The complete genome sequence of a single-stranded RNA virus from the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois)

The complete genome sequence of a single-stranded RNA virus infecting the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), was identified by sequencing cDNA prepared from insects collected from the Mississippi Delta. The 9655 nucleotide positive-sense single-stranded RNA genome of the L. lineolaris single-stranded RNA virus (LyLV-1) contained a single open reading frame of 8958 nucleotides encoding a 2986 amino acid genome polypeptide. The open reading frame was flanked by untranslated regions of 603 and 69 nucleotides at the 5'- and 3'- ends of the genome, respectively. Database searches and homology based modeling was used to identify four capsid proteins (VP1-VP4), helicase/AAA-ATPase, cysteine protease (C3P), protease 2A, and the RNA-directed RNA polymerase (RdRp). In addition, a region with weak similarity to the eukaryotic structural maintenance of chromosome (SMC) domain was identified near the amino-terminal of the polyprotein and adjacent to the VP1 domain. The amino acid sequence of LyLV-1 was approximately 44.4% similar to that of sacbrood virus (SBV) of the honey bee. The genomic organization of both viruses showed remarkable similarity with the exception of highly divergent amino acid regions flanking fairly conserved structural and non-structural polypeptide regions. High similarity to the SBV genome and similarities in the genome organization and amino acid sequence with the viruses of the family Iflaviridae suggested that LyLV-1 was a novel member of this family. Virus particles were 39 nm in diameter and appeared to transmit vertically via eggs. Although this virus may only cause covert infections under normal conditions, the potential for using this virus in biological control of L. lineolaris is discussed.     

Delineating the role of alterations in lipid metabolism to the pathogenesis of inherited skeletal and cardiac muscle disorders: Thematic Review Series: Genetics of Human Lipid Diseases

As the specific composition of lipids is essential for the maintenance of membrane integrity, enzyme function, ion channels, and membrane receptors, an alteration in lipid composition or metabolism may be one of the crucial changes occurring during skeletal and cardiac myopathies. Although the inheritance (autosomal dominant, autosomal recessive, and X-linked traits) and underlying/defining mutations causing these myopathies are known, the contribution of lipid homeostasis in the progression of these diseases needs to be established. The purpose of this review is to present the current knowledge relating to lipid changes in inherited skeletal muscle disorders, such as Duchenne/Becker muscular dystrophy, myotonic muscular dystrophy, limb-girdle myopathic dystrophies, desminopathies, rostrocaudal muscular dystrophy, and Dunnigan-type familial lipodystrophy. The lipid modifications in familial hypertrophic and dilated cardiomyopathies, as well as Barth syndrome and several other cardiac disorders associated with abnormal lipid storage, are discussed. Information on lipid alterations occurring in these myopathies will aid in the design of improved methods of screening and therapy in children and young adults with or without a family history of genetic diseases.     

Teaching an old scaffold new tricks: monobodies constructed using alternative surfaces of the FN3 scaffold

The fibronectin type III domain (FN3) has become one of the most widely used non-antibody scaffolds for generating new binding proteins. Because of its structural homology to the immunoglobulin domain, combinatorial libraries of FN3 designed to date have primarily focused on introducing amino acid diversity into three loops that are equivalent to antibody complementarity-determining regions. Here, we report an FN3 library that utilizes alternative positions for presenting amino acid diversity. We diversified positions on a β-sheet and surface loops that together form a concave surface. The new library produced binding proteins (termed "monobodies") to multiple target proteins, generally with similar efficacy as the original, loop-focused library. The crystal structure of a monobody generated from the new library in complex with its target, the Abl SH2 domain, revealed that a concave surface of the monobody, as intended in our design, bound to a convex surface of the target with the interface area being among the largest of published structures of monobody-target complexes. This mode of interaction differs from a common binding mode for single-domain antibodies and antibody mimics in which recognition loops recognize clefts in targets. Together, this work illustrates the utilization of different surfaces of a single immunoglobulin-like scaffold to generate binding proteins with distinct characteristics.     

Chronic shift-lag alters the circadian clock of NK cells and promotes lung cancer growth in rats

Prolonged subjection to unstable work or lighting schedules, particularly in rotating shift-workers, is associated with an increased risk of immune-related diseases, including several cancers. Consequences of chronic circadian disruption may also extend to the innate immune system to promote cancer growth, as NK cell function is modulated by circadian mechanisms and plays a key role in lysis of tumor cells. To determine if NK cell function is disrupted by a model of human shift-work and jet-lag, Fischer (344) rats were exposed to either a standard 12:12 light-dark cycle or a chronic shift-lag paradigm consisting of 10 repeated 6-h photic advances occurring every 2 d, followed by 5-7 d of constant darkness. This model resulted in considerable circadian disruption, as assessed by circadian running-wheel activity. NK cells were enriched from control and shifted animals, and gene, protein, and cytolytic activity assays were performed. Chronic shift-lag altered the circadian expression of clock genes, Per2 and Bmal1, and cytolytic factors, perforin and granzyme B, as well as the cytokine, IFN-γ. These alterations were correlated with suppressed circadian expression of NK cytolytic activity. Further, chronic shift-lag attenuated NK cell cytolytic activity under stimulated in vivo conditions, and promoted lung tumor growth following i.v. injection of MADB106 tumor cells. Together, these findings suggest chronic circadian disruption promotes tumor growth by altering the circadian rhythms of NK cell function.     

CYTOLOGICAL AND PHYLOGENETIC DIVERSITY IN FRESHWATER ESOPTRODINIUM/BERNARDINIUM SPECIES (DINOPHYCEAE)1

The genera Esoptrodinium Javornický and Bernardinium Chodat comprise freshwater, athecate dinoflagellates with an incomplete cingulum but differing reports regarding cingulum orientation and the presence of chloroplasts and an eyespot. To examine this reported diversity, six isolates were collected from different freshwater ponds and brought into clonal culture. The isolates were examined using LM to determine major cytological differences, and rDNA sequences were compared to determine relatedness and overall phylogenetic position within the dinoflagellates. All isolates were athecate with a left‐oriented cingulum that did not fully encircle the cell, corresponding to the current taxonomic concept of Esoptrodinium. However, consistent cytological differences were observed among clonal isolates. Most isolates exhibited unambiguous pale green chloroplasts and a distinct bright‐red eyespot located at the base of the longitudinal flagellum. However, one isolate had cryptic chloroplasts that were difficult to observe using LM, and another had an eyespot that was so reduced as to be almost undetectable. Another isolate lacked visible chloroplasts but did possess the characteristic eyespot. Nuclear rDNA phylogenies strongly supported a monophyletic Esoptrodinium clade containing all isolates from this study together with a previous sequence from Portugal, within the Tovelliaceae. Esoptrodinium subclades were largely correlated with cytological differences, and the data suggested that independent chloroplast and eyespot reduction and/or loss may have occurred within this taxon. Overall, the isolates encompassed the majority of cytological diversity reported in previous observations of Bernardinium/Esoptrodinium in field samples. Systematic issues with the current taxonomic distinction between Bernardinium and Esoptrodinium are discussed.     

Peroxiredoxin II Regulates Effector and Secondary Memory CD8+ T Cell Responses

Reactive oxygen intermediates (ROI) generated in response to receptor stimulation play an important role in cellular responses. However, the effect of increased H₂O₂ on an antigen-specific CD8⁺ T cell response was unknown. Following T cell receptor (TCR) stimulation, the expression and oxidation of peroxiredoxin II (PrdxII), a critical antioxidant enzyme, increased in CD8⁺ T cells. Deletion of PrdxII increased ROI, S phase entry, division, and death during in vitro division. During primary acute viral and bacterial infection, the number of effector CD8⁺ T cells in PrdxII-deficient mice was increased, while the number of memory cells were similar to those of the wild-type cells. Adoptive transfer of P14 TCR transgenic cells demonstrated that the increased expansion of effector cells was T cell autonomous. After rechallenge, effector CD8⁺ T cells in mutant animals were more skewed to memory phenotype than cells from wild-type mice, resulting in a larger secondary memory CD8⁺ T cell pool. During chronic viral infection, increased antigen-specific CD8⁺ T cells accumulated in the spleens of PrdxII mutant mice, causing mortality. These results demonstrate that PrdxII controls effector CD8⁺ T cell expansion, secondary memory generation, and immunopathology.     

Acute impact of intermittent pneumatic leg compression frequency on limb hemodynamics, vascular function, and skeletal muscle gene expression in humans

The mechanisms by which intermittent pneumatic leg compression (IPC) treatment effectively treats symptoms associated with peripheral artery disease remain speculative. With the aim of gaining mechanistic insight into IPC treatment, the purpose of this study was to investigate the effect of IPC frequency on limb hemodynamics, vascular function, and skeletal muscle gene expression. In this two study investigation, healthy male subjects underwent an hour of either high-frequency (HF; 2-s inflation/3-s deflation) or low-frequency (LF; 4-s inflation/16-s deflation) IPC treatment of the foot and calf. In study 1 (n = 11; 23.5 ± 4.7 yr), subjects underwent both HF and LF treatment on separate days. Doppler/ultrasonography was used to measure popliteal artery diameter and blood velocity at baseline and during IPC treatment. Flow-mediated dilation (FMD) and peak reactive hyperemia blood flow (RHBF) were determined before and after IPC treatment. In study 2 (n = 19; 22.0 ± 4.6 yr), skeletal muscle biopsies were taken from the lateral gastrocnemius of the treated and control limb at baseline and at 30- and 150-min posttreatment. Quantitative PCR was used to assess mRNA concentrations of genes associated with inflammation and vascular remodeling. No treatment effect on vascular function was observed. Cuff deflation resulted in increased blood flow (BF) and shear rate (SR) in both treatments at the onset of treatment compared with baseline (P < 0.01). BF and SR significantly diminished by 45 min of HF treatment only (P < 0.01). Both treatments reduced BF and SR and elevated oscillatory shear index compared with baseline (P < 0.01) during cuff inflation. IPC decreased the mRNA expression of cysteine-rich protein 61 from baseline and controls (P <0 .01) and connective tissue growth factor from baseline (P < 0.05) in a frequency-dependent manner. In conclusion, a single session of IPC acutely impacts limb hemodynamics and skeletal muscle gene expression in a frequency-dependent manner but does not impact vascular function.     

Mimicking dynamic in vivo environments with stimuli-responsive materials for cell culture

Traditional synthetic substrates and matrices for cell culture have proven to be of only limited utility in efforts to understand and control cell behavior, in large part because they fail to capture the multifarious biochemical, mechanical, geometric and dynamic characteristics of in vivo environments. However, recent advances in materials chemistry and engineering have begun to provide researchers with a toolbox to mimic the complex characteristics of natural extracellular matrices (ECMs), providing new pathways to explore cell-matrix interactions and direct cell fate under physiologically realistic conditions. In this review, we describe recent developments in stimuli-responsive materials as dynamic substrates and matrices for cell culture, and highlight their use in furthering our understanding of how cells respond to temporal variations in their environment.     

Assessment Strategies and the Effects of Fighting Experience on Future Contest Performance in the Green Anole (Anolis carolinensis)

Social experiences can be useful sources of information for animals charged with making fitness‐related decisions. Fighting experience can alter an animal's perception of its fighting ability possibly leading to changes in future contest decisions, which may increase/decrease their probability of winning future contests. Winner and loser effects have been revealed in a wide array of animals, but studies using reptilian models are rare. This study investigated the impact of fighting experience on future contest performance and outcome in the green anole lizard and investigated the assessment strategies used by anoles during contests of different intensities. To determine whether the green anole expresses winner or loser effects, focal animals engaged in a primary contest with a smaller (larger) opponent to gain a winning (losing) experience; opponent size asymmetries were a significant predictor of contest outcome. Focal individuals were isolated for 2 d before being given a secondary contest with a size‐matched, naïve opponent. We found no evidence of winner or loser effects 2 d following a previous contest. Although previous contest outcome did not dictate future contest success, dynamics of the previous contest did. Highly aggressive primary contest losers won a significant proportion of the secondary contests, while less aggressive losers were more apt to lose the secondary contest. Secondary contest success of prior winners was not influenced by earlier contest performance. Further analyses of contest dynamics reveal that individuals may use different assessment strategies depending on the intensity of the contest. Our results demonstrate that future contest success may be driven more by individual performance in a prior contest and less by prior contest outcome.