CB1 Cannabinoid Receptors Couple to Focal Adhesion Kinase to Control Insulin Release

Endocannabinoid signaling has been implicated in modulating insulin release from β cells of the endocrine pancreas. β Cells express CB₁ cannabinoid receptors (CB₁Rs), and the enzymatic machinery regulating anandamide and 2-arachidonoylglycerol bioavailability. However, the molecular cascade coupling agonist-induced cannabinoid receptor activation to insulin release remains unknown. By combining molecular pharmacology and genetic tools in INS-1E cells and in vivo, we show that CB₁R activation by endocannabinoids (anandamide and 2-arachidonoylglycerol) or synthetic agonists acutely or after prolonged exposure induces insulin hypersecretion. In doing so, CB₁Rs recruit Akt/PKB and extracellular signal-regulated kinases 1/2 to phosphorylate focal adhesion kinase (FAK). FAK activation induces the formation of focal adhesion plaques, multimolecular platforms for second-phase insulin release. Inhibition of endocannabinoid synthesis or FAK activity precluded insulin release. We conclude that FAK downstream from CB₁Rs mediates endocannabinoid-induced insulin release by allowing cytoskeletal reorganization that is required for the exocytosis of secretory vesicles. These findings suggest a mechanistic link between increased circulating and tissue endocannabinoid levels and hyperinsulinemia in type 2 diabetes.     

Insights into β-Lactamases from Burkholderia Species,Two Phylogenetically Related yet Distinct Resistance Determinants

Burkholderia cepacia complex and Burkholderia pseudomallei are opportunistic human pathogens. Resistance to β-lactams among Burkholderia spp. is attributable to expression of β-lactamases (e.g. PenA in B. cepacia complex and PenI in B. pseudomallei). Phylogenetic comparisons reveal that PenA and PenI are highly related. However, the analyses presented here reveal that PenA is an inhibitor-resistant carbapenemase, most similar to KPC-2 (the most clinically significant serine carbapenemase), whereas PenI is an extended spectrum β-lactamase. PenA hydrolyzes β-lactams with k_cat values ranging from 0.38 ± 0.04 to 460 ± 46 s⁻¹ and possesses high k_cat/k_inact values of 2000, 1500, and 75 for β-lactamase inhibitors. PenI demonstrates the highest k_cat value for cefotaxime of 9.0 ± 0.9 s⁻¹. Crystal structure determination of PenA and PenI reveals important differences that aid in understanding their contrasting phenotypes. Changes in the positioning of conserved catalytic residues (e.g. Lys-73, Ser-130, and Tyr-105) as well as altered anchoring and decreased occupancy of the deacylation water explain the lower k_cat values of PenI. The crystal structure of PenA with imipenem docked into the active site suggests why this carbapenem is hydrolyzed and the important role of Arg-220, which was functionally confirmed by mutagenesis and biochemical characterization. Conversely, the conformation of Tyr-105 hindered docking of imipenem into the active site of PenI. The structural and biochemical analyses of PenA and PenI provide key insights into the hydrolytic mechanisms of β-lactamases, which can lead to the rational design of novel agents against these pathogens.     

Early Gag Immunodominance of the HIV-Specific T-Cell Response during Acute/Early Infection Is Associated with Higher CD8+ T-Cell Antiviral Activity and Correlates with Preservation of the CD4+ T-Cell Compartment

The important role of the CD8⁺ T-cell response on HIV control is well established. Moreover, the acute phase of infection represents a proper scenario to delineate the antiviral cellular functions that best correlate with control. Here, multiple functional aspects (specificity, ex vivo viral inhibitory activity [VIA] and polyfunctionality) of the HIV-specific CD8⁺ T-cell subset arising early after infection, and their association with disease progression markers, were examined. Blood samples from 44 subjects recruited within 6 months from infection (primary HIV infection [PHI] group), 16 chronically infected subjects, 11 elite controllers (EC), and 10 healthy donors were obtained. Results indicated that, although Nef dominated the anti-HIV response during acute/early infection, a higher proportion of early anti-Gag T cells correlated with delayed progression. Polyfunctional HIV-specific CD8⁺ T cells were detected at early time points but did not associate with virus control. Conversely, higher CD4⁺ T-cell set points were observed in PHI subjects with higher HIV-specific CD8⁺ T-cell VIA at baseline. Importantly, VIA levels correlated with the magnitude of the anti-Gag cellular response. The advantage of Gag-specific cells may result from their enhanced ability to mediate lysis of infected cells (evidenced by a higher capacity to degranulate and to mediate VIA) and to simultaneously produce IFN-γ. Finally, Gag immunodominance was associated with elevated plasma levels of interleukin 2 (IL-2) and macrophage inflammatory protein 1β (MIP-1β). All together, this study underscores the importance of CD8⁺ T-cell specificity in the improved control of disease progression, which was related to the capacity of Gag-specific cells to mediate both lytic and nonlytic antiviral mechanisms at early time points postinfection.     

Interspecific relationships in co‐occurring populations of social parasites and their host ants

Myrmica ant colonies host numerous insect species, including the larvae of Maculinea butterflies and Microdon myrmicae hoverflies. Little is known about the interspecific relationships among these social parasites and their host ants occurring in sympatric populations. We investigated communities of social parasites to assess the strategies allowing them to share the same pool of resources (i.e. Myrmica colonies). The present study was carried out at five sites inhabited by different social parasite communities, each comprising varying proportions of Maculinea teleius, Maculinea nausithous, Maculinea alcon, and Microdon myrmicae. We investigated their spatial distributions, host segregation, the degree of chemical similarity between social parasites and hosts, and temporal overlaps in colony resource exploitation. Spatial segregation among social parasites was found in two populations and it arises from microhabitat preferences and biological interactions. Local conditions can drive selection on one social parasite to use a Myrmica host species that is not exploited by other social parasites. Myrmica scabrinodis and Myrmica rubra nests infested by larvae of two social parasite species were found and the most common co‐occurrence was between Ma. teleius and Mi. myrmicae. The successful coexistence of these two species derives from their exploitation of the host colony resources at different times of the year. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 699–709.     

Docosahexaenoic acid and tetracyclines as promising neuroprotective compounds with poly(ADP-ribose) polymerase inhibitory activities for oxidative/genotoxic stress treatment

The human genome is exposed to oxidative/genotoxic stress by several endogenous and exogenous compounds. These events evoke DNA damage and activate poly(ADP-ribose) polymerase-1 (PARP-1), the key enzyme involved in DNA repair. The massive stress and over-activation of this DNA-bound enzyme can be responsible for an energy crisis and neuronal death. The last data indicated that product of PARP-1, i.e. poly(ADP-ribose) (PAR), acts as a signalling molecule and plays a significant role in nucleus-mitochondria cross-talk. PAR translocated to the mitochondria can be involved in mitochondrial permeability, the release of an apoptosis-inducing factor (AIF). Its translocation into the nucleus leads to chromatin condensation, fragmentation and cell death. The exact mechanism of this novel death pathway has not yet fully been understood.     

Comparative histomorphology of intrinsic vibrissa musculature among primates: implications for the evolution of sensory ecology and “face touch”

Macrovibrissae are specialized tactile sensory hairs present in most mammalian orders, used in maxillary mechanoreception or “face touch.” Some mammals have highly organized vibrissae and are able to “whisk” them. Movement of vibrissae is influenced by intrinsic vibrissa musculature, striated muscle bands that attach directly to the vibrissa capsule. It is unclear if primates have organized vibrissae or intrinsic vibrissa musculature and it is uncertain if they can move their vibrissae. The present study used histomorphological techniques to compare vibrissae among 19 primates and seven non‐primate mammalian taxa. Upper lips of these mammals were sectioned and processed for histochemical analysis. While controlling for phylogenetic effects the following hypotheses were tested: 1) mammals with well‐organized vibrissae possess intrinsic vibrissa musculature and 2) intrinsic vibrissa musculature is best developed in nocturnal, arboreal taxa. Our qualitative analyses show that only arboreal, nocturnal prosimians possess intrinsic musculature. Not all taxa that possessed organized vibrissae had intrinsic vibrissa musculature. Phylogenetic comparative analyses revealed a 70% probability that stem mammals, primates, and haplorhines possessed intrinsic vibrissa musculature and well‐organized vibrissae. These two traits most likely coevolved according to a discrete phylogenetic analysis. These results indicate that nocturnal, arboreal primates have the potential to more actively use their vibrissae in spatial recognition and navigation tasks than diurnal, more terrestrial species, but there is a clear phylogenetic signal involved in the evolution of primate vibrissae and “face touch.” Am J Phys Anthropol, 2013. © 2012 Wiley Periodicals, Inc.     

Global Analysis of DNA Methylation Variation in Adipose Tissue from Twins Reveals Links to Disease-Associated Variants in Distal Regulatory Elements

Epigenetic modifications such as DNA methylation play a key role in gene regulation and disease susceptibility. However, little is known about the genome-wide frequency, localization, and function of methylation variation and how it is regulated by genetic and environmental factors. We utilized the Multiple Tissue Human Expression Resource (MuTHER) and generated Illumina 450K adipose methylome data from 648 twins. We found that individual CpGs had low variance and that variability was suppressed in promoters. We noted that DNA methylation variation was highly heritable (h²_median = 0.34) and that shared environmental effects correlated with metabolic phenotype-associated CpGs. Analysis of methylation quantitative-trait loci (metQTL) revealed that 28% of CpGs were associated with nearby SNPs, and when overlapping them with adipose expression quantitative-trait loci (eQTL) from the same individuals, we found that 6% of the loci played a role in regulating both gene expression and DNA methylation. These associations were bidirectional, but there were pronounced negative associations for promoter CpGs. Integration of metQTL with adipose reference epigenomes and disease associations revealed significant enrichment of metQTL overlapping metabolic-trait or disease loci in enhancers (the strongest effects were for high-density lipoprotein cholesterol and body mass index [BMI]). We followed up with the BMI SNP rs713586, a cg01884057 metQTL that overlaps an enhancer upstream of ADCY3, and used bisulphite sequencing to refine this region. Our results showed widespread population invariability yet sequence dependence on adipose DNA methylation but that incorporating maps of regulatory elements aid in linking CpG variation to gene regulation and disease risk in a tissue-dependent manner.     

Analysis of Age-Related Global DNA Methylation in Chicken

DNA methylation is an epigenetic modification that plays an important role in the normal development and function of organisms. The level of DNA methylation is species-, tissue-, and organelle-specific, and the methylation pattern is determined during embryogenesis. DNA methylation has also been correlated with age. The aim of this study was to determine the global DNA methylation levels and their correlation with age in the chicken, using a Polish autosexing chicken breed, Polbar. A quantitative technique based on an immunoenzymatic assay was used for global DNA methylation analysis. The results show increased global DNA methylation levels with older Polbar embryos. Global DNA methylation levels decrease with the age of hens in the postembryonic stage. This study expands the current knowledge of the Polbar epigenome and the general knowledge of the function of epigenetic mechanisms in birds.