Sequencing,Annotation, and Characterization of the Influenza Ferret Infectome

Ferrets have become an indispensable tool in the understanding of influenza virus virulence and pathogenesis. Furthermore, ferrets are the preferred preclinical model for influenza vaccine and therapeutic testing. Here we characterized the influenza infectome during the different stages of the infectious process in ferrets with and without prior specific immunity to influenza. RNA from lung tissue and lymph nodes from infected and naïve animals was subjected to next-generation sequencing, followed by de novo data assembly and annotation of the resulting sequences; this process generated a library comprising 13,202 ferret mRNAs. Gene expression profiles during pandemic H1N1 (pdmH1N1) influenza virus infection were analyzed by digital gene expression and solid support microarrays. As expected during primary infection, innate immune responses were triggered in the lung tissue; meanwhile, in the lymphoid tissue, genes encoding antigen presentation and maturation of effector cells of adaptive immunity increased dramatically. After 5 days postinfection, the innate immune gene expression was replaced by the adaptive immune response, which correlates with viral clearance. Reinfection with homologous pandemic influenza virus resulted in a diminished innate immune response, early adaptive immune gene regulation, and a reduction in clinical severity. The fully annotated ferret infectome will be a critical aid to the understanding of the molecular events that regulate disease severity and host-influenza virus interactions among seasonal, pandemic, and highly pathogenic avian influenzas.     

Testosterone and cortisol concentrations vary with reproductive status in wild female red deer

Although hormones are key regulators of many fitness and life history traits, the causes of individual level variation in hormones, particularly in wild systems, remain understudied. Whilst we know that androgen and glucocorticoid levels vary within and among individuals in mammalian populations, how this relates to key reproductive processes such as gestation and lactation, and their effects on a female''s measurable hormone levels are poorly understood in wild systems. Using fecal samples collected from females in a wild red deer population between 2001 and 2013, we explore how fecal androgen (FAM) and cortisol (FCM) metabolite concentrations change with age and season, and how individual differences relate to variation in reproductive state. Both FAM and FCM levels increase toward parturition, although this only affects FCM levels in older females. FCM levels are also higher when females suckle a male rather than a female calf, possibly due to the higher energetic costs of raising a son. This illustrates the importance of accounting for a female''s life history and current reproductive status, as well as temporal variation, when examining individual differences in hormone levels. We discuss these findings in relation to other studies of mammalian systems and in particular to the relatively scarce information on variation in natural levels of hormones in wild populations.     

Gender-dependent modulation of alpha 1-adrenergic responses in rat mesenteric arteries

The purpose of this study was to test the hypothesis that pathways modulating vasoconstriction in rat mesenteric resistance arteries are gender dependent. Net contractile responses to phenylephrine were significantly increased by endothelium disruption in arteries from males but not females. This gender-dependent effect was stimulus specific, because disruption of endothelium increased reactivity to serotonin comparably in arteries from both genders. Ovariectomy unmasked an increase in net alpha(1)-adrenergic contractile responsiveness after endothelium disruption, suggesting alpha(1)-adrenergic-stimulated production of endothelial vasodilators is suppressed in control females by gonadal sex steroids. Production of modulatory endothelium-derived vasodilators in males is balanced by production of vasoconstricting arachidonic acid metabolites. This was revealed by decreased alpha(1)-adrenergic contractile responses in arteries from males after pretreatment with indomethacin or the cyclooxygenase-1 selective inhibitor SC-560. The indomethacin-induced effect persisted after endothelium disruption, indicating smooth muscle as the source of cyclooxygenase-1-derived vasoconstrictors and was attenuated after orchiectomy. This study indicates gender differences in the expression of two pathways modulating alpha(1)-adrenergic sensitivity in mesenteric arteries: an endothelium-dependent vasodilator pathway and a balancing smooth muscle cyclooxygenase-1-dependent vasoconstrictor pathway. One consequence of these differences is that endothelial damage produces a selective increase in alpha(1)-adrenergic agonist reactivity in arteries from males.     

Structure and axon outgrowth inhibitor binding of the Nogo-66 receptor and related proteins

The myelin-derived proteins Nogo, MAG and OMgp limit axonal regeneration after injury of the spinal cord and brain. These cell-surface proteins signal through multi-subunit neuronal receptors that contain a common ligand-binding glycosylphosphatidylinositol-anchored subunit termed the Nogo-66 receptor (NgR). By deletion analysis, we show that the binding of soluble fragments of Nogo, MAG and NgR to cell-surface NgR requires the entire leucine-rich repeat (LRR) region of NgR, but not other portions of the protein. Despite sharing extensive sequence similarity with NgR, two related proteins, NgR2 and NgR3, which we have identified, do not bind Nogo, MAG, OMgp or NgR. To investigate NgR specificity and multi-ligand binding, we determined the crystal structure of the biologically active ligand-binding soluble ectodomain of NgR. The molecule is banana shaped with elongation and curvature arising from eight LRRs flanked by an N-terminal cap and a small C-terminal subdomain. The NgR structure analysis, as well as a comparison of NgR surface residues not conserved in NgR2 and NgR3, identifies potential protein interaction sites important in the assembly of a functional signaling complex.