell) technology in cancer therapy is extremely promising, especially in the treatment of patients with B-cell acute lymphoblastic leukemia. In contrast, due to the hostile immunosuppressive microenvironment of a solid tumor, CAR T-cell accessibility and survival continue to pose a considerable challenge, which leads to their limited therapeutic efficacy. In this study, we constructed two anti-MUC1 CAR-T cell lines. One set of CAR-T cells contained SM3 single chain variable fragment(sc Fv) sequence specifically targeting the MUC1 antigen and co-expressing interleukin(IL) 12(named SM3-CAR). The other CAR-T cell line carried the SM3 sc Fv sequence modified to improve its binding to MUC1 antigen(named p SM3-CAR) but did not co-express IL-12. When those two types of CAR-T cells were injected intratumorally into two independent metastatic lesions of the same MUC1+ seminal vesicle cancer patient as part of an interventional treatment strategy, the initial results indicated no side-effects of the MUC1 targeting CAR-T cell approach, and patient serum cytokines responses were positive. Further evaluation showed that p SM3-CAR effectively caused tumor necrosis, providing new options for improved CAR-T therapy in solid tumors.     

cDNA cloning and expression analysis of the juvenile hormone acid methyltransferase from seabuckthorn carpenterworm,Holcocerus hippophaecolus (Lepidoptera: Cossidae)

In this study, we report the cDNA cloning and sequence determination of Hh‐JHAMT from the seabuckthorn carpenterworm, Holcocerus hippophaecolus, by using rapid amplification of cDNA ends. The full‐length cDNA of putative Hh‐JHAMT was 1659 bp and contained a highly conserved Motif I, SAM motif I, which showed that Hh‐JHAMT like enzyme was a member of SAM‐dependent MTases. Moreover, putative Hh‐JHAMT had high homology to the other members of the JHAMT peptide family: 59% with Spodoptera litura, 54% with Bombyx mori and 54% with Helicoverpa armigera. Multiple alignments and phylogenetic analysis revealed that Hh‐JHAMT was closely related to JHAMT from Lepidoptera. Real‐time quantitative PCR experiments showed that Hh‐JHAMT mRNA expression was highest in the corpora allata (CA) complex, and was also detected at high levels during earlier larval and adult stages. The JHAMT mRNA level gradually declined during larval development, and the lowest amount of expression was observed in the pupal stage, while it increased to a higher level during adult stages. The pattern of Hh‐JHAMT expression was similar to the mode of JH biosynthesis. These results provided information concerning molecular characteristics of Hh‐JHAMT, whose expression profile suggests that the Hh‐JHAMT gene might be changed with larval development, metamorphosis and adult reproduction of the H. hippophaecolus.