A southern California freeway is a physical and social barrier to gene flow in carnivores

Roads present formidable barriers to dispersal. We examine movements of two highly mobile carnivores across the Ventura Freeway near Los Angeles, one of the busiest highways in the United States. The two species, bobcats and coyotes, can disappear from habitats isolated and fragmented by roads, and their ability to disperse across the Ventura Freeway tests the limits of vertebrates to overcome anthropogenic obstacles. We combine radio-telemetry data and genetically based assignments to identify individuals that have crossed the freeway. Although the freeway is a significant barrier to dispersal, we find that carnivores can cross the freeway and that 5-32% of sampled carnivores crossed over a 7-year period. However, despite moderate levels of migration, populations on either side of the freeway are genetically differentiated, and coalescent modelling shows their genetic isolation is consistent with a migration fraction less than 0.5% per generation. These results imply that individuals that cross the freeway rarely reproduce. Highways and development impose artificial home range boundaries on territorial and reproductive individuals and hence decrease genetically effective migration. Further, territory pile-up at freeway boundaries may decrease reproductive opportunities for dispersing individuals that do manage to cross. Consequently, freeways are filters favouring dispersing individuals that add to the migration rate but little to gene flow. Our results demonstrate that freeways can restrict gene flow even in wide-ranging species and suggest that for territorial animals, migration levels across anthropogenic barriers need to be an order of magnitude larger than commonly assumed to counteract genetic differentiation.     

Small molecules that enhance the catalytic efficiency of HLA-DM

HLA-DM (DM) plays a critical role in Ag presentation to CD4 T cells by catalyzing the exchange of peptides bound to MHC class II molecules. Large lateral surfaces involved in the DM:HLA-DR (DR) interaction have been defined, but the mechanism of catalysis is not understood. In this study, we describe four small molecules that accelerate DM-catalyzed peptide exchange. Mechanistic studies demonstrate that these small molecules substantially enhance the catalytic efficiency of DM, indicating that they make the transition state of the DM:DR/peptide complex energetically more favorable. These compounds fall into two functional classes: two compounds are active only in the presence of DM, and binding data for one show a direct interaction with DM. The remaining two compounds have partial activity in the absence of DM, suggesting that they may act at the interface between DM and DR/peptide. A hydrophobic ridge in the DMbeta1 domain was implicated in the catalysis of peptide exchange because the activity of three of these enhancers was substantially reduced by point mutations in this area.     

IL-4 and IL-13 form a negative feedback circuit with surfactant protein-D in the allergic airway response

The innate immune molecule surfactant protein-D (SP-D) plays an important regulatory role in the allergic airway response. In this study, we demonstrate that mice sensitized and challenged with either Aspergillus fumigatus (Af) or OVA have increased SP-D levels in their lung. SP-D mRNA and protein levels in the lung also increased in response to either rIL-4 or rIL-13 treatment. Type II alveolar epithelial cell expression of IL-4Rs in mice sensitized and challenged with Af, and in vitro induction of SP-D mRNA and protein by IL-4 and IL-13, but not IFN-gamma, suggested a direct role of IL-4R-mediated events. The regulatory function of IL-4 and IL-13 was further supported in STAT-6-deficient mice as well as in IL-4/IL-13 double knockout mice that failed to increase SP-D production upon allergen challenge. Interestingly, addition of rSP-D significantly inhibited Af-driven Th2 cell activation in vitro whereas mice lacking SP-D had increased numbers of CD4(+) cells with elevated IL-13 and thymus- and activation-regulated chemokine levels in the lung and showed exaggerated production of IgE and IgG1 following allergic sensitization. We propose that allergen exposure induces elevation in SP-D protein levels in an IL-4/IL-13-dependent manner, which in turn, prevents further activation of sensitized T cells. This negative feedback regulatory circuit could be essential in protecting the airways from inflammatory damage after allergen inhalation.     

FGF21 N- and C-termini play different roles in receptor interaction and activation

Fibroblast growth factor-21 (FGF21) signaling requires the presence of β-Klotho, a co-receptor with a very short cytoplasmic domain. Here we show that FGF21 binds directly to β-Klotho through its C-terminus. Serial C-terminal truncations of FGF21 weakened or even abrogated its interaction with β-Klotho in a Biacore assay, and led to gradual loss of potency in a luciferase reporter assay but with little effect on maximal response. In contrast, serial N-terminal truncations of FGF21 had no impact on β-Klotho binding. Interestingly, several of them exhibited characteristics of partial agonists with minimal effects on potency. These data demonstrate that the C-terminus of FGF21 is critical for binding to β-Klotho and the N-terminus is critical for fibroblast growth factor receptor (FGFR) activation.

Structured summary

MINT-6799939: FGFR1c (uniprotkb:P11362) binds (MI:0407) to β-Klotho (uniprotkb: Q86Z14) by surface plasmon resonance (MI:0107)MINT-6799907, MINT-6799922: FGF21 (uniprotkb: Q9NSA1) binds (MI:0407) to β-Klotho (uniprotkb: Q86Z14) by surface plasmon resonance (MI:0107)     

Crystal Structure of Iodotyrosine Deiodinase, a Novel Flavoprotein Responsible for Iodide Salvage in Thyroid Glands

The flavoprotein iodotyrosine deiodinase (IYD) salvages iodide from mono- and diiodotyrosine formed during the biosynthesis of the thyroid hormone thyroxine. Expression of a soluble domain of this membrane-bound enzyme provided sufficient material for crystallization and characterization by x-ray diffraction. The structures of IYD and two co-crystals containing substrates, mono- and diiodotyrosine, alternatively, were solved at resolutions of 2.0, 2.45, and 2.6 Å, respectively. The structure of IYD is homologous to others in the NADH oxidase/flavin reductase superfamily, but the position of the active site lid in IYD defines a new subfamily within this group that includes BluB, an enzyme associated with vitamin B₁₂ biosynthesis. IYD and BluB also share key interactions involving their bound flavin mononucleotide that suggest a unique catalytic behavior within the superfamily. Substrate coordination to IYD induces formation of an additional helix and coil that act as an active site lid to shield the resulting substrate·flavin complex from solvent. This complex is stabilized by aromatic stacking and extensive hydrogen bonding between the substrate and flavin. The carbon-iodine bond of the substrate is positioned directly over the C-4a/N-5 region of the flavin to promote electron transfer. These structures now also provide a molecular basis for understanding thyroid disease based on mutations of IYD.The micronutrient iodide is essential for the biosynthesis of thyroxine (3,3′,5,5′-tetraiodothyronine), a hormone used by a wide range of organisms as a master control of metabolic rate. In mammals, iodide homeostasis in the thyroid gland is critical for generating thyroxine and is achieved by sequestering and salvaging iodide. Both of these functions are critical for human health, and congenital defects in either may lead to hypothyroidism (1, 2). Sequestration of iodide from the circulatory system is accomplished by a Na⁺/I⁻ symporter located in the plasma membrane of thyroid follicular cells (2). Salvage of iodide is accomplished by iodotyrosine deiodinase (IYD)³ located in the apical plasma membrane surrounding the thyroid colloid in which thyroglobulin is stored and processed (3). Proteolysis of mature thyroglobulin releases thyroxine as well as mono- and diiodotyrosine (MIT and DIT, respectively). IYD catalyzes a reductive deiodination of MIT and DIT selectively to prevent loss of iodide that would otherwise occur by excretion of these amino acids. The gene encoding IYD has recently been identified (3, 4) and has provided an initial basis for correlating its mutation with hypothyroidism and goiter observed in certain patients (1). The crystal structure described in this work now supersedes the previous structural models.IYD represents one of only two enzymes known to promote reductive dehalogenation in mammals (Fig. 1). The other enzyme, iodothyronine deiodinase, acts alternatively to activate and deactivate thyroxine by deiodinating the outer or inner ring, respectively (5). Interestingly two distinct strategies based on two distinct protein architectures have been recruited for catalyzing essentially the same deiodination. Iodothyronine deiodinase is a member of the thioredoxin structural superfamily and utilizes an active site selenocysteine for its catalysis. In contrast, IYD is a member of the NADH oxidase/flavin reductase superfamily (3, 4) and requires neither selenocysteine nor cysteine for catalysis (6). Instead a bound flavin mononucleotide (FMN) is required (7). Flavin is integral to many catalytic functions and even certain biological halogenation reactions (8–11). However, its role in reductive dehalogenation is quite unusual and not yet characterized. Flavin has also been implicated in just one bacterial reductive dehalogenation, although little information is available on this additional system other than its use of flavin-adenine dinucleotide (FAD) rather than FMN (12).Open in a separate windowFIGURE 1.Reductive deiodination for iodide salvage catalyzed by IYD (A) and metabolism of the hormone thyroxine by iodothyronine deiodinase (ID) (B).Structural characterization of IYD is critical for identifying the active site properties that support its unique catalytic ability and substrate selectivity. Moreover this information contributes a molecular understanding of how mutations in IYD can disrupt function to cause iodide deficiency and hypothyroidism. Such mutants are particularly tragic because their detection often occurs only after developmental damage to patients has occurred (1). Studies on IYD to date have been limited by its membrane association and transient expression in mammalian cells (3, 4). However, deletion of its N-terminal membrane anchor yields a soluble and active enzyme (6) that has been expressed in Sf9 cells in sufficient quantities to support the crystallographic analysis reported here.     

Six months of disuse during hibernation does not increase intracortical porosity or decrease cortical bone geometry,strength, or mineralization in black bear (Ursus americanus) femurs

Disuse typically uncouples bone formation from resorption, leading to bone loss which compromises bone mechanical properties and increases the risk of bone fracture. Previous studies suggest that bears can prevent bone loss during long periods of disuse (hibernation), but small sample sizes have limited the conclusions that can be drawn regarding the effects of hibernation on bone structure and strength in bears. Here we quantified the effects of hibernation on structural, mineral, and mechanical properties of black bear (Ursus americanus) cortical bone by studying femurs from large groups of male and female bears (with wide age ranges) killed during pre-hibernation (fall) and post-hibernation (spring) periods. Bone properties that are affected by body mass (e.g. bone geometrical properties) tended to be larger in male compared to female bears. There were no differences (p>0.226) in bone structure, mineral content, or mechanical properties between fall and spring bears. Bone geometrical properties differed by less than 5% and bone mechanical properties differed by less than 10% between fall and spring bears. Porosity (fall: 5.5±2.2%; spring: 4.8±1.6%) and ash fraction (fall: 0.694±0.011; spring: 0.696±0.010) also showed no change (p>0.304) between seasons. Statistical power was high (>72%) for these analyses. Furthermore, bone geometrical properties and ash fraction (a measure of mineral content) increased with age and porosity decreased with age. These results support the idea that bears possess a biological mechanism to prevent disuse and age-related osteoporoses.     

The roles of host evolutionary relationships (genus: Nasonia) and development in structuring microbial communities

The comparative structure of bacterial communities among closely related host species remains relatively unexplored. For instance, as speciation events progress from incipient to complete stages, does divergence in the composition of the species’ microbial communities parallel the divergence of host nuclear genes? To address this question, we used the recently diverged species of the parasitoid wasp genus Nasonia to test whether the evolutionary relationships of their bacterial microbiotas recapitulate the Nasonia phylogenetic history. We also assessed microbial diversity in Nasonia at different stages of development to determine the role that host age plays in microbiota structure. The results indicate that all three species of Nasonia share simple larval microbiotas dominated by the γ‐proteobacteria class; however, bacterial species diversity increases as Nasonia develop into pupae and adults. Finally, under identical environmental conditions, the relationships of the microbial communities reflect the phylogeny of the Nasonia host species at multiple developmental stages, which suggests that the structure of an animal's microbial community is closely allied with divergence of host genes. These findings highlight the importance of host evolutionary relationships on microbiota composition and have broad implications for future studies of microbial symbiosis and animal speciation.     

Effect of ANXA2 gene single nucleotide polymorphism (SNP) on the development of osteonecrosis in Indian sickle cell patient: a PCR-RFLP approach

Osteonecrosis is a serious complication in sickle cell patients. The common sites of the necrosis are femoral head, head of the humerus and acetabulam. Annexin A2 (ANXA2) protein mainly functions in bone formation and bone resorption. Alteration of ANXA2 gene may affect the manifestations of osteonecrosis in the patients. PCR-RFLP is a common applicable technique for the detection of known mutation/polymorphisms. Here we are presenting application of the PCR-RFLP technique for determination of the ANXA2 gene single nucleotide polymorphism frequency and their clinical association among Indian sickle cell patients. Five known SNPs of ANXA2 gene (rs7170178, rs73435133, rs73418020, rs72746635 and rs73418025) were determined using the HpyCH4V, DdeI, HpyCH4III and Sau 961 restriction enzyme respectively. Restriction enzyme DdeI was common for rs73435133 and rs72746635 SNP. Only the rs7170178 SNP was detected among patient and control and the other four SNPs were absent in the studied groups. The frequency of ANXA2 gene rs7170178 SNP (A/G, G/G) was comparatively higher in sickle cell patients than controls and it was clinically associated with sickle cell osteonecrosis. The P value of heterozygotes (A/G) and homozygotes (G/G) genotypes were <0.001 and 0.001 respectively, which were highly significant. This study established the application of PCR-RFLP in detection of ANXA2 SNPs in sickle cell patients.     

Ultrasensitive Responses and Specificity in Cell Signaling

Background  

Interconnected cell signaling pathways are able to efficiently and accurately transmit a multitude of different signals, despite an inherent potential for undesirable levels of cross-talk. To ensure that an appropriate response is produced, biological systems have evolved network-level mechanisms that insulate pathways from crosstalk and prevent 'leaking' or 'spillover' between pathways. Many signaling pathways have been shown to respond in an ultrasensitive (switch-like) fashion to graded input, and this behavior may influence specificity. The relationship of ultrasensitivity to signaling specificity has not been extensively explored.