Integrated small RNA profiling and degradome analysis of Anthurium andraeanum cultivars with different-colored spathes

MicroRNAs (miRNAs) are known to play vital roles in coloration of leaves, flowers, and fruits in plants. However, their functions in spathe coloration are poorly known. Anthurium andraeanum is a popular ornamental plant with various spathe colors. In this study, small RNA and degradome libraries from three A. andraeanum cultivars with different-colored spathes were constructed and sequenced. Illumina sequencing resulted in 94 conserved miRNAs, and 34 novel miRNAs in total were then identified based on precursor sequences and hairpin structures. Differential expression analysis showed that 52, 51, and 49 miRNAs were differentially expressed in comparisons of orange- versus white-colored spathe, purple- versus white-colored spathe, and purple- versus orange-colored spathe, respectively. The expression patterns of miRNAs and their corresponding targets involved in spathe coloration were further analyzed, and displayed that miR156b and miR529 were highly abundant in the spathes with higher anthocyanin content. These two miRNAs co-targeted a gene encoding SPL17, which may function as a negative regulator in anthocyanin accumulation. In addition, miR408 was also abundantly expressed in purple- and orange-colored spathes, and its typical targets were also identified. This comprehensive integrated analysis provides insight into the miRNA-mediated genetic regulation in spathe coloration of A. andraeanum.

     

Identification of a putative azadirachtin-binding complex from Drosophila Kc167 cells

Cell-proliferation in Drosophila Kc167 cells was inhibited by 50% when cell cultures contained 1.7 x 10(-7) M azadirachtin for 48 h (a tertranortriterpenoid from the neem tree Azadirachta indica). Drosophila Kc167 cells exhibited direct nuclear damage within 6-h exposure to azadirachtin (5 x 10(-7) M and above) or within 24 h when lower concentrations were used (1 x 10(-9) M). Fractionation of an extract of Drosophila Kc167 cells combined with ligand overlay technique resulted in the identification of a putative azadirachtin binding complex. Identification of the members of this complex by Peptide Mass Fingerprinting (PMF) and N-terminal sequencing identified heat shock protein 60 (hsp60) as one of its components.     

Functional analysis of the interferon-stimulated response element of porcine circovirus type 2 and its role during viral replication in vitro and in vivo

ABSTRACT: Background, Porcine circovirus type 2 (PCV2) is associated with post-weaning multi-systemic wasting syndrome (PMWS) in young weaned pigs. Immune stimulation was found to activate the replication of PCV2 and exacerbate the clinical outcome of the infection. Proper amount of interferon-alpha (IFN-alpha) is able to enhance PCV2 infection and production in Porcine kidney-15 (PK-15) cells when administered after inoculation; Methods, in the present study, luciferase reporter assays, construction of mutant viruses, Analysis the replication efficiency and the response to IFN-alpha treatment in PK-15 cells and animal experiments were carried out to analyze the function of interferon-stimulated response element (ISRE) of PCV2 and its role during viral replication in vitro and in vivo; Results, a functional viral ISRE sequence, 5`-CTGAAAACGAAAGA-3`, was identified in Rep gene promoter (Prep) of PCV2. PCV2 Prep is composed of two mini promoters, the proximal one span the sequence +1 to -106, containing an ISRE while the distal mini promoter is composed of three tandem GC box like sites locate at -85 to -194. It was demonstrated that viral ISRE is necessary for porcine IFN-alpha initiated luciferase expression enhancement and it plays an important role in affecting the replication efficiency of PCV2 in vivo and in vitro; Conclusions, These findings provide a theoretical basis for the Phenomenon of immunostimulation is able to enhance PCV2 infection, and improve the understanding of the complicated mechanisms involved in the host and pathogen interactions of PCV2.     

Protein interaction analysis of ST14 domains and their point and deletion mutants

ST14 (suppression of tumorigenicity 14) is a transmembrane serine protease that contains a serine protease catalytic (SP) domain, an SEA domain, two complement subcomponent C1r/s (CUB) domains, and four low density lipoprotein receptor class A domains. Glutathione S-transferase fusion proteins with SP, CUB, and low density lipoprotein receptor domains and their corresponding mutants were generated to analyze protein interactions with these domains. Modified glutathione S-transferase pull-down assays demonstrated the interaction between the SP domain and hepatocyte growth factor activator inhibitor-1. With the same method, a CUB domain-interacting protein was isolated and turned out to be the transmembrane protein with epidermal growth factor-like and two follistatin-like domains 1 (TMEFF1). Quantitative real time PCR revealed that the expression of the TMEFF1 gene was dependent on the transfection of the ST14 gene in the RKO cell line. Our results also suggested that ST14 and TMEFF1 were co-expressed in the human breast cancer cell line MCF7, human placenta, kidney, and liver tissues. Interestingly, these two genes were co-up-regulated in kidney tumors versus normal tissues, consistent with our results that showed the dependence of TMEFF1 expression on ST14 in RKO cells. Finally, homology modeling studies suggested that TMEFF1 might form a complex with ST14 by an interaction between epidermal growth factor and CUB domains.     

K562 cell growth activity and metabolism characteristics in APA microencapsulated culture and modeling study

Cell microencapsulation is likely to play a major role in cell and transplantation therapies in the next decade. The microcapsules provide a special microenvironment in which cells always have different behaviors compared with free non-encapsulated culture. In this work, the behaviors of K562 leukemia cells were studied once entrapped in solid and liquefied APA microcapsules as well as in free non-encapsulated culture. Glucose pulse culture was employed to characterize the growth and metabolism of microencapsulated K562 cells. And mathematical modeling was presented to develop a basis for the deeper understanding of cells responses to different culture environments. Based on the results of experiments and modeling, it was found that cells presented a better growing pattern and maintain the activity at a higher level for extending time. The concentration of lactate was higher in solid microcapsules culture than that of liquefied microcapsules culture, but the cell number was lower. And the lactate yield coefficients (lactate/glucose) were 0.8129, 0.6978 and 0.601 for free non-encapsulated, solid microcapsules and liquefied microcapsules culture, respectively. An increase of glucose concentration led a decrease of cell activity, The glucose consumption ratio were 99.9%, 86.8%, 49.4% and 28.6% with the decrease in its concentration from 2 to 4, 6, 10 g/L, however, the lactate yield coefficient were 0.7184, 0.6654, 0.8239 and 0.9693, respectively.     

Construction, Characterization, and Chromosomal Mapping of a Fosmid Library of the White-Cheeked Gibbon （Nomascus leucogenys）

Gibbons have experienced extensive karyotype rearrangements during evolution and represent an ideal model for studying the underlying molecular mechanism of evolutionary chromosomal rearrangements. It is anticipated that the cloning and sequence characterization of evolutionary chromosomal breakpoints will provide vital insights into the molecular force that has driven such a radical karyotype reshuffle in gibbons. We constructed and characterized a high-quality fosmid li- brary of the white-cheeked gibbon (Nomascus leucogenys) containing 192,000 non- redundant clones with an average insert size of 38 kb and 2.5-fold genome coverage. By end sequencing of 100 randomly selected fosmid clones, we generated 196 se- quence tags for the library. These end-sequenced fosmid clones were then mapped onto the chromosomes of the white-cheeked gibbon by fluorescence in situ hy- bridization, and no spurious chimeric clone was detected. BLAST search against the human genome showed a good correlation between the number of hit clones and the number of chromosomes, an indication of unbiased chromosomal distribu- tion of the fosmid library. The chromosomal distribution of the mapped clones is also consistent with the BLAST search result against human and white-cheeked gibbon genomes. The fosmid library and the mapped clones will serve as a valu- able resource for further studying gibbons' chromosomal rearrangements and the underlying molecular mechanism as well as for comparative genomic study in the lesser apes.     

IL-6: TNF-α之后的类风湿关节炎治疗关键靶点

IL-6是一种重要的细胞因子,在类风湿关节炎发病过程中发挥重要作用。本文对已上市的IL-6受体单克隆抗体tocilizumab对类风湿关节炎的临床疗效和安全性进行了总结,并和TNF-α阻断药物进行了对比,证明tocilizumab在药物疗效和副作用方面与TNF-α阻断药物相比各有优劣。另外也对在研的IL-6通路阻断单抗的临床试验结果进行了总结。结合本中心近年的研究和总结,IL-6是继TNF-α之后的另一个重要的类风湿关节炎治疗关键靶点,该类药物的上市为以后类风湿关节炎的个性化治疗提供了更多的选择。     

基于核型和分子系统学方法对中国猪尾鼠分类与分布的讨论

猪尾鼠属（Typhlomys）是啮齿类中的孑遗类群,片段化分布于我国南方和越南最北部的山地森林。前期我们基于形态和分子系统学的方法将猪尾鼠属现生类群划分为4个物种。但该分类系统仍存在争议和有待于解决的问题：一方面,有学者对大娄山猪尾鼠等是否应属于种级分类单元抱有疑问;另一方面,秦岭、南岭和广西周边的猪尾鼠种群的分类地位仍未有定论。因此,本研究采用核型分析以及分子系统学分析的方法,对我国猪尾鼠的分类及分布进行进一步梳理。本研究首次报道了中华猪尾鼠和大娄山猪尾鼠的G带核型和Ag-NORs的数目和分布,其中中华猪尾鼠2n=36,核型为14(M, SM) + 20 (A),XY (SM, A),Ag-NORs 6个;大娄山猪尾鼠2n=56,核型为2 (SM) + 52 (A),XX (SM, SM ),Ag-NORs 4个。上述核型与已报道的沙巴猪尾鼠核型（2n=38）存在显著差异,支持上述类群均为独立物种。基于线粒体Cyt b,ND2和COⅠ基因的进化关系分析,秦岭种群应属于大娄山猪尾鼠、南岭种群应属于中华猪尾鼠。而来自红河以东的云南南部的种群以及贵州南部的种群情况复杂,暗示云南、贵州和广西的喀斯特地区可能还存在未知的分类单元。