Extensive Variation in Gene Copy Number at the Killer Immunoglobulin-Like Receptor Locus in Humans

Abstract

Killer immunoglobulin-like receptors (KIRs) are involved in the regulation of natural killer cell cytotoxicity. Within the human genome seventeen KIR genes are present, which all contain a large number of allelic variants. The high level of homology among KIR genes has hampered KIR genotyping in larger cohorts, and determination of gene copy number variation (CNV) has been difficult. We have designed a multiplex ligation-dependent probe amplification (MLPA) technique for genotyping and CNV determination in one single assay and validated the results by next-generation sequencing and with a KIR gene-specific short tandem repeat assay. In this way, we demonstrate in a cohort of 120 individuals a high level of CNV for all KIR genes except for the framework genes KIR3DL3 and KIR3DL2. Application of our MLPA assay in segregation analyses of families from the Centre d’Etude du Polymorphisme Humaine, previously KIR-genotyped by classical techniques, confirmed an earlier reported duplication and resulted in the identification of a novel duplication event in one of these families. In summary, our KIR MLPA assay allows rapid and accurate KIR genotyping and CNV detection, thus rendering improved transplantation programs and oncology treatment feasible, and enables more detailed studies on the role of KIRs in human (auto)immunity and infectious disease.

Deciphering the killer‐cell immunoglobulin‐like receptor system at super‐resolution for natural killer and T‐cell biology

“…KIR genotyping methodology. (a) Representative haplotype-pairs of two individuals are shown along with the expected results from different typing approaches; (i) presence/absence typing by PCR-sequence-specific primers (SSP) or sequence-specific oligonucleotide (SSO) probes, (ii) copy number typing by quantitative PCR (qPCR), 96 multiplex ligation-dependent probe amplification (MLPA) 97 and digital PCR (dPCR), 98 (iii) allele and copy number typing by pyrosequencing and next-generation sequencing (NGS), (iv) imputation infers KIR genotypes from single nucleotide polymorphism (SNP) data. (b) Schemes illustrating different typing approaches.…”

Killer‐cell immunoglobulin‐like receptor assessment algorithms in haemopoietic progenitor cell transplantation: current perspectives and future opportunities

“…The NAHR that governs CNV of KIR appears to be spatially restricted, with an inconsistent degree of CNV between KIR genes. The genes at the extremities of the cluster, such as KIR3DL2, 3DL3, 2DS2, 2DS1 and 2DS4, present limited duplication in comparison with the high frequency of duplication observed in KIR3DL1S1 at the centre of the cluster . The tight restriction of NAHR within the limits of the KIR gene complex suggests this mechanism is of central importance to the genetic evolution of the KIR gene cluster, and may explain why the primary hotspot for recombination is located at the centre of the gene cluster.…”

KIR3DL1 and KIR3DL2 gene copy number variation in axial spondyloarthritis

“…The HLA‐B27 molecule is known to be recognized by KIR3DL1 and KIR3DL2 , and binding of HLA‐B27 to KIR3DL2 activates Th17 cells in AS . The KIR locus is subject to genetic variation, which is because of gene polymorphisms and gene copy number variation (CNV), as we have previously shown . Although KIR genes and polymorphisms have been studied in SpA, nothing is known of gene CNV in relation to this disease.…”

“…Using a multiplex ligation‐dependent probe amplification assay , we genotyped Caucasian AS and axSpA patients and healthy controls for both the presence and CNV of each KIR gene. The study was statistically powered to detect an increase of two KIR3DL1 copies of 20% in patients compared with controls; from 60% in controls to 80% in patients (power 80%, confidence interval 95%, 82 cases, 82 controls).…”