Identification of a single killer immunoglobulin-like receptor (KIR) gene in the porcine leukocyte receptor complex on chromosome 6q

Human killer immunoglobulin-like receptors (KIR) are expressed on natural killer (NK) cells and are involved in their immunoreactivity. While KIR with a long cytoplasmic tail deliver an inhibitory signal when bound to their respective major histocompatibility complex class I ligands, KIR with a short cytoplasmic tail can activate NK responses. The expansion of the KIR gene family originally appeared to be a phenomenon restricted to primates (human, apes, and monkeys) in comparison to rodents, which via convergent evolution have numerous C-type lectin-like Ly49 molecules that function analogously. Further studies have shown that multiple KIR are also present in cow and horse. In this study, we have identified by comparative genomics the first and possibly only KIR gene, named KIR2DL1, in the domesticated pig (Sus scrofa) allowing further evolutionary comparisons to be made. It encodes a protein with two extracellular immunoglobulin domains (D0 + D2), and a long cytoplasmic tail containing two inhibitory motifs. We have mapped the pig KIR2DL1 gene to chromosome 6q. Flanked by LILRa, LILRb, and LILRc, members of the leukocyte immunoglobulin-like receptor (LILR) family, on the centromeric end, and FCAR, NCR1, NALP7, NALP2, and GP6 on the telomeric end, pig demonstrates conservation of synteny with the human leukocyte receptor complex (LRC). Both the porcine KIR and LILR genes have diverged sufficiently to no longer be clearly orthologous with known human LRC family members.     

The genomic organization and evolution of the natural killer immunoglobulin-like receptor (KIR) gene cluster

Natural killer (NK) immunoglobulin-like receptors (KIRs) are a family of polymorphic receptors which interact with specific motifs on HLA class I molecules and modulate NK cytolytic activity. In this study, we analyzed a recently sequenced subgenomic region on chromosome 19q13.4 containing eight members of the KIR receptor repertoire. Six members are clustered within a 100-kb continuous sequence. These genes include a previously unpublished member of the KIR gene family 2DS6, as well as 2DL1, 2DL4, 3DL1, 2DS4, 3DL2, from centromere to telomere. Two additional KIR genes, KIRCI and 2DL3, which may be located centromeric of this cluster were also analyzed. We show that the KIR genes have undergone repeated gene duplications. Diversification between the genes has occurred postduplication primarily as a result of retroelement indels and gene truncation. Using pre- and postduplication Alu sequences identified within these genes as evolutionary molecular clocks, the evolution and duplication of this gene cluster is estimated to have occurred 30–45 million years ago, during primate evolution. A proposed model of the duplication history of the KIR gene family leading to their present organization is presented. Received: 25 November 1999 / Revised: 10 January 2000     

Complexity in cattle <Emphasis Type="Italic">KIR</Emphasis> genes: transcription and genome analysis

Killer immunoglobulin (Ig)-like receptors (KIRs) are the major functional natural killer (NK) cell receptors in human. The presence of KIR genes has only recently been demonstrated in other (non-primate) species, and their expression, genomic arrangement, and function in these species have yet to be investigated. In this study, we describe the KIR gene family in cattle. KIR sequences were amplified from cDNA derived from four animals. Seventeen new sequences were identified in total. Some are alleles of two previously described genes, and the remainder are representative of at least four additional genes. These cDNA data, together with analysis of the cattle genome sequence, confirm that, as in humans, cattle have multiple inhibitory and activating KIR genes, with variable haplotype composition, and putative framework genes. In contrast to human, the majority of the cattle KIR genes encode three Ig-domain KIRs; most of the inhibitory genes encode only one immunoreceptor tyrosine-based inhibitory motif (ITIM), and the activating genes encode molecules with arginine rather than the more usual lysine in the transmembrane domain. A divergent gene, 2DL1, encodes a two Ig-domain KIR with an unusual D0-D2 structure, and a distinct signaling domain with two ITIMs. Similarity to pig and human two Ig-domain (D0-D2) KIRs suggest these may be more related to an ancestral gene than the other cattle KIR genes. Cattle have multiple NKG2A-related genes and at least one Ly49 gene; thus, the data presented here suggest that they have the potential to express more major histocompatibility complex-binding NK receptors than other species.     

Haplotype block and superblock structures of the alpha1-adrenergic receptor genes reveal echoes from the chromosomal past

A significant proportion of the human genome is contained within haplotype blocks across which pairwise linkage disequilibrium (LD) is very high. However, LD is also often high between markers at more remote distances, and within different haplotype blocks. Here, we evaluate the origins of haplotype block structure in the three genes for alpha1 adrenergic receptors (alpha1-AR) in the human genome ( ADRA1A, ADRA1B and ADRA1D) by genotyping dense single-nucleotide polymorphism (SNP) marker maps, and show that LD signals between distant markers are due to the presence of extended haplotype superblocks in individuals with ancient chromosomes which have escaped historic recombination. ARs mediate the physiological effects of epinephrine and norepinephrine, and are targets of many therapeutic drugs. This work has identified haplotype backgrounds of alpha1-AR missense variants, haplotype block structures in US Caucasians and African Americans, and haplotype tag SNPs for each block, and we present strong evidence for ancient haplotype block superstructure at these genes which has been partially disrupted by recombination, and evidence for reinstatement of linkage disequilibrium by subsequent recombination events. ADRA1A is comprised of four haplotype blocks in US Caucasians, while in African Americans Block 1 is split. ADRA1B has four blocks in US Caucasians, but in African Americans only the first two blocks are present. ADRA1D has two blocks in US Caucasians, and the first block is replaced by two smaller blocks in African Americans. For both ADRA1A and ADRA1B, haplotype superstructures may represent a novel, higher-level hierarchy in the human genome, which may reduce redundancy of testing by further aggregation of genotype data.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by W. R. McCombie     

Role of HLA antigens in Rh (D) alloimmunized pregnant women from Mumbai, Maharashtra, India

Immunogenetic studies in various diseases provide potential genetic markers. We have studied the incidence of HLA A, B, C, DR and DQ loci antigen in Rh (D) antigen isoimmunized mothers compared to those nonimmunized isoimmunized Rh negative mothers. Seventy six mothers who were immunized to Rh (D) antigen due to pregnancy (responders) and fifty four mothers who did not develop Rh (D) isoimmunization despite positive pregnancies (nonresponders) were selected for the study. Standard methods of serological HLA typing, ABO and Rh (D) groups, and screening for Rh D antibodies were used. 392 unrelated individuals from the population were compared as controls. In addition 45 unrelated individuals from the same population were typed for HLA DRB and DQB gene using PCR-SSP kits. The genotype frequencies of HLA A2, A3, A28, B13, B17, B35, B52, B60, Cw2, Cw6, DR4, and DQ3 were significantly increased, while the frequencies of the HLA A11, A29, A31, B7, B37, B51, Cw1 and DR9 were decreased in the responder women when compared to the non-responder women. HLA A30 (19) split antigen was not identified in immunized women while HLA A23 (9) split antigen was not identified in non immunized women. HLA A3, B17, Cw2 and DR4 showed a significant relative risk among the immunized responder women. When compared with Rh immunized women (responders) reported from USA, England and Hungary the phenotype frequencies of HLA A11, A24, A28, B5, B17, B40, DR2 and DR5 were increased while HLA A23, B8, B18, and DR6 were decreased in the Indian Rh immunized women. Two locus haplotype frequency analysis observed among the responders women revealed that among the significant haplotypes expressed A2–B5, B7–Cw1, DR2–DQ1 were highly significant haplotypes in positive linkage, while A1–B5, and A1–B7 were in significant negative linkage disequilibrium. The haplotype frequencies were ≤one when these common hapoltypes were compared with control population. Thus in the present study it is evident that the inheritance of HLA A3, B17, Cw2 and DR4 increases the relative risk factor by 2.6 times among Indian Rh isoimmunized women. Further, it is evident that there are significant differences in the observed HLA antigen frequencies and two locus haplotypes in Rh isoimmunized women when compared to women from USA, UK and Hungary due to extreme HLA polymorphism in different populations of the world     

Biased distribution of single nucleotide polymorphisms (SNPs) in porcine Toll-like receptor 1 (TLR1), TLR2, TLR4, TLR5, and TLR6 genes

Toll-like receptors (TLRs) recognize various microbial components and induce immune responses. Polymorphisms in TLRs may influence their recognition of pathogen-derived molecules; swine TLRs are predicted to be associated with responses to infectious diseases such as pneumonia. In this study, we searched for single nucleotide polymorphisms (SNPs) in the coding sequences of porcine TLR1, TLR2, TLR4, TLR5, and TLR6 genes in 96 pigs from 11 breeds and elucidated 21, 11, 7, 13, and 11 SNPs, respectively, which caused amino acid substitutions in the respective TLRs. Distribution of these nonsynonymous SNPs was biased; many were located in the leucine-rich repeats, particularly in TLR1. These data demonstrated that the heterogeneity of TLR genes was preserved in various porcine breeds despite intensive breeding that was carried out for livestock improvement. It suggests that the heterogeneity in TLR genes is advantageous in increasing the possibility of survival in porcine populations.Electronic SupplementaryMaterial Supplementary material is available for this article at     

过去二十年贵州黑叶猴分布与种群动态及致危因子分析(英文)

本文通过整理相关文献与政府报告数据并结合我们在贵州的最新调查与监测数据,总结过去20年间黑叶猴在贵州的分布、种群变动及其所面临的主要威胁,并在此基础上提出相关的保护管理建议。贵州黑叶猴野生种群在过去20年间增长了约10%～20%:从20世纪90年代的约109群1000只增加到现在的约132～137群1160～1200只。但同期黑叶猴已从5个原有的分布点绝迹,现仅存于大沙河、柏箐、麻阳河、宽阔水和野钟等5个保护区,总面积约912km2。现有的5个隔离分布点保存有全球黑叶猴野生种群(1800～2000)的约62%,是黑叶猴物种保存的最关键地区,并应该给于保护管理的优先考虑。偷猎压力在贵州相对较小:在过去的20年间仅有3次偷猎的官方记录。黑叶猴在贵州所面临的主要威胁是栖息地的丧失与退化。其主要表现形式有:以农作物和烟草种植为目的的耕地扩大,以做饭、取暖以及烟叶烘烤为目的的薪材需求,以及家畜(主要是山羊)的过度放牧。     

Ultrastructure and chemistry of the cell wall of the moss Rhacocarpus purpurascens (Rhacocarpaceae): a puzzling architecture among plants

The architectural, compositional and functional characteristics of the cell walls of the leaves of the moss Rhacocarpus purpurascens (Brid.) Par. have been analysed by scanning and transmission electron microscopy, wall-extraction methods, nuclear magnetic resonance (NMR) spectroscopy, and water-retention experiments. Four-layered cell walls with a peculiar architecture which, so far, appears to be unique among plants were apparent. The architecture of the walls was not affected by sequential wall-extraction procedures. Subsequent analysis of the residual pre-extracted walls by classical spectro-photometrical methods revealed that the walls are composed of mainly lignin, hemicellulose and cellulose in a ratio of about 9:8:5, determining their integrity. This was supported by NMR spectroscopy. The resonance spectrum showed various characteristics typical of lignin; however, some specific peaks associated with lignin were missing. The walls exhibited no particular properties for external water conduction but seem to be adapted to rapid absorption of fog, dew, or rain. Received: 3 June 1997 / Accepted: 25 February 1998     

Research progress on distribution,migration, transformation of antibiotics and antibiotic resistance genes (ARGs) in aquatic environment

Antimicrobial and antibiotics resistance caused by misuse or overuse of antibiotics exposure is a growing and significant threat to global public health. The spread and horizontal transfer of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) by the selective pressure of antibiotics in an aquatic environment is a major public health issue. To develop a better understanding of potential ecological risks die to antibiotics and ARGs, this study mainly summarizes research progress about: (i) the occurrence, concentration, fate, and potential ecological effects of antibiotics and ARGs in various aquatic environments, (ii) the threat, spread, and horizontal gene transfer (HGT) of ARGs, and (iii) the relationship between antibiotics, ARGs, and ARB. Finally, this review also proposes future research direction on antibiotics and ARGs.     

ABO,Rh(D) and haptoglobin distribution in a sample from the sultanate of Oman

The distribution of phenotypic frequencies of ABO, Rh(D) system and Hp polymorphism in a sample from Muscat, Oman, revealed the following gene frequencies: P=0.164, q=0.110, r=0.726; D=0.738, d=0.262; Hp¹=0.289 and Hp²=0.711. These frequencies are in good agreement with the distribution in the Euroasiatic continent.     

Examining the critical roles of human CB2 receptor residues Valine 3.32 (113) and Leucine 5.41 (192) in ligand recognition and downstream signaling activities

We performed molecular modeling and docking to predict a putative binding pocket and associated ligand–receptor interactions for human cannabinoid receptor 2 (CB2). Our data showed that two hydrophobic residues came in close contact with three structurally distinct CB2 ligands: CP-55,940, SR144528 and XIE95-26. Site-directed mutagenesis experiments and subsequent functional assays implicated the roles of Valine residue at position 3.32 (V113) and Leucine residue at position 5.41 (L192) in the ligand binding function and downstream signaling activities of the CB2 receptor. Four different point mutations were introduced to the wild type CB2 receptor: V113E, V113L, L192S and L192A. Our results showed that mutation of Val113 with a Glutamic acid and Leu192 with a Serine led to the complete loss of CB2 ligand binding as well as downstream signaling activities. Substitution of these residues with those that have similar hydrophobic side chains such as Leucine (V113L) and Alanine (L192A), however, allowed CB2 to retain both its ligand binding and signaling functions. Our modeling results validated by competition binding and site-directed mutagenesis experiments suggest that residues V113 and L192 play important roles in ligand binding and downstream signaling transduction of the CB2 receptor.     

Analysis of serotonin receptor 2A gene (HTR2A): Association study with autism spectrum disorder in the Indian population and investigation of the gene expression in peripheral blood leukocytes

Several studies suggest involvement of serotoninergic system in the pathophysiology of Autism Spectrum Disorder (ASD). The 5-HT receptor binding studies using ³H-lysergic acid diethylamide (³H-LSD) and linkage analysis provided evidences to consider HTR2A as a potential candidate gene for ASD. The three SNPs, −1438A/G (rs6311), 102T/C (rs6313) and 1354C/T (rs6314) of HTR2A have been well studied in the etiology of various neuropsychiatric disorders. But studies on association of this gene with ASD are limited to two reports from American and Korean populations. Additionally there are reports, which demonstrated paternal imprinting of HTR2A with expression from only one allele. So far no reports are available on HTR2A and its association with any neuropsychiatric disorders from Indian population. Therefore, the present study investigates association of the above mentioned three markers of HTR2A with ASD in Indian population using population and family-based approaches. The study also deals with allelic expression pattern of HTR2A in Peripheral Blood Leukocytes (PBLs) to understand the parental imprinting status. The genotyping analyses were carried out for probands, parents and controls. The subsequent association analyses did not show association of these markers with ASD. So, HTR2A is unlikely to be a genetic marker for ASD in Indian population. The expression analyses showed absence of monoallelic expression, suggesting lack of parental imprinting of HTR2A gene. However, we noticed methylation of the CpG sites at −1438A/G and 102T/C loci of HTR2A gene. Further bioinformatics analysis revealed absence of CpG islands in the promoter of the gene supporting biallelic expression pattern of HTR2A in PBLs.     

Senescence and steroid hormone receptor reactivities in accessory sex glands of elderly rats (Sprague-Dawley) following exogenous hormonal therapy

The aim of this study was to characterize the stromal and epithelial distribution of AR, ERα and ERβ reactivities in the different accessory sex glands of elderly rats and during strong hormonal changes. Ten month old male rats were divided into six senile groups and submitted to treatment: Senile/Control group (SC); Senile/Testosterone group (ST): Senile/Estrogen group (SE); Castrated group (CA); Castrated/Testosterone group (CT); Castrated/Estrogen group (CE). After a 30-day treatment, the prostatic ventral lobe (VL), dorsal lobe (DL) and coagulating gland (CG) samples were processed for immunohistochemistry and Western Blotting. The results showed that AR immunoreactivity was characterized in the epithelium of VL and DL in senile/control rats and senile rats submitted to exogenous hormonal therapy. AR reactivity in the coagulating gland was verified predominantly in the stromal cells in the different experimental groups. ERα reactivity occurred predominantly in the stromal compartment in all accessory sex glands. In the DL and CG, ERα immunoreactivities were intense in the groups which received testosterone (ST) and estrogen (SE). ERβ immunoreactivity in the CG was verified in the stromal compartment in the different experimental groups, showing a positive response to both increased testosterone and estrogen levels. ERβ reactivity, in the DL, was intensified in the stroma of senile rats with higher serum testosterone levels, and in senile rats with increased serum estrogen levels, especially in the glandular epithelium. Thus, the results revealed different distribution pattern of steroid hormone receptors in each one of the prostatic lobes in senescence, especially in the prostate dorsal lobe and coagulating gland, which is a fundamental factor due to the fact that major prostatic diseases occur in a later period of life.     

Ins(1,4,5)P3 receptor type 1 associates with AKAP9 (AKAP450 variant) and protein kinase A type IIβ in the Golgi apparatus in cerebellar granule cells

Background information. Interconnections between the Ca²⁺ and cAMP signalling pathways can determine the specificity and diversity of the cellular effects mediated by these second messengers. Most cAMP effects are mediated by PKA (protein kinase A), which is anchored close to its membranous substrates by AKAPs (A kinase‐anchoring proteins). In many cell types, the activation of InsP₃R (inositol 1,4,5‐trisphosphate receptor), an endoplasmic reticulum Ca²⁺ channel, is a key event of Ca²⁺ signalling. The phosphorylation of InsP₃R1 by PKA stimulates Ca²⁺ mobilization. This control is thought to be tight, involving the association of PKA with InsP₃R1. The InsP₃R1 isoform predominates in central nervous tissue and its concentration is highest in the cerebellar microsomes. We investigated the complex formed by InsP₃R1 and PKA in this fraction, vith a view to identifying its components and determining its distribution in the cerebellar cortex. Results. Immunoprecipitation experiments showed that InsP₃R1 associated with PKA type IIβ and AKAP450, the longer variant of AKAP9, in sheep cerebellar microsomes. The co‐purification of AKAP450 with InsP₃R1 on heparin‐agarose provided further evidence of the association of these proteins. Immunohistofluorescence experiments on slices of cerebellar cortex showed that AKAP450 was colocalized with InsP₃R1 and RIIβ (regulatory subunit of PKA IIβ) in granule cells, but not in Purkinje cells. AKAP450 was localized in the Golgi apparatus of these two cell types whereas InsP₃R1 was detected in this organelle only in granule cells. Conclusions. Taken together these results suggest that InsP₃R1 forms a complex with AKAP450 and PKAIIβ, localized in the Golgi apparatus of cerebellar granule cells. In contrast, the association of InsP₃R1 with PKA in Purkinje cells would require a different macromolecular complex.     

Association of adiponectin (AdipoQ) and sulphonylurea receptor (ABCC8) gene polymorphisms with Type 2 Diabetes in North Indian population of Punjab

In Type 2 Diabetes (T2D), adiponectin (AdipoQ) and sulphonylurea receptor genes (ABCC8) are important targets for candidate gene association studies. The single nucleotide polymorphisms (SNPs) in these genes have been associated with features of the metabolic syndrome across various populations. The present case–control study undertaken in the population of Punjab, evaluates the association of + 45T>G polymorphism in AdipoQ gene; and Exon16-3C>T as well as Exon18C>T polymorphisms in ABCC8 gene with T2D. These SNPs were genotyped in 200 T2D cases and 200 non-diabetic healthy controls using the PCR-RFLP method. The frequency of the minor G-allele for AdipoQ+ 45(T>G) polymorphism was significantly higher in T2D cases (29.0%) than in controls (21.5%) [P = 0.02, OR = 1.49 (1.07–2.04)]. The genetic model analysis revealed that the G-allele cumulatively provides nearly 1.59–1.78 fold increased risk to T2D under the additive (P = 0.009; OR = 1.59, 1.12–2.25 at 95% CI), dominant (TG/GG vs. TT) (P = 0.034, OR = 1.64, 1.04–2.56 at 95% CI) and codominant model (TG vs. TT/GG) (P = 0.014; OR = 1.78, 1.12–2.82 at 95% CI) after adjusting for confounding factors. However, no difference in the distribution of genotype and allele frequencies was observed for both the ABCC8 polymorphisms. The distribution of obesity profiles (BMI, WC and WHR) was also significantly different between cases and controls (P < 0.05). Higher BMI and central obesity were observed to increase the risk of T2D. G-allele of + 45(T>G) polymorphism in the adiponectin gene appears to be associated with increased risk of T2D, but the polymorphisms in sulphonylurea receptor gene do not seem to be associated with T2D in the population of Punjab.