DLA-DRB1, DQA1, and DQB1 alleles and haplotypes in North American Gray Wolves

The canine major histocompatibility complex contains highly polymorphic genes, many of which are critical in regulating immune response. Since domestic dogs evolved from Gray Wolves (Canis lupus), common DLA class II alleles should exist. Sequencing was used to characterize 175 Gray Wolves for DLA class II alleles, and data from 1856 dogs, covering 85 different breeds of mostly European origin, were available for comparison. Within wolves, 28 new alleles were identified, all occurring in at least 2 individuals. Three DLA-DRB1, 8 DLA-DQA1, and 6 DLA-DQB1 alleles also identified in dogs were present. Twenty-eight haplotypes were identified, of which 2 three-locus haplotypes, and many DLA-DQA1/DQB1 haplotypes, are also found in dogs. The wolves studied had relatively few dog DLA alleles and may therefore represent a remnant population descended from Asian wolves. The single European wolf included carried a haplotype found in both these North American wolves and in many dog breeds. Furthermore, one wolf DQB1 allele has been found in Shih Tzu, a breed of Asian origin. These data suggest that the wolf ancestors of Asian and European dogs may have had different gene pools, currently reflected in the DLA alleles present in dog breeds.     

Functional specialization of mouse higher visual cortical areas

The mouse is emerging as an important model for understanding how sensory neocortex extracts cues to guide behavior, yet little is known about how these cues are processed beyond primary cortical areas. Here, we used two-photon calcium imaging in awake mice to compare visual responses in primary visual cortex (V1) and in two downstream target areas, AL and PM. Neighboring V1 neurons had diverse stimulus preferences spanning five octaves in spatial and temporal frequency. By contrast, AL and PM neurons responded best to distinct ranges of stimulus parameters. Most strikingly, AL neurons preferred fast-moving stimuli while PM neurons preferred slow-moving stimuli. By contrast, neurons in V1, AL, and PM demonstrated similar selectivity for stimulus orientation but not for stimulus direction. Based on these findings, we predict that area AL helps guide behaviors involving fast-moving stimuli (e.g., optic flow), while area PM?helps guide behaviors involving slow-moving objects.     

Translation initiator EIF4G1 mutations in familial Parkinson disease

Genome-wide analysis of a multi-incident family with autosomal-dominant parkinsonism has implicated a locus on chromosomal region 3q26-q28. Linkage and disease segregation is explained by a missense mutation c.3614G>A (p.Arg1205His) in eukaryotic translation initiation factor 4-gamma (EIF4G1). Subsequent sequence and genotype analysis identified EIF4G1 c.1505C>T (p.Ala502Val), c.2056G>T (p.Gly686Cys), c.3490A>C (p.Ser1164Arg), c.3589C>T (p.Arg1197Trp) and c.3614G>A (p.Arg1205His) substitutions in affected subjects with familial parkinsonism and idiopathic Lewy body disease but not in control subjects. Despite different countries of origin, persons with EIF4G1 c.1505C>T (p.Ala502Val) or c.3614G>A (p.Arg1205His) mutations appear to share haplotypes consistent with ancestral founders. eIF4G1 p.Ala502Val and p.Arg1205His disrupt eIF4E or eIF3e binding, although the wild-type protein does not, and render mutant cells more vulnerable to reactive oxidative species. EIF4G1 mutations implicate mRNA translation initiation in familial parkinsonism and highlight a convergent pathway for monogenic, toxin and perhaps virally-induced Parkinson disease.     

Induction of autoantibodies to CCR5 in macaques and subsequent effects upon challenge with an R5-tropic simian/human immunodeficiency virus

Antibodies against CCR5, the major coreceptor for human immunodeficiency virus type 1 (HIV-1), may have antiviral potential as viral fusion inhibitors. In this study, we generated a virus-like particle (VLP)-based vaccine that effectively breaks B-cell tolerance and elicits autoantibodies against CCR5 in pig-tailed macaques. Initial studies in mice identified a polypeptide comprising the N-terminal domain of pig-tailed macaque CCR5 fused to streptavidin that, when conjugated at high density to bovine papillomavirus major capsid protein L1 VLPs, induced high-titer immunoglobulin G (IgG) that bound to a macaque CCR5-expressing cell line in vitro. In macaques, CCR5 peptide-conjugated VLP preparations induced high-avidity anti-CCR5 IgG autoantibody responses, and all five immunized macaques generated IgG that could block infection of CCR5-tropic simian/human immunodeficiency virus SHIV(SF162P3) in vitro. Although the anti-CCR5 IgG titers declined with time, autoantibody levels were boosted upon revaccination. Vaccinated macaques remained healthy for a period of over 3 years after the initial immunization, and no decline in the number of CCR5-expressing T cells was detected. To test the prophylactic efficacy of CCR5 autoantibodies, immunized macaques were challenged with SHIV(SF162P3). Although the plasma-associated virus in half of six control macaques declined to undetectable levels, viral loads were lower, declined more rapidly, and eventually became undetectable in all five macaques in which CCR5 autoantibodies had been elicited. In addition, in the four vaccinated macaques with higher autoantibody titers, viral loads and time to control of viremia were significantly decreased relative to controls, indicating the possibility that CCR5 autoantibodies contributed to the control of viral replication.     

Characterization of the Butyrivibrio fibrisolvens glgB gene, which encodes a glycogen-branching enzyme with starch-clearing activity.

A Butyrivibrio fibrisolvens H17c glgB gene, was isolated by direct selection for colonies that produced clearing on starch azure plates. The gene was expressed in Escherichia coli from its own promoter. The glgB gene consisted of an open reading frame of 1,920 bp encoding a protein of 639 amino acids (calculated Mr, 73,875) with 46 to 50% sequence homology with other branching enzymes. A limited region of 12 amino acids showed sequence similarity to amylases and glucanotransferases. The B. fibrisolvens branching enzyme was not able to hydrolyze starch but stimulated phosphorylase alpha-mediated incorporation of glucose into alpha-1,4-glucan polymer 13.4-fold. The branching enzyme was purified to homogeneity by a simple two-step procedure; N-terminal sequence and amino acid composition determinations confirmed the deduced translational start and amino acid sequence of the open reading frame. The enzymatic properties of the purified enzyme were investigated. The enzyme transferred chains of 5 to 10 (optimum, 7) glucose units, using amylose and amylopetin as substrates, to produce a highly branched polymer.     

Characterization of the Aspergillus nidulans septin (asp) gene family

Members of the septin gene family are involved in cytokinesis and the organization of new growth in organisms as diverse as yeast, fruit fly, worm, mouse, and human. Five septin genes have been cloned and sequenced from the model filamentous fungus A. nidulans. As expected, the A. nidulans septins contain the highly conserved GTP binding and coiled-coil domains seen in other septins. On the basis of hybridization of clones to a chromosome-specific library and correlation with an A. nidulans physical map, the septins are not clustered but are scattered throughout the genome. In phylogenetic analysis most fungal septins could be grouped with one of the prototypical S. cerevisiae septins, Cdc3, Cdc10, Cdc11, and Cdc12. Intron-exon structure was conserved within septin classes. The results of this study suggest that most fungal septins belong to one of four orthologous classes.     

Cell-cell and cell-surface interactions mediated by cellulose and a novel exopolysaccharide contribute to Pseudomonas putida biofilm formation and fitness under water-limiting conditions

The composition of the exopolysaccharide matrix of Pseudomonas putida mt2 biofilms is relatively undefined as well as the contributions of each polymer to ecological fitness. Here, we describe the role of two putative exopolysaccharide gene clusters, putida exopolysaccharide A (pea) and bacterial cellulose (bcs) in biofilm formation and stability, rhizosphere colonization and matrix hydration under water-limiting conditions. Our findings suggest that pea is involved in the production of a novel glucose, galactose, and mannose-rich polymer that contributes to cell-cell interactions necessary for pellicle and biofilm formation and stability. In contrast, Bcs plays a minor role in biofilm formation and stability, although it does contribute to rhizosphere colonization based on a competition assay. We show that pea expression is highly induced transiently under water-limiting conditions but only slightly by high osmolarity, as determined by qRT-PCR. In contrast, both forms of water stress highly induced bcs expression. Cells deficient in making one or more exopolysaccharide experienced greater dehydration-mediated cell-envelope stress, leading to increased alginate promoter activity. However, this did not lead to increased exopolysaccharide production, except in bcs or pea mutants unable to produce alginate, indicating that P. putida compensates by producing, presumably more Pea or Bcs exopolysaccharides, to facilitate biofilm hydration. Collectively, the data suggest that Pea and Bcs contribute to biofilm formation and in turn their presence contributes to fitness under water-limiting conditions, but not to the extent of alginate.     

The effect of high temperatures on tissue lactate,pyruvate and venous blood properties in the rat

Brain, muscle and liver pyruvate and L(+) lactate were determined in rats (1) control, T_re=37.2°C; (2) thermal stress T_re=40.8°C; (3) terminal thermal stress T_re=43.8°C, and (4) hypoxic death at reduced oxygen tension. A second experiment was conducted to examine venous blood acid-base parameters in heated rats at imminent death. Group 2 failed to show a significant change in brain lactate concentration, but muscle lactate decreased and liver lactate increased significantly (27 and 48%, respectively). Group 3 showed significant increases of 65 and 125% in the lactate content of brain and liver, respectively, but in this instance no significance was attributed to the 12% decrease in muscle lactate. Hypoxia in Group 4 resulted in the greatest increases in tissue lactate in brain, muscle and liver (130, 50 and 171%, respectively). The pyruvate concentrations of brain and liver in Group 2 exhibited no change, but muscle pyruvate decreased significantly (59%). Group 3 brain and muscle pyruvate decreased significantly by 57 and 74%, while liver pyruvate remained unchanged. Hypoxia (4) produced no significant differences in pyruvate levels from those observed in Group 3. The changes in venous blood properties of rats heated until respiratory movement ceased suggested acute and severe metabolic acidosis while animals exhibiting heat induced spasms and loss of coordination showed only slight decreases in blood pH and bicarbonate levels. Brain water content did not change, but muscle was dehydrated and liver tissue water content increased in rats exposed to lethal temperatures. The results indicate that hypoxia probably occurs in rat tissues at high temperatures, but not to a degree that would result in death.

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Zusammenfassung Gehirn, Muskel und Leberpyruvat and 1-Laktat wurden bestimmt in Ratten (1) Kontrollen, T_re=37,2°C; (2) Hitze T_re=40,8°C; (3) tödliche Belastung T_re=43,8°C und (4) Tod durch Hypoxie bei reduziertem PO₂. In einem zweiten Experiment wurde der Säure Basengehalt des venösen Blutes in erhitzten Tieren vor Eintritt des Todes bestimmt. Gruppe 2 zeigte keine signifikante Änderung in der Gehirnlaktatkonzentration, dagegen fiel das Muskellaktat (–27%) und das Leberlaktat stieg significant(+48%). Gruppe 3 zeigte einen signifikanten Anstieg des Gehirn- (+65%) und Leberlaktats (+125%), während das Muskellaktat fiel (–11%). Hypoxie in Gruppe 4 bewirkte den grössten Anstieg des Laktats im Gehirn (+130%), Muskel (+50%) und Leber (+171%). Die Pyruvatkonzentration in Gruppe 2 zeigte nur im Muskel einen Anstieg auf +59%. In Gruppe 3 fielen das Gehirn- (–57%) und Muskelpyruvat (–74%) signifikant, ohne Änderung in der Leber. Ähnliche Resultate ergaben sich bei Hypoxie in Gruppe 4. Vor dem Eintritt des Hitzetodes war im Blut hochgradige Acidose nachweisbar, während Tiere mit Hitzespasmus und Koordinationsstörungen nur gering niedrigere pH- und Bikarbonatwerte aufwiesen. Der Gehirnwassergehalt blieb unverändert, der Muskel war dehydriert und die Leber zeigte einen höheren Wassergehalt bei letal hohen Temperaturen. Die Ergebnisse zeigen, dass Hypoxia bei Hyperthermie auftritt, doch nicht in dem Ausmasse, um zum Tod zu führen.

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Resume On a déterminé les pyruvates et lactates du cerveau, des muscles et du foie de rats présentant 4 particularités différentes: (1) T_re (température rectale) = 37,2°C (contrôle); (2) T_re = 40,8°C (contrainte de chaleur); (3) T_re = 43,8°C (contraite de chaleur limite) et (4) morts par hypoxie par suite de la réduction de la pression partielle de l'oxygène. Dans un second essai, on a déterminé le rapport acides/bases dans le sang veineux de rats sous contrainte de chaleur, juste avant leur trépas. Dans le groupe 2, on n'a pas trouvé de modification significative de la concentration des lactates du cerveau. Par contre, celle des muscles a diminué (–27%) et celle du foie a augmenté (+48%) et cela de façon significative. Dans le groupe 3, on constate une hausse du taux de lactates dans le cerveau (+65%) et dans le foie (+125%), alors que celui des muscles diminue (–11%). L'hypoxie du groupe 4 a provoqué une forte augmentation des lactates aussi bien dans le cerveau (+130%), les muscles (+50%) que le foie (+171%). La concentration des pyruvates ne fut sensiblement modifiée dans le groupe 2 que pour les muscles (+59%). Dans le groupe 3, on en note une diminution significative dans le cerveau (–57%) et dans les muscles (–74%) alors que le taux du foie reste inchangé. On a obtenu des résultats analogues par l'hypoxie dans le groupe 4. On a pu constater une acidose aigue du sang juste avant la mort de chaleur, alors que les animaux atteints de spasmes et de perturbations dans la coordination ne présentaient que peu de variations du pH et du taux de bicarbonate. Dans un état létal par hautes températures, la teneur en eau du cerveau est restée inchangée. Les muscles étaient par contre déshydratés et le foie contenait davantage d'eau. Ces résultats ont montré que, lorsque la température augmente, les animaux souffrent d'hypoxie, mais pas au point d'en mourir.