DGR mutagenic transposition occurs via hypermutagenic reverse transcription primed by nicked template RNA.

Diversity-generating retroelements (DGRs) are molecular evolution machines that facilitate microbial adaptation to environmental changes. Hypervariation occurs via a mutagenic retrotransposition process from a template repeat (TR) to a variable repeat (VR) that results in adenine-to-random nucleotide conversions. Here we show that reverse transcription of the Bordetella phage DGR is primed by an adenine residue in TR RNA and is dependent on the DGR-encoded reverse transcriptase (bRT) and accessory variability determinant (Avd ), but is VR-independent. We also find that the catalytic center of bRT plays an essential role in site-specific cleavage of TR RNA for cDNA priming. Adenine-specific mutagenesis occurs during reverse transcription and does not involve dUTP incorporation, indicating it results from bRT-catalyzed misincorporation of standard deoxyribonucleotides. In vivo assays show that this hybrid RNA-cDNA molecule is required for mutagenic transposition, revealing a unique mechanism of DNA hypervariation for microbial adaptation

A new topology of the HK97-like fold revealed in Bordetella bacteriophage by cryoEM at 3.5 A resolution.

Bacteriophage BPP-1 infects and kills Bordetella species that cause whooping cough. Its diversity-generating retroelement (DGR) provides a naturally occurring phage-display system, but engineering efforts are hampered without atomic structures. Here, we report a cryo electron microscopy structure of the BPP-1 head at 3.5 Å resolution. Our atomic model shows two of the three protein folds representing major viral lineages: jellyroll for its cement protein (CP) and HK97-like ('Johnson') for its major capsid protein (MCP). Strikingly, the fold topology of MCP is permuted non-circularly from the Johnson fold topology previously seen in viral and cellular proteins. We illustrate that the new topology is likely the only feasible alternative of the old topology. β-sheet augmentation and electrostatic interactions contribute to the formation of non-covalent chainmail in BPP-1, unlike covalent inter-protein linkages of the HK97 chainmail. Despite these complex interactions, the termini of both CP and MCP are ideally positioned for DGR-based phage-display engineering. DOI: http://dx.doi.org/10.7554/eLife.01299.001

The consistency method of linguistic information and other four preference information in group decision-making

The transformation method between the linguistic judgement matrix and other four forms of preference information is researched in this paper. The four forms of preference information are preference ordering, utility value, reciprocal judgement matrices and complementary judgement matrices. First, based on the definition of preference information, the mutual transformation methods of linguistic judgement matrix and other four forms of preference information are given. Then new transformation equations are obtained. Next, it is proved that when the linguistic judgement matrix has complete consistency, the other four forms of preference information will have complete consistency too. Finally, a numerical analysis is offered to show that these methods are feasible and effective

Selenium Attenuates Chronic Heat Stress-Induced Apoptosis via the Inhibition of Endoplasmic Reticulum Stress in Mouse Granulosa Cells

Heat stress induces apoptosis in various cells. Selenium, an essential micronutrient, has beneficial effects in maintaining the cellular physiological functions. However, its potential protective action against chronic heat stress (CHS)-induced apoptosis in granulosa cells and the related molecular mechanisms are not fully elucidated. In this study, we investigated the roles of selenium in CHS-induced apoptosis in mouse granulosa cells and explored its underlying mechanism. The heat treatment for 6–48 h induced apoptosis, potentiated caspase 3 activity, increased the expression levels of apoptosis-related gene BAX and ER stress markers, glucose-regulated protein 78 (GRP78), and CCAAT/enhancer binding protein homologous protein (CHOP) in mouse granulosa cells. The treatment with ER stress inhibitor 4-PBA significantly attenuated the adverse effects caused by CHS. Selenium treatment significantly attenuated the CHS- or thapsigargin (Tg, an ER stress activator)-induced apoptosis, potentiation of caspase 3 activity, and the increased protein expression levels of BAX, GRP78, and CHOP. Additionally, treatment of the cells with 5 ng/mL selenium significantly ameliorated the levels of estradiol, which were decreased in response to heat exposure. Consistently, administering selenium supplement alleviated the hyperthermia-caused reduction in the serum estradiol levels in vivo. Together, our findings indicate that selenium has protective effects on CHS-induced apoptosis via inhibition of the ER stress pathway. The current study provides new insights in understanding the role of selenium during the process of heat-induced cell apoptosis

Activation of CREBZF Increases Cell Apoptosis in Mouse Ovarian Granulosa Cells by Regulating the ERK1/2 and mTOR Signaling Pathways

CREBZF, a multifunction transcriptional regulator, participates in the regulation of numerous cellular functions. The aims of the present study were to detect the localization of CREBZF expression in the ovary and explore the role of CREBZF and related mechanisms in the apoptosis of ovarian granulosa cells. We found by immunohistochemistry that CREBZF was mainly located in granulosa cells and oocytes during the estrous cycle. Western blot analysis showed that SMILE was the main isoform of CREBZF in the ovary. The relationship between apoptosis and CREBZF was assessed via CREBZF overexpression and knockdown. Flow cytometry analysis showed that CREBZF induced cell apoptosis in granulosa cells. Western bolt analysis showed that overexpression of CREBZF upregulated BAX and cleaved Caspase-3, while it downregulated BCL-2. Furthermore, overexpression of CREBZF inhibited the ERK1/2 and mTOR signaling pathways through the phosphorylation of intracellular-regulated kinases 1/2 (ERK1/2) and p70 S6 kinase (S6K1). Moreover, we found that CREBZF also activated autophagy by increasing LC3-II. In summary, these results suggest that CREBZF might play a proapoptotic role in cell apoptosis in granulosa cells, possibly by regulating the ERK1/2 and mTOR signaling pathways

The Determination Method of Satisfactory Consistency of the Interval Number Pairwise Comparisons Matrix Based on Submatrix

The decision-maker obtains the pairwise comparisons matrix by comparing two entities. In the process of comparing the two entities, the relationship between the two entities and other entities is not considered. In this way, the judgment may be illogical. This paper mainly studies the satisfactory consistency of the interval number pairwise comparisons matrix based on cyclic matrix. Firstly, the illogical judgment entity in the process of the decision-maker’s judgment is expressed by the cyclic matrix. There are three entities and four entities to form the cyclic matrix. The relationship and various forms of the cyclic cycle formed by the four entities and the three entities are discussed; then, the satisfactory consistency of the interval number pairwise comparisons matrix is determined by judging whether there is a cyclic matrix in the submatrix of the interval number pairwise comparisons matrix. Finally, two examples are given to verify the rationality and effectiveness of the method

The Bordetella bacteriophage DGR employs similar mechanisms for retrotransposition in heterologous species

Diversity-generating retroelements (DGRs) are a unique group of retroelements found in bacteria, archaea and their viruses. They mediate hyperdiversification of protein-encoding DNA sequences in facilitate the adaptation of their hosts to changing environments. The prototype DGR was discovered in the Bordetella bacteriophage BPP-1 and consists of three genes, mtd (major tropism determinant), avd (accessory variability determinant) and brt (Bordetella reverse transcriptase), and two imperfect repeats, variable repeat (VR) and template repeat (TR). VR is located at the 3' end of mtd, which encodes the phage distal tail fiber protein responsible for receptor recognition. Diversification of mtd results from unidirectional transfer of sequence information from TR to VR during which adenine residues in TR are converted into random nucleotides in VR, leading to phage tropic variants that recognize different receptor molecules. Here, we show that the BPP-1 DGR is also functional in heterologous bacterial species - Escherichia coli and Pseudomonas aeruginosa, and uses a similar mechanism for cDNA synthesis. However, efficiency of DGR mutagenic homing is affected by target sequence orientation in plasmids. Interestingly, overexpression of Avd and bRT has differential effects on DGR homing into targets inserted in different vectors. Surprisingly, homing into plasmid targets in E. coli is found to be largely independent of IMH (initiation of mutagenic homing) and the DNA stem-loop, elements important for its homing into native phage targets.Santa S. Naorem, Jin Han, Christa Jackson, Bingyue Zhang, James Guo and Huatao Guo ; Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, Missouri 6521