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.
Highlights
The immune system is a complex network that protects the human body against internal and external threats
Killer immunoglobulin-like receptors (KIRs) Genotyping by multiplex ligation-dependent probe amplification (MLPA) Methodology In designing the set of probes for the KIR MLPA we used the following principles
In some cases two of those single nucleotide polymorphisms (SNPs) were combined together in one probe set that consists of three probe parts, to make the probe segregating between the KIR genes
Summary
The immune system is a complex network that protects the human body against internal and external threats. It consists of an innate and an adaptive system. Natural killer (NK) cells constitute the first line of innate immunity against viral infections and tumor development. Recent studies have shown that NK cells can be educated during development and that memory NK cells are able to mount a more effective cytokine response upon reactivation, suggesting adaptive functions [1,2]. The importance of human NK cells has become clear through studies concerning cancer, viral infections and donor/recipient panels in transplantation settings. The cytotoxicity of NK cells is regulated through large families of molecules with a homologous structure, belonging to the C-type lectin-like molecules and immunoglobulin-like receptors [3]
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