Allele frequencies of human complement factor I in a sample from Iwate, northern Japan, with the description of geographical cline.

Serum samples from 270 healthy blood donors of Iwate prefecture, northern Japan, were examined for polymorphism of factor I (IF) by using polyacrylamide gel isoelectric focusing followed by semidry horizontal electroblotting with enzyme immunoassay. In 270 individuals four different patterns were observed, and these were controlled by two common alleles, IF*A and IF*B, and one rare allele, IF*A2. Allele frequencies were estimated to be 0.1019, 0.8963 and 0.0018 for IF*A, IF*B and IF*A2, respectively. The data of IF allele frequencies thus far reported in Japan excluding Okinawa Island were compared, and a statistically significant (p less than 0.01) geographical cline was detected for IF*A and IF*B alleles.     

PI (α1) polymorphism in the Japanese: Confirmation of PI*M4 and description of new PI variants

PI phenotyping by separator isoelectric focusing (SIEF) was performed on a total of 1000 unrelated Japanese individuals from two different areas of Western Japan. The PI M1M4 subtype was observed together with the six common PI M subtypes. PI*M4 was confirmed to be present but rare in the Japanese. Several new PI variants were identified by comparison runs of each variant with previously reported genetic variants. The significance of treatment of serum with dithiothreitol (DTT) followed by iodacetic acid (IAC) in determination of PI variants is also described.     

I (C3b/C4b inactivator) typing by agarose gel isoelectric focusing and immunoblotting technique

Genetic polymorphism of I (C3b/C4b inactivator) was studied by the method of agarose gel isoelectric focusing followed by an immunoblotting technique. Serum or plasma samples were pretreated with neuraminidase. The method is rapid, and gives the simple and reliable patterns of I. The allele frequencies calculated from healthy Japanese individuals living in the western part of Japan were: IF* A = 0.126 and IF*B = 0.874.     

Phosphoglucomutase-1 subtypes: polymorphic occurrence of PGM1*7+ and geographical variation in Japan

The distribution of gene frequencies in the phosphoglucomutase-1 (PGM1) system was investigated in two Japanese populations from Yamaguchi (Western Japan) and Okinawa (Southern Japan) using an improved isoelectric focusing method permitting the successful detection of the most anodal variant PGM1 3+. PGM1*7+ occurred with a polymorphic frequency of 0.012-0.021. A difference in the gene frequency was observed between the two populations. In comparison with neighboring populations, the Yamaguchi population was similar to Mongolians and Koreans in North China, and Okinawa to Zhuang in South China.     

Gc types in western Japan: report of a new variant Gc 1C35

Gc types were examined in a total of 1,000 unrelated Japanese individuals from Western Japan. By isoelectric focusing the six common subtypes and several rare types were observed. In addition, a new variant with a mobility between the Gc 1S and 1C2 was identified in 2 individuals. A family investigation confirmed the inheritance of the corresponding allele Gc* 1C35.     

HLA genes and haplotypes in Ryukyuans suggest recent gene flow to the Okinawa Islands.

Polymorphism of HLA genes was investigated in a population sample of Ryukyuans living on the main island of Okinawa (n = 197), in the southwestern islands of Japan. Serological typing was applied to class I loci (HLA-A, -B, and -C) and to HLA-DRB1; nucleotide sequence-level typing was performed using PCR microtiter plate hybridization and PCR single-strand conformation polymorphism methods. Ryukyuans showed a higher frequency of DRB1*0405 and lower frequencies of DRB1*1502 and DRB1*1302 compared with Hondo Japanese living on main islands. Principal components and phylogenetic analyses of 12 East Asian populations, including Ryukyuans, were performed based on the allele frequencies of HLA-A, -B, and -DRB1. In the principal components analysis 3 Japanese populations (Ryukyuans, Hondo Japanese, and Ainu) formed a cluster and showed the highest affinity to 2 Korean populations. In the phylogenetic tree Ryukyuans and Ainu were neighbors, but the genetic distance between them was larger than the distances between Ryukyuans and Hondo Japanese and between Ryukyuans and Korean populations. The geographic cline of the predominant haplotype in Ryukyuans, A*24-B*54-DRB1*0405, suggests that an ancestral population possessing A*24-B*54-DRB1*0405 moved into the Okinawa Islands after the divergence of Ryukyuans from the Ainu. Such a recent gene flow, probably from South China to the Okinawa Islands, is considered the major cause of difference in genetic characteristics between Ryukyuans and the Ainu.     

CYP2A6 polymorphism reveals differences in Japan and the existence of a specific variant in Ovambo and Turk populations

CYP2A6 is a polymorphic enzyme, and CYP2A6 genotype has been shown to be associated with smoking habits and lung cancer. We investigated CYP2A6 polymorphism in Japanese from four different geographic areas of Japan and in the Ovambo and Turk populations. Using two polymerase chain reaction restriction fragment length polymorphisms (PCR-RFLPs), we identified the functionally important variants of CYP2A6: *1A, *1B, *1F, *1G, *4A, and *4D. In the Japanese population the highest frequencies of the CYP2A6*1A allele were observed in subjects from the Fukuoka (Kyushu Island) and Ehime (Shikoku Island) prefectures, whereas subjects in Shimane and Tottori (both located on the Japan Sea side of Honshu Island) showed the highest frequencies of the CYP2A6*1B allele. In the Tottori and Shimane groups no subject was homozygous for the CYP2A6*4A allele, a whole gene deletion type that is prevalent among Asians. In the Ovambo and Turk populations the CYP2A6*1A allele was predominant. Furthermore, two alleles undetected in the Japanese were observed in these latter two ethnic groups: CYP2A6*1G was found solely in the Ovambos, and CYP2A6*1F was found solely in the Turks. The present study is the first to show interprefecture differences in CYP2A6 polymorphism in Japanese who live in relatively close but distinct geographic areas; this is also the first study to evaluate CYP2A6 variations among these Japanese and the Ovambo and Turk populations. The distribution results of these alleles could help to define the true significance of CYP2A6 polymorphism as a genetic susceptibility marker in worldwide populations.     

How specific MHC class I and class II combinations affect disease resistance against infectious salmon anaemia in Atlantic salmon (Salmo salar)

The aim was to evaluate the performance of selected individual MHC class I and class II alpha (A) alleles, and combinations of these on disease resistance against infectious salmon anaemia (ISA). The material consisting of 1966 fish from seven families, contained five MHC class I alleles and four MHC class II A alleles. Which representing given class II A and class II beta (B) haplotypes, totalling 19 MHC class I and class II A genotypes. The fish were challenged with infectious salmon anaemia virus (ISAV), the virus causing ISA. Dead fish were collected daily during the challenge experiment and the survivors were collected at termination. All fish were genotyped for MHC class I and class II A. The total mortality in the material was 85.14%. For MHC class I, UBA*0201 and UBA*0301 were significantly the most resistant alleles, while UBA*0601 for class I and DAA*0301 for class II A were the significantly most susceptible alleles. The analysis of combined MHC class I and class II A genotypes detected that fish with the genotype UBA*0201/*0301;DAA*0201/*0201 were the most resistant fish with a hazard ratio (HR) at 0.750, while the fish with the genotypes UBA*0601/*0801;DAA*0501/*0501 and UBA*0201/*0301;DAA*0301/*0501 were the most susceptible fish with HR of 1.334 and 1.425. In addition, Cox regression analysis within family detected combined MHC class I and class II A genotypes that contributed significantly to resistance and susceptibility. The study confirmed the expectation of performance of individual MHC class I and class II A alleles, and also detected an effect of MHC class I and class II A in combinations.     

Aberrant expression of HLA-B*3565Q is associated with a disrupted disulfide bond

The identification of expression variants is a challenge in HLA diagnostics. We here describe the identification of the novel allele HLA-B*3565Q. The serological HLA class I type, as determined by a lymphocytotoxicity test, was A11,24; B38; Bw4; Cw−; whereas PCR-sequence-specific primers resulted in A*11,*24, B*35,*38; Cw*12, thus suggesting the presence of a nonexpressed B*35 allele. To clarify the lack of serological HLA-B35 reactivity, exons 2 and 3 were sequenced following haplotype-specific amplification. At position 564 from the beginning of the coding region (exon 3), a transversion (C→G) was observed, which, at the amino acid level, results in a substitution from cysteine to tryptophane at position 164 of the mature polypeptide. Because this position is essential for the formation of a disulfide bond linking the cysteine residues at positions 101 and 164, which is strongly conserved in functional class I molecules of vertebrates, the disruption of this bond is very likely to be the reason for the lack of serological detectability. We later found the same novel allele in a second unrelated individual, of whom we were able to establish a lymphoblastoid cell line (B-LCL). Serological testing of this B-LCL indicated a very low aberrant expression of HLA-B*3565Q, which cannot be expected to be detected by standard serology techniques. Holger-Andreas Elsner and Peter A. Horn contributed equally to this work. The nucleotide sequence data reported in this paper have been published in the European Molecular Biology Laboratory, GenBank, and DNA Data Bank of Japan Nucleotide Sequence Database under the accession numbers AJ278746, AJ278747, and AJ879892. The name B*3565Q was officially assigned by the WHO Nomenclature Committee in December 2005. This follows the agreed policy that, subject to the conditions stated in the most recent Nomenclature Report (Marsh et al. 2005), names will be assigned to new sequences as they are identified. Lists of such new names will be published in the following WHO Nomenclature Report.     

Coherent interaction of dynamical attractors for object-based selective attention

I investigate essential neuronal mechanisms of visual attention based on object-based theory and a biased-competition scheme. A neural network model is proposed that consists of two feature networks, FI and FII, and one object network, OJ. The FI and FII networks send feedforward projections to the OJ network and receive feedback projections from the OJ network in a convergent/divergent manner. The OJ network integrates information about sensory features originated from the FI and FII networks into information about objects. I let the feature networks and the object network memorize individual features and objects according to the Hebbian learning rule and create the point attractors corresponding to these features and objects as long-term memories in the network dynamics. When the model tries to attend to objects that are superimposed, the point attractors relevant to the two objects emerge in each network. After a short interval (hundreds of milliseconds), the point attractors relevant to one of the two objects are selected and the other point attractors are completely suppressed. I suggest that coherent interactions of dynamical attractors relevant to the selected object may be the neuronal substrate for object-based selective attention. Bottom-up (FI-to-OJ and FI-to-OJ) neuronal mechanisms separate candidate objects from the background, and top-down (OJ-to-FI and OJ-to-FII) mechanisms resolve object-competition by which one relevant object is selected from candidate objects.     

A sensitive method for detecting the fermentation-inhibition antibody to Mycoplasma pneumoniae.

The fermentation-inhibition (FI) test for Mycoplasma pneumoniae was improved by using a combination of guinea pig complement and gamma globulin-depleted horse serum in place of unheated whole horse serum employed in the conventional assay system. As the test antigen for the new FI assay system, M. pneumoniae filtrated through a 3.0 microns membrane filter was used. Owing to the strong augmenting effect of guinea pig complement, the FI activity of rabbit immune serum was increased 32-fold in the new system compared with the conventional system. Furthermore, IgM antibody, which is barely detectable by the conventional system, could easily be titrated by the new system. With this sensitive method, rapid rise of FI titer was clearly demonstrable in most children with acute M. pneumoniae infections, and a prevalence of FI or growth-inhibitory antibody among healthy adults in Japan (82%) was revealed.     

The influence of HLA class I alleles and heterozygosity on the outcome of human T cell lymphotropic virus type I infection

The inflammatory disease human T cell lymphotropic virus type I (HTLV-I)-associated myelopathy (HAM/TSP) occurs in only 1-2% of HTLV-I-infected individuals and is associated with a high provirus load of HTLV-I. We hypothesize that a person's risk of developing HAM/TSP depends upon the efficiency of their immune response to the virus, which differs between individuals because of polymorphism in genes that influence this response. Previously we showed that the possession of HLA-A*02 was associated with a lower risk of HAM/TSP, and with a lower provirus load in healthy carriers of HTLV-I. However, HLA-A*02 did not account for all the observed difference in the risk of HAM/TSP. Here we present evidence, in the same study population in Japan, that HLA-Cw*08 was also associated with disease protection (probability value, two-tailed test = 0.002) and with a lower proviral load in healthy carriers. Possession of the A*02 and/or Cw*08 genes prevented 36% of potential HAM/TSP cases. In contrast, HLA-B*5401 was associated with a higher susceptibility to HAM/TSP (probability value, two-tailed test = 0.0003) and with a higher provirus load in HAM/TSP patients. At a given provirus load, B*5401 appeared to increase the risk of disease. The fraction of HAM/TSP cases attributable to B*5401 was 17%. Furthermore, individuals who were heterozygous at all three HLA class I loci have a lower HTLV-I provirus load than those who were homozygous at one or more loci. These results are consistent with the proposal that a strong class I-restricted CTL response to HTLV-I reduces the proviral load and hence the risk of disease.     

A dip in the chlorophyll fluorescence induction at 0.2-2 s in Trebouxia-possessing lichens reflects a fast reoxidation of photosystem I. A comparison with higher plants

An unusual dip (compared to higher plant behaviour under comparable light conditions) in chlorophyll fluorescence induction (FI) at about 0.2-2 s was observed for thalli of several lichen species having Trebouxia species (the most common symbiotic green algae) as their native photobionts and for Trebouxia species cultured separately in nutrient solution. This dip appears after the usual O(J)IP transient at a wide range of excitation light intensities (100-1800 micromol photons m(-2) s(-1)). Simultaneous measurements of FI and 820-nm transmission kinetics (I(820)) with lichen thalli showed that the decreasing part of the fluorescence dip (0.2-0.4 s) is accompanied by a decrease of I(820), i.e., by a reoxidation of electron carriers at photosystem I (PSI), while the subsequent increasing part (0.4-2 s) of the dip is not paralleled by the change in I(820). These results were compared with that measured with pea leaves-representatives of higher plants. In pea, PSI started to reoxidize after 2-s excitation. The simultaneous measurements performed with thalli treated with methylviologen (MV), an efficient electron acceptor from PSI, revealed that the narrow P peak in FI of Trebouxia-possessing lichens (i.e., the I-P-dip phase) gradually disappeared with prolonged MV treatment. Thus, the P peak behaves in a similar way as in higher plants where it reflects a traffic jam of electrons induced by a transient block at the acceptor side of PSI. The increasing part of the dip in FI remained unaffected by the addition of MV. We have found that the fluorescence dip is insensitive to antimycin A, rotenone (inhibitors of cyclic electron flow around PSI), and propyl gallate (an inhibitor of plastid terminal oxidase). The 2-h treatment with 5 microM nigericin, an ionophore effectively dissipating the pH-gradient across the thylakoid membrane, did not lead to significant changes either in FI nor I(820) kinetics. On the basis of the presented results, we suggest that the decreasing part of the fluorescence dip in FI of Trebouxia-lichens reflects the activation of ferredoxin-NADP(+)-oxidoreductase or Mehler-peroxidase reaction leading to the fast reoxidation of electron carriers in thylakoid membranes. The increasing part of the dip probably reflects a transient reduction of plastoquinone (PQ) pool that is not associated with cyclic electron flow around PSI. Possible causes of this MV-insensitive PQ reduction are discussed.     

Motifs in outer membrane protein sequences: applications for discrimination

Discriminating outer membrane proteins (OMPs) from other folding types of globular and membrane proteins is an important problem for predicting their secondary and tertiary structures and detecting outer membrane proteins from genomic sequences as well. In this work, we have systematically analyzed the distribution of amino acid residues in the sequences of globular and outer membrane proteins with several motifs, such as A*B, A**B, etc. We observed that the motifs E*L, A*K and L*E occur frequently in globular proteins while S*S, N*S and R*D predominantly occur in OMPs. We have devised a statistical method based on frequently occurring motifs in globular and OMPs and obtained an accuracy of 96% and 82% for correctly identifying OMPs and excluding globular proteins, respectively. Further, we noticed that the motifs of transmembrane helical (TMH) proteins are different from that of OMPs. While I*A, I*L and L*I prefer in TMH proteins S*S, N*S and N*N predominantly occur in OMPs. The information about the occurrence of A*B motifs in TMH and OMPs could discriminate them with an accuracy of 80% for excluding OMPs and 100% for identifying OMPs. The influence of protein size and structural class for discrimination is discussed.     

Mitochondrial DNA variation in the Viking age population of Norway

The medieval Norsemen or Vikings had an important biological and cultural impact on many parts of Europe through raids, colonization and trade, from about AD 793 to 1066. To help understand the genetic affinities of the ancient Norsemen, and their genetic contribution to the gene pool of other Europeans, we analysed DNA markers in Late Iron Age skeletal remains from Norway. DNA was extracted from 80 individuals, and mitochondrial DNA polymorphisms were detected by next-generation sequencing. The sequences of 45 ancient Norwegians were verified as genuine through the identification of damage patterns characteristic of ancient DNA. The ancient Norwegians were genetically similar to previously analysed ancient Icelanders, and to present-day Shetland and Orkney Islanders, Norwegians, Swedes, Scots, English, German and French. The Viking Age population had higher frequencies of K*, U*, V* and I* haplogroups than their modern counterparts, but a lower proportion of T* and H* haplogroups. Three individuals carried haplotypes that are rare in Norway today (U5b1b1, Hg A* and an uncommon variant of H*). Our combined analyses indicate that Norse women were important agents in the overseas expansion and settlement of the Vikings, and that women from the Orkneys and Western Isles contributed to the colonization of Iceland.     

Antichymotrypsin interaction with chymotrypsin. Intermediates on the way to inhibited complex formation.

Serpins form enzymatically inactive covalent complexes (designated E*I*) with their target proteinases, corresponding most likely to the acyl enzyme that resembles the normal intermediate in substrate turnover. Formation of E*I* involves large changes in the conformation of the reactive center loop (residues P17 to P9') and of the serpin molecule in general. The "hinge" region of the reactive center loop, including residues P10-P14, shows facile movement in and out of beta-sheet A, and this movement appears to be crucial in determining whether E*I* is formed (the inhibitor pathway) or whether I is rapidly hydrolyzed to I* (the substrate pathway). Here, we report stopped-flow and rapid quench studies investigating the pH dependence of the conversion of the alpha1-antichymotrypsin.alpha-chymotrypsin encounter complex, E.I, to E*I*. These studies utilize fluorescent derivatives of cysteine variants of alpha1-antichymotrypsin at the P11 and P13 residues. Our results demonstrate three identifiable intermediates, EIa, EIb, and EIc, between E.I and E*I* and permit informed speculation regarding the nature of these intermediates. Partitioning between inhibitor and substrate pathways occurs late in the process of E*I* formation, most likely from a species occurring between EIc and E*I*.     

Acquisition of complement inhibitor serine protease factor I and its cofactors C4b-binding protein and factor H by Prevotella intermedia

Infection with the Gram-negative pathogen Prevotella intermedia gives rise to periodontitis and a growing number of studies implies an association of P. intermedia with rheumatoid arthritis. The serine protease Factor I (FI) is the central inhibitor of complement degrading complement components C3b and C4b in the presence of cofactors such as C4b-binding protein (C4BP) and Factor H (FH). Yet, the significance of complement inhibitor acquisition in P. intermedia infection and FI binding by Gram-negative pathogens has not been addressed. Here we show that P. intermedia isolates bound purified FI as well as FI directly from heat-inactivated human serum. FI bound to bacteria retained its serine protease activity as shown in degradation experiments with (125)I-labeled C4b. Since FI requires cofactors for its activity we also investigated the binding of purified cofactors C4BP and FH and found acquisition of both proteins, which retained their activity in FI mediated degradation of C3b and C4b. We propose that FI binding by P. intermedia represents a new mechanism contributing to complement evasion by a Gram-negative bacterial pathogen associated with chronic diseases.     

Orosomucoid (ORM) typing by isoelectric focusing: an analysis of ORM haplotypes

A new separator isoelectric focusing method for typing of orosomucoid (ORM) was developed. This method provided a superior resolution of ORM patterns: two close bands of ORM1*5.2 products were clearly separated. A total of 364 subjects from Okinawa (Japan) were classified into 21 ORM phenotypes determined by 6 ORM1 and 7 ORM2 alleles including a polymorphic silent allele, ORM2*QO, and 2 new rare variants, ORM2*18 and ORM2*19. These phenotypes were also explained by 12 ORM haplotypes, half of which were polymorphic.     

Fixed interval and fixed time treadle pressing in the pigeon: A comparison with FI and FT keypecking

Experiment I used non-naive pigeons having previously performed on both keypecking and treadlepressing Fixed Interval schedules. In condition IT, treadlepressing was reinforced on successive Fixed Interval 60 seconds, Fixed Time 60 seconds and Fixed Interval 60 seconds schedules. Subsequently (condition IK), the same subjects pecked a key on an identical schedule sequence (FI60, FT60, FI60). In Experiment II, separate groups of naïve subjects were assigned either to treadlepressing (condition IIT) or keypecking (condition IIK) and to the same schedule sequence (FI60, FT60, FI60). Treadle pressing and keypecking decreased greatly in Fixed Time schedules. Curvature indices, pauses and running rates were less sensitive than response rates to the switching from one schedule to the other. Experiments I and II yielded similar results, experimental history accounting only for minor differences. The results were discussed in relation to interspecies differences in the temporal regulation of behavior and operant versus respondent control of the response and schedule-induced behaviour.     

Placental glucose dehydrogenase polymorphism in Japanese

The polymorphism of glucose dehydrogenase (GDH) was investigated in 516 Japanese placentae. The allele frequencies were GDH*1 = 0.510, GDH*2 = 0.488 and GDH*3 = 0.002. GDH*3 appears to increase from Japan via Southeast Asia and India to Europe.