Variation in the Gene TAS2R13 is Associated with Differences in Alcohol Consumption in Patients with Head and Neck Cancer

Variation in responsiveness to bitter-tasting compounds has been associated with differences in alcohol consumption. One strong genetic determinant of variation in bitter taste sensitivity is alleles of the TAS2R gene family, which encode chemosensory receptors sensitive to a diverse array of natural and synthetic compounds. Members of the TAS2R family, when expressed in the gustatory system, function as bitter taste receptors. To better understand the relationship between TAS2R function and alcohol consumption, we asked if TAS2R variants are associated with measures of alcohol consumption in a head and neck cancer patient cohort. Factors associated with increased alcohol intake are of strong interest to those concerned with decreasing the incidence of cancers of oral and pharyngeal structures. We found a single nucleotide polymorphism (SNP) located within the TAS2R13 gene (rs1015443 [C1040T, Ser259Asn]), which showed a significant association with measures of alcohol consumption assessed via the Alcohol Use Disorders Identification Test (AUDIT). Analyses with other SNPs in close proximity to rs1015443 suggest that this locus is principally responsible for the association. Thus, our results provide additional support to the emerging hypothesis that genetic variation in bitter taste receptors can impact upon alcohol consumption.     

Aurapex penicillata gen. sp. nov. from native Miconia theaezans and Tibouchina spp. in Colombia

Conidiomata of a fungus resembling Chrysoporthe cubensis, a serious canker pathogen of Eucalyptus spp. (Myrtaceae, Myrtales) in tropical and subtropical parts of the world, was found on Eucalyptus grandis in Colombia. Fruiting structures of the fungus could be distinguished from those of C. cubensis by their distinctly orange conidiomatal necks. This fungus also was found on several plant species native to Colombia including Tibouchina urvilleana, T. lepidota and Miconia theaezans (Melastomataceae, Myrtales). Morphological comparisons, as well as those based on sequences of the ITS1/ITS2 region of the ribosomal DNA repeat and the beta-tubulin gene, were used to characterize this fungus. Its pathogenicity was assessed on various plants from which it has been collected, either in field or greenhouse trials. Phylogenetic analyses showed that isolates reside in a clade distinct from the four clades accommodating Chrysoporthe, Cryphonectria, Endothia and Rostraureum. Members of this clade are distinguished by the presence of orange conidiomatal necks with black bases and a unique internal stromatal structure. No teleomorph has been found for this fungus, for which we have provided the name Aurapex penicillata gen. sp. nov. A. penicillata produced only small lesions after inoculation on young T. urvilleana, M. theaezans and E. grandis trees and appears not to be a serious pathogen.     

NMR backbone resonance assignments of the prodomain variants of BDNF in the urea denatured state

Brain derived neurotrophic factor (BDNF) is a member of the neurotrophin family of proteins which plays a central role in neuronal survival, growth, plasticity and memory. A single Val66Met variant has been identified in the prodomain of human BDNF that is associated with anxiety, depression and memory disorders. The structural differences within the full-length prodomain Val66 and Met66 isoforms could shed light on the mechanism of action of the Met66 and its impact on the development of neuropsychiatric-associated disorders. In the present study, we report the backbone ¹H, ¹³C, and ¹⁵N NMR assignments of both full-length Val66 and Met66 prodomains in the presence of 2 M urea. These conditions were utilized to suppress residual structure and aid subsequent native state structural investigations aimed at mapping and identifying variant-dependent conformational differences under native-state conditions.     

Effects of ATP depletion and phosphate analogues on P-glycoprotein conformation in live cells.

P-glycoprotein (Pgp), a membrane pump often responsible for the multidrug resistance of cancer cells, undergoes conformational changes in the presence of substrates/modulators, or upon ATP depletion, reflected by its enhanced reactivity with the UIC2 monoclonal antibody. When the UIC2-shift was elicited by certain modulators (e.g. cyclosporin A or vinblastine, but not with verapamil or Tween 80), the subsequent binding of other monoclonal anti-Pgp Ig sharing epitopes with UIC2 (e.g. MM12.10) was abolished [Nagy, H., Goda, K., Arceci, R., Cianfriglia, M., Mechetner, E. & Szabó Jr, G. (2001) Eur. J. Biochem. 268, 2416-2420]. To further study the relationship between UIC2-shift and the suppression of MM12.10 binding, we compared, on live cells, how ATP depletion and treatment of cells with phosphate analogues (sodium orthovanadate, beryllium fluoride and fluoro-aluminate) that trap nucleotides at the catalytic site, affect the two phenomena. Similarly to modulators or ATP depleting agents, all the phosphate analogues increased daunorubicin accumulation in Pgp-expressing cells. Prelabeling of ATP depleted cells with UIC2 completely abolished the subsequent binding of MM12.10, in accordance with the enhanced binding of the first mAb. Vanadate and beryllium fluoride, but not fluoro-aluminate, reversed the effect of cyclosporin A, preventing UIC2 binding and allowing for labeling of cells with MM12.10. Thus, changes in UIC2 reactivity are accompanied by complementary changes in MM12.10 binding also in response to direct modulation of the ATP-binding site, confirming that conformational changes intrinsic to the catalytic cycle are reflected by both UIC2-related phenomena. These data also fit a model where the UIC2 epitope is available for antibody binding throughout the catalytic cycle including the step of ATP binding, to become unavailable only in the catalytic transition state.     

Structural and Genetic Basis for the Serological Differentiation of Pasteurella multocida Heddleston Serotypes 2 and 5

Pasteurella multocida is classified into 16 serotypes according to the Heddleston typing scheme. As part of a comprehensive study to define the structural and genetic basis of this scheme, we have determined the structure of the lipopolysaccharide (LPS) produced by P. multocida strains M1404 (B:2) and P1702 (E:5), the type strains for serotypes 2 and 5, respectively. The only difference between the LPS structures made by these two strains was the absence of a phosphoethanolamine (PEtn) moiety at the 3 position of the second heptose (Hep II) in M1404. Analysis of the lpt-3 gene, required for the addition of this PEtn residue, revealed that the gene was intact in P1702 but contained a nonsense mutation in M1404. Expression of an intact copy of lpt-3 in M1404 resulted in the attachment of a PEtn residue to the 3 position of the Hep II residue, generating an LPS structure identical to that produced by P1702. We identified and characterized each of the glycosyltransferase genes required for assembly of the serotype 2 and 5 LPS outer core. Monoclonal antibodies raised against serotype 2 LPS recognized the serotype 2/5-specific outer core LPS structure, but recognition of this structure was inhibited by the PEtn residue on Hep II. These data indicate that the serological classification of strains into Heddleston serotypes 2 and 5 is dependent on the presence or absence of PEtn on Hep II.Pasteurella multocida is a gram-negative pathogen that causes serious diseases in animals and humans. It is the causative agent of fowl cholera (7), hemorrhagic septicemia in cattle (9), atrophic rhinitis in pigs (6), and dog and cat bite infections in humans (28).P. multocida isolates may be grouped serologically based on capsular antigens into five serogroups—A, B, D, E, and F—using a passive hemagglutination test with erythrocytes sensitized with capsular antigen. Structural information is available for the capsular polysaccharides synthesized by serogroups A (hyaluronic acid) (22), D (heparin) (10), and F (chondroitin) (10). The genes involved in biosynthesis of the capsules have been identified for all five serogroups (27), and capsule is a critical virulence factor for serogroups A (8) and B (3).Lipopolysaccharide (LPS) is also an important virulence factor in P. multocida (13) and can be used for the identification of strains, with two main somatic typing systems reported (14, 17). The Namioka system is based on a tube agglutination test and is able to recognize 11 serotypes (17), whereas the Heddleston system uses a gel diffusion precipitation test and can recognize 16 serotypes; the Heddleston system is currently the preferred method (14). Current classification of P. multocida strains combines capsular typing with Heddleston somatic typing. Strains are given a designation in which the first letter indicates the capsular group and the number designates the Heddleston LPS serotype (e.g., A:1 indicates a strain that is capsular group A and LPS serotype 1). LPS produced by each of the 16 Heddleston serotype strains has been examined previously for sugar content and reactivity with LPS antisera (21). The LPS isolated from serotype 2 and 5 strains was virtually identical in sodium dodecyl sulfate-polyacrylamide gel electrophoresis migration profile (19), sugar composition, and serological reactivity with anti-LPS antibodies (21). Interestingly, serotypes 2 and 5 were the only serotypes found to elaborate two isomers of heptose in their LPS, namely l-glycero-d-manno-heptose (ld-Hep) and d-glycero-d-manno-heptose (dd-Hep) (21). The aims of this study were to determine whether the LPS molecules made by these two serotypes were structurally distinct and to compare the LPS structures with those previously determined for P. multocida serotypes 1 and 3 (24-26). Furthermore, we identified the transferase genes responsible for the assembly of the outer core LPS structure in each of these strains and characterized the function of each glycosyltransferase.     

C-Terminal Fragments of Amyloid-β Peptide Cause Cholinergic Axonal Degeneration by a Toxic Effect Rather Than by Physical Injury in the Nondemented Human Brain

Previous experimental studies have indicated that amyloid-b peptide (A) may cause axonal degeneration in the brain of individuals with Alzheimer's disease (AD) by physical injury, mass lesion, or membrane perturbation. In this study, acetylcholinesterase histochemical, and A and tau immunohistochemical double-staining were performed in nondemented elderly human hippocampal and entorhinal brain samples, to demonstrate the presence of dystrophic neurites caused by the C-terminal or N-terminal fragments of A. The early interactions between the A-stained senile plaques (SPs) and the enzyme-positive axons were investigated. The double-stained samples revealed that A deposition occurs first, followed by the development of cholinergic axonal damage. Most of the dystrophic axonal processes are incorporated in the peripheral area of the SPs and are positive for phosphorylated tau [pS²⁰²] and tau-5. The result suggests that C-terminal fragments are more harmful than N-terminal fragments of A and may induce the development of dystrophic neurites by a toxic effect rather than by physical injury.     

N-Terminal deletions modify the Cu2+ binding site in amyloid-beta

Copper is implicated in the in vitro formation and toxicity of Alzheimer's disease amyloid plaques containing the beta-amyloid (Abeta) peptide (Bush, A. I., et al. (2003) Proc. Natl. Acad. Sci. U.S.A. 100, 11934). By low temperature electron paramagnetic resonance (EPR) spectroscopy, the importance of the N-terminus in creating the Cu(2+) binding site in native Abeta has been examined. Peptides that contain the proposed binding site for Cu(2+)-three histidines (H6, H13, and H14) and a tyrosine (Y10)-but lack one to three N-terminal amino acids, do not bind Cu(2+) in the same coordination environment as the native peptide. EPR spectra of soluble Abeta with stoichiometric amounts of Cu(2+) show type 2 Cu(2+) EPR spectra for all peptides. The ligand donor atoms to Cu(2+) are 3N1O when Cu(2+) is bound to any of the Abetapeptides (Abeta16, Abeta28, Abeta40, and Abeta42) that contain the first 16 amino acids of full-length Abeta. When a Y10F mutant of Abeta is used, the coordination environment for Cu(2+) remains 3N1O and Cu(2+) EPR spectra of this mutant are identical to the wild-type spectra. Isotopic labeling experiments show that water is not the O-atom donor to Cu(2+) in Abeta fibrils or in the Y10F mutant. Further, we find that Cu(2+) cannot be removed from Cu(2+)-containing fibrils by washing with buffer, but that Cu(2+) binds to fibrils initially assembled without Cu(2+) in the same coordination environment as in fibrils assembled with Cu(2+). Together, these results indicate (1) that the O-atom donor ligand to Cu(2+) in Abeta is not tyrosine, (2) that the native Cu(2+) binding site in Abeta is sensitive to small changes at the N-terminus, and (3) that Cu(2+) binds to Abetafibrils in a manner that permits exchange of Cu(2+) into and out of the fibrillar architecture.     

Radiofrequency and extremely low-frequency electromagnetic field effects on the blood-brain barrier

During the last century, mankind has introduced electricity and during the very last decades, the microwaves of the modern communication society have spread a totally new entity--the radiofrequency fields--around the world. How does this affect biology on Earth? The mammalian brain is protected by the blood-brain barrier, which prevents harmful substances from reaching the brain tissue. There is evidence that exposure to electromagnetic fields at non thermal levels disrupts this barrier. In this review, the scientific findings in this field are presented. The result is a complex picture, where some studies show effects on the blood-brain barrier, whereas others do not. Possible mechanisms for the interactions between electromagnetic fields and the living organisms are discussed. Demonstrated effects on the blood-brain barrier, as well as a series of other effects upon biology, have caused societal anxiety. Continued research is needed to come to an understanding of how these possible effects can be neutralized, or at least reduced. Furthermore, it should be kept in mind that proven effects on biology also should have positive potentials, e.g., for medical use.     

Land-use effects on structural and functional composition of benthic and leaf-associated macroinvertebrates in four Andean streams

The replacement of native forests by pastures takes place widely in the Andes. The effects of such land-use change on aquatic assemblages are poorly understood. We conducted a comparative analysis of the effects of forest conversion to pastures on the taxonomic, structural, and functional composition of macroinvertebrates (benthic and leaf-associated) in montane and upper montane streams (ecosystem type) of the south Ecuadorian Andes. Taxonomic composition of benthic and leaf-associated macroinvertebrates was different between ecosystem type and land use. Also, major differences in the structural and functional composition of benthic and leaf-associated macroinvertebrates were mainly promoted by land use in both ecosystem types. Forested streams showed higher diversity than pasture streams, sustaining more shredder, scraper, and predatory invertebrates. We also observed differences in the macroinvertebrate communities between benthic and leaf-bag samples. Leaf bags had lower diversity and more collector invertebrates than benthic samples. This study highlights the large effect of riparian forest conversion to pasture land on macroinvertebrate communities, and the importance of using appropriate sampling techniques to characterize aquatic assemblages. We also recommend the maintenance and restoration of riparian vegetation to mitigate the effects of deforestation on stream communities and ecosystem processes.