80 research outputs found

    Defining KIR and HLA Class I Genotypes at Highest Resolution via High-Throughput Sequencing.

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    The physiological functions of natural killer (NK) cells in human immunity and reproduction depend upon diverse interactions between killer cell immunoglobulin-like receptors (KIRs) and their HLA class I ligands: HLA-A, HLA-B, and HLA-C. The genomic regions containing the KIR and HLA class I genes are unlinked, structurally complex, and highly polymorphic. They are also strongly associated with a wide spectrum of diseases, including infections, autoimmune disorders, cancers, and pregnancy disorders, as well as the efficacy of transplantation and other immunotherapies. To facilitate study of these extraordinary genes, we developed a method that captures, sequences, and analyzes the 13 KIR genes and HLA-A, HLA-B, and HLA-C from genomic DNA. We also devised a bioinformatics pipeline that attributes sequencing reads to specific KIR genes, determines copy number by read depth, and calls high-resolution genotypes for each KIR gene. We validated this method by using DNA from well-characterized cell lines, comparing it to established methods of HLA and KIR genotyping, and determining KIR genotypes from 1000 Genomes sequence data. This identified 116 previously uncharacterized KIR alleles, which were all demonstrated to be authentic by sequencing from source DNA via standard methods. Analysis of just two KIR genes showed that 22% of the 1000 Genomes individuals have a previously uncharacterized allele or a structural variant. The method we describe is suited to the large-scale analyses that are needed for characterizing human populations and defining the precise HLA and KIR factors associated with disease. The methods are applicable to other highly polymorphic genes.This study was supported by U.S. National Institutes of Health grants U01 AI090905, R01 20 GM109030, R01 AI17892 and U19 AI119350. Authors Steven Norberg and Mostafa Ronaghi are 21 employees of Illumina Inc.This is the author accepted manuscript. It is currently under an indefinite embargo pending publication by Elsevier

    Conservation, Extensive Heterozygosity, and Convergence of Signaling Potential All Indicate a Critical Role for KIR3DL3 in Higher Primates

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    Natural killer (NK) cell functions are modulated by polymorphic killer cell immunoglobulin-like receptors (KIR). Among 13 human KIR genes, which vary by presence and copy number, KIR3DL3 is ubiquitously present in every individual across diverse populations. No ligand or function is known for KIR3DL3, but limited knowledge of expression suggests involvement in reproduction, likely during placentation. With 157 human alleles, KIR3DL3 is also highly polymorphic and we show heterozygosity exceeds that of HLA-B in many populations. The external domains of catarrhine primate KIR3DL3 evolved as a conserved lineage distinct from other KIR. Accordingly, and in contrast to other KIR, we show the focus of natural selection does not correspond exclusively to known ligand binding sites. Instead, a strong signal for diversifying selection occurs in the D1 Ig domain at a site involved in receptor aggregation, which we show is polymorphic in humans worldwide, suggesting differential ability for receptor aggregation. Meanwhile in the cytoplasmic tail, the first of two inhibitory tyrosine motifs (ITIM) is conserved, whereas independent genomic events have mutated the second ITIM of KIR3DL3 alleles in all great apes. Together, these findings suggest that KIR3DL3 binds a conserved ligand, and a function requiring both receptor aggregation and inhibitory signal attenuation. In this model KIR3DL3 resembles other NK cell inhibitory receptors having only one ITIM, which interact with bivalent downstream signaling proteins through dimerization. Due to the extensive conservation across species, selection, and other unusual properties, we consider elucidating the ligand and function of KIR3DL3 to be a pressing question

    A KIR B centromeric region present in Africans but not Europeans protects pregnant women from pre-eclampsia.

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    In sub-Saharan Africans, maternal mortality is unacceptably high, with >400 deaths per 100,000 births compared with <10 deaths per 100,000 births in Europeans. One-third of the deaths are caused by pre-eclampsia, a syndrome arising from defective placentation. Controlling placentation are maternal natural killer (NK) cells that use killer-cell immunoglobulin-like receptor (KIR) to recognize the fetal HLA-C molecules on invading trophoblast. We analyzed genetic polymorphisms of maternal KIR and fetal HLA-C in 484 normal and 254 pre-eclamptic pregnancies at Mulago Hospital, Kampala, Uganda. The combination of maternal KIR AA genotypes and fetal HLA-C alleles encoding the C2 epitope associates with pre-eclampsia [P = 0.0318, odds ratio (OR) = 1.49]. The KIR genes associated with protection are located in centromeric KIR B regions that are unique to sub-Saharan African populations and contain the KIR2DS5 and KIR2DL1 genes (P = 0.0095, OR = 0.59). By contrast, telomeric KIR B genes protect Europeans against pre-eclampsia. Thus, different KIR B regions protect sub-Saharan Africans and Europeans from pre-eclampsia, whereas in both populations, the KIR AA genotype is a risk factor for the syndrome. These results emphasize the importance of undertaking genetic studies of pregnancy disorders in African populations with the potential to provide biological insights not available from studies restricted to European populations.This work was supported by the Wellcome Trust (090108/Z/09/Z, 085992/Z/08/Z, 089821/Z/09/Z), the British Heart Foundation (PG/ 09/077/27964), the Centre for Trophoblast Research at the University of Cambridge, a Wellcome Trust Uganda PhD Fellowship in Infection and Immunity held by Annettee Nakimuli, funded by a Wellcome Trust Strategic Award (084344), the US National Institutes of Health (AI017892), and the UK Medical Research Council (G0901682).This is the accepted manuscript of a paper published in PNAS (A Nakimuli, O Chazara, SE Hiby, L Farrell, S Tukwasibwe, J Jayaraman, JA Traherne, J Trowsdale, F Colucci, Emma Lougee, RW Vaughan, AM Elliott, J Byamugishaa, P Kaleebu, F Mirembe, N Nemat-Gorgani, P Parham, PJ Norman, A Moffett, PNAS 2015, 112, 845-850

    Large Proteins Have a Great Tendency to Aggregate but a Low Propensity to Form Amyloid Fibrils

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    The assembly of soluble proteins into ordered fibrillar aggregates with cross-β structure is an essential event of many human diseases. The polypeptides undergoing aggregation are generally small in size. To explore if the small size is a primary determinant for the formation of amyloids under pathological conditions we have created two databases of proteins, forming amyloid-related and non-amyloid deposits in human diseases, respectively. The size distributions of the two protein populations are well separated, with the systems forming non-amyloid deposits appearing significantly larger. We have then investigated the propensity of the 486-residue hexokinase-B from Saccharomyces cerevisiae (YHKB) to form amyloid-like fibrils in vitro. This size is intermediate between the size distributions of amyloid and non-amyloid forming proteins. Aggregation was induced under conditions known to be most effective for amyloid formation by normally globular proteins: (i) low pH with salts, (ii) pH 5.5 with trifluoroethanol. In both situations YHKB aggregated very rapidly into species with significant β-sheet structure, as detected using circular dichroism and X-ray diffraction, but a weak Thioflavin T and Congo red binding. Moreover, atomic force microscopy indicated a morphology distinct from typical amyloid fibrils. Both types of aggregates were cytotoxic to human neuroblastoma cells, as indicated by the MTT assay. This analysis indicates that large proteins have a high tendency to form toxic aggregates, but low propensity to form regular amyloid in vivo and that such a behavior is intrinsically determined by the size of the protein, as suggested by the in vitro analysis of our sample protein

    An archaic HLA class I receptor allele diversifies natural killer cell-driven immunity in First Nations peoples of Oceania

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    Genetic variation in host immunity impacts the disproportionate burden of infectious diseases that can be experienced by First Nations peoples. Polymorphic human leukocyte antigen (HLA) class I and killer cell immunoglobulin-like receptors (KIRs) are key regulators of natural killer (NK) cells, which mediate early infection control. How this variation impacts their responses across populations is unclear. We show that HLA-A∗24:02 became the dominant ligand for inhibitory KIR3DL1 in First Nations peoples across Oceania, through positive natural selection. We identify KIR3DL1∗114, widespread across and unique to Oceania, as an allele lineage derived from archaic humans. KIR3DL1∗114+NK cells from First Nations Australian donors are inhibited through binding HLA-A∗24:02. The KIR3DL1∗114 lineage is defined by phenylalanine at residue 166. Structural and binding studies show phenylalanine 166 forms multiple unique contacts with HLA-peptide complexes, increasing both affinity and specificity. Accordingly, assessing immunogenetic variation and the functional implications for immunity are fundamental toward understanding population-based disease associations

    Testicular extramedullary plasmacytoma

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    Extramedullary plasmacytomas are plasma cell tumors that arise outside the bone marrow. They are most often located in the head and neck. Extramedullary plasmacytomas account for approximately 3 percent of plasma cell malignancies. The diagnosis of an extramedullary plasmacytoma depends upon the demonstration of a monoclonal plasma cell tumor at an extramedullary site with no evidence of multiple myeloma based upon bone marrow examination and roentgenographic studies, absence of an M-protein in serum and/ or urine, and no related organ or tissue impairment. These tumors are highly responsive to local radiation therapy and rarely progress or recur. A 77 year old man referred for gradual enlargement of scrotum in April, 2004. He had no fever, weight loss or loss of appetite during the period. No tenderness, warmth or erythem of scrotum was observed on physical examination. No lymphadenopathy was detected. Later he had temporary relief because of antibiotic therapy . 7 months later, he referred again following the aggravation of his symptom. Ultrasonography of his scrotum was done and enlargement in the left testis was reported. Lab data including CBC,diff,ESR,AFP were normal but βHCG titer was reported higher than normal . He underwent radical orchiectomy and the specimen was sent for pathological study. Spermatocytic seminoma and plasmacytoma were suspected, but plasmacytoma was reported after the IHC . In order to rule out multiple myeloma, whole body bone scanning with sestamibi and urine Bence Jones protein were examined, all of which were negative. In bone marrow aspiration, the cellularity was normal and no pathologic lesion was reported. Considering clinical and paraclinical evidences, the diagnosis was extramedullary plasmacytoma
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