Proline transport carrier-defective mutants of Escherichia coli K-12: properties and mapping.

A series of mutants of Escherichia coli K-12 requiring a high concentration of L-proline for growth were isolated from a proline auxotroph strain, JE2133. Genetic studies of the mutants, PT19, PT21, and PT22, showed that all the mutations (proT) were point mutations, and these were mapped at 82 min on the E. coli genetic map. Intact cells and cytoplasmic membrane vesicles of these mutants were specifically defective in L-proline transport activity. Strain PT21 had no detectable activity of the L-proline transport carrier at all, and strains PT19 and PT22 had only 1/35 and 1/70, respectively, of the transport activity of the parental strain. The mutants were also shown to have a defect in proline-binding function of the carrier by measuring specific binding of proline to sonically disrupted membranes. These results indicate that the gene proT determines the function of proline carrier in the cytoplasmic membrane.     

Carbohydrate Components of Influenza C Virions

The carbohydrate components of influenza C virions grown in chicken kidney (CK) cells were analyzed by gel filtration following exhaustive digestion with Pronase. The [³H]glucosamine-labeled glycopeptides were larger and more heterogeneous than those of influenza A/WSN virions; three major size classes (G₁, G₂, and G₃) were resolved. Treatment with Vibrio cholerae neuraminidase caused a decrease in size of G₁ and G₂, along with release of about 16% of the ³H label. The released sugar components were identified as N-acetylneuraminic acid by thin-layer chromatography. Peak G₃ was highly labeled with [³H]mannose, whereas G₁ and G₂ contained lower levels of mannose. The three major viral glycoproteins gp88, gp65, and gp30 were isolated from sodium dodecyl sulfate-polyacrylamide gels, and their glycopeptide components were analyzed after Pronase digestion. The three size classes of glycopeptides were obtained from any of the three glycoproteins; however, the relative amounts of the three components varied among the glycoproteins. Host cell-derived components, which appear to be mucopolysaccharides and glycoproteins, were found associated with influenza C virions grown in CK cells. These components contained glycopeptides that were mainly of sizes similar to peak G₂ from influenza C virions. Previous studies have shown that influenza A/WSN virus grown in several cell types contained only two size classes of glycopeptides. Two size classes comparable to peaks G₂ and G₃ from influenza C virions were also observed in influenza A/WSN grown in CK cells. Thus the large G₁ glycopeptides appear to be characteristic of influenza C virions.     

Population structure and genetic diversity in insular populations of Nasutitermes takasagoensis (Isoptera: Termitidae) analyzed by AFLP markers

Dispersal ability and degree of inbreeding in a population can indirectly be assessed using genetic markers. In general, it was suggested that winged termites are not able to fly distances greater than several hundred meters. Here, amplified fragment length polymorphism (AFLP) was used to analyze genetic diversity, population substructure, and gene flow among insular populations of the termite Nasutitermes takasagoensis (Isoptera: Termitidae) in the Yaeyama Islands, Okinawa, Japan. Samples were collected from 77 nests on seven islands of the Yaeyama Group. Using three primer combinations a total of 155 bands were generated with 78 (50%) polymorphic bands. Genetic distance and G(st) values among insular populations were calculated. Relatively high genetic diversity and low values of G (st), suggest there is moderate subpopulation structure. Based on these results, we discussed two possibilities; first, winged termites are able to fly over distances of several kilometers, and second, these results were obtained because insular populations share a recent common origin.     

Kinetic Analysis of Conformational Changes of GroEL Based on the Fluorescence of Tyrosine 506

The conformational changes of GroEL during the ATPase cycle in the presence of GroES were studied by measuring the fluorescence intensity time course of intrinsic tyrosine Y506, which is located near the nucleotide-binding site. A GroEL solution containing GroES was mixed with an ATP solution to initiate the reaction cycle. The tyrosine fluorescence intensity relative to that without the nucleotide reached 112% within the dead time of the apparatus (>15 s^?1) and further increased to 123% at 0.57 s^?1 followed by a decrease to 102% at 0.32 s^?1. An initial conformational change and a second intermediate state were expected to occur in ATP-bound GroEL because similar changes were observed for the ATPase-deficient D398A mutant. The conformational change to the third intermediate state corresponded to a process during or after ATP hydrolysis because D398A had no decreasing phase. The second intermediate state before ATP hydrolysis was characterized for the first time.     

Geographic color variation of Phelotrupes auratus (Coleoptera,Geotrupidae) in the Kinki region,central Japan: A quantitative spectrophotometric analysis

We investigated geographic color variation of the beetle Phelotrupes auratus in the Kinki region of central Japan using a spectrophotometer. The reflectance spectrum of the dorsal surface of the elytron was measured for beetles collected from 23 sites. We focused on λmax(α), the wavelength at the peak (α peak) between 400 and 700 nm (human visual sensitivity range), for analyses of color variation. In populations distributed on the Kii Peninsula, in the southern part of the Kinki region, average λmax(α) values were lower (480–497 nm) than in populations distributed in the area west of Lake Biwa, the western part of the Kinki region (618–633 nm). For populations distributed in the areas south and southeast of Lake Biwa, a geographic cline of λmax(α) was observed approximately along an east–west transect, with average λmax values varying continuously from 624 to 557 nm. The easternmost populations along this cline had almost the same λmax(α) values as the populations distributed west of Lake Biwa. The coefficient of variation for λmax(α) tended to be larger in populations with intermediate averages than those with lower or higher average values.     

Immunolocalization of 8-5′ and 8-8′ linked structures of lignin in cell walls of Chamaecyparis obtusa using monoclonal antibodies

Mouse monoclonal antibodies were generated against dehydrodiconiferyl alcohol- or pinoresinol-p-aminohippuric acid (pAHA)-bovine serum albumin (BSA) conjugate as probes that specifically react with 8-5′ or 8-8′ linked structure of lignin in plant cell walls. Hybridoma clones were selected that produced antibodies that positively reacted with dehydrodiconiferyl alcohol- or pinoresinol-pAHA–BSA and negatively reacted with pAHA–BSA and guaiacylglycerol-beta-guaiacyl ether-pAHA–BSA conjugates containing 8-O-4′ linkage. Eight clones were established for each antigen and one of each clone that positively reacted with wood sections was selected. The specificity of these antibodies was examined by competitive ELISA tests using various lignin dimers with different linkages. The anti-dehydrodiconiferyl alcohol antibody reacted specifically with dehydrodiconiferyl alcohol and did not react with other model compounds containing 8-O-4′, 8-8′, or 5-5′ linkages. The anti-pinoresinol antibody reacted specifically with pinoresinol and syringaresinol and did not react with the other model compounds containing 8-O-4′, 8-5′, or 5-5′ linkages. The antibodies also did not react with dehydrodiconiferyl alcohol acetate or pinoresinol acetate, indicating that the presence of free phenolic or aliphatic hydroxyl group was an important factor in their reactivity. In sections of Japanese cypress (Chamaecyparis obtusa), labeling by the anti-dehydrodiconiferyl alcohol antibody was found in the secondary walls of phloem fibers and in the compound middle lamellae, and secondary walls of tracheids. Weak labeling by the anti-pinoresinol antibody was found in secondary walls of phloem fibers and secondary walls and compound middle lamellae of developed tracheids. These labelings show the localization of 8-5′ and 8-8′ linked structure of lignin in the cell walls.     

High-level expression of the Geranylgeranyl diphosphate synthase gene in the frontal gland of soldiers in Reticulitermes speratus (Isoptera: Rhinotermitidae)

Defensive strategies of termite soldiers are roughly classified as either mechanical, using mandibles and/or the whole head, or chemical, using frontal gland secretion. Soldiers of the genus Nasutitermes (Termitidae, Nasutitermitinae), which is one of the most derived termite genera, use only chemical defenses, and diterpene defensive secretions were suggested to be synthesized through geranylgeranyl diphosphate (GGPP). On the other hand, soldiers of the genus Reticulitermes (Rhinotermitidae, Heterotermitinae) mainly use mechanical defenses, but also use supplementary chemical defenses involving frontal gland secretions, including diterpene alcohol. In this study, to confirm whether the GGPP is used for diterpene synthesis in a representative of an earlier-branching termite lineage, the GGPP synthase gene (RsGGPPS) was identified in the rhinotermitid Reticulitermes speratus (Kolbe). The relative expression level of RsGGPPS in soldiers was three-fold higher than in workers. Furthermore, RsGGPPS gene expression was detected in epithelial class 1 gland cells around the frontal-gland reservoir. Although GGPP is used for various essential cellular roles in animals, RsGGPPS is suggested to be used not only for these essential roles but also for diterpene synthesis in order to produce defensive secretions. Chemical structures of the diterpene identified from Reticulitermes and Nasutitermes are extremely different from each other, and the two genera are phylogenetically distant from each other. Thus, these two lineages may have independently acquired the abilities of diterpene synthesis from GGPP.     

Association between Catechol-O-Methyltrasferase Val108/158Met Genotype and Prefrontal Hemodynamic Response in Schizophrenia

Background

“Imaging genetics” studies have shown that brain function by neuroimaging is a sensitive intermediate phenotype that bridges the gap between genes and psychiatric conditions. Although the evidence of association between functional val108/158met polymorphism of the catechol-O-methyltransferase gene (COMT) and increasing risk for developing schizophrenia from genetic association studies remains to be elucidated, one of the most topical findings from imaging genetics studies is the association between COMT genotype and prefrontal function in schizophrenia. The next important step in the translational approach is to establish a useful neuroimaging tool in clinical settings that is sensitive to COMT variation, so that the clinician could use the index to predict clinical response such as improvement in cognitive dysfunction by medication. Here, we investigated spatiotemporal characteristics of the association between prefrontal hemodynamic activation and the COMT genotype using a noninvasive neuroimaging technique, near-infrared spectroscopy (NIRS).

Methodology/Principal Findings

Study participants included 45 patients with schizophrenia and 60 healthy controls matched for age and gender. Signals that are assumed to reflect regional cerebral blood volume were monitored over prefrontal regions from 52-channel NIRS and compared between two COMT genotype subgroups (Met carriers and Val/Val individuals) matched for age, gender, premorbid IQ, and task performance. The [oxy-Hb] increase in the Met carriers during the verbal fluency task was significantly greater than that in the Val/Val individuals in the frontopolar prefrontal cortex of patients with schizophrenia, although neither medication nor clinical symptoms differed significantly between the two subgroups. These differences were not found to be significant in healthy controls.

Conclusions/Significance

These data suggest that the prefrontal NIRS signals can noninvasively detect the impact of COMT variation in patients with schizophrenia. NIRS may be a promising candidate translational approach in psychiatric neuroimaging.     

Effects of metabotropic glutamate receptor 3 genotype on phonetic mismatch negativity

Background

The genetic and molecular basis of glutamatergic dysfunction is one key to understand schizophrenia, with the identification of an intermediate phenotype being an essential step. Mismatch negativity (MMN) or its magnetic counterpart, magnetic mismatch field (MMF) is an index of preattentive change detection processes in the auditory cortex and is generated through glutamatergic neurotransmission. We have previously shown that MMN/MMF in response to phoneme change is markedly reduced in schizophrenia. Variations in metabotropic glutamate receptor (GRM3) may be associated with schizophrenia, and has been shown to affect cortical function. Here we investigated the effect of GRM3 genotypes on phonetic MMF in healthy men.

Methods

MMF in response to phoneme change was recorded using magnetoencephalography in 41 right-handed healthy Japanese men. Based on previous genetic association studies in schizophrenia, 4 candidate SNPs (rs6465084, rs2299225, rs1468412, rs274622) were genotyped.

Results

GRM3 rs274622 genotype variations significantly predicted MMF strengths (p = 0.009), with C carriers exhibiting significantly larger MMF strengths in both hemispheres compared to the TT subjects.

Conclusions

These results suggest that variations in GRM3 genotype modulate the auditory cortical response to phoneme change in humans. MMN/MMF, particularly those in response to speech sounds, may be a promising and sensitive intermediate phenotype for clarifying glutamatergic dysfunction in schizophrenia.