Complex expression of natural killer receptor genes in single natural killer cells

Abstract

Human natural killer (NK) cells express several inhibitory and non-inhibitory NK receptors per cell. Understanding the expression patterns of these receptor genes in individual cells is important to understanding their function. Using a single-cell reverse transcription-polymerase chain reaction (RT-PCR) method, we analysed the expression of nine NK receptor genes in 38 resting CD56+ NK cells from peripheral blood of normal donors. We observed highly diverse patterns of receptor expression in these cells. No NK receptor is expressed universally in every CD56+ NK cell. The expressed receptor types per cell varied from two to eight. We specifically analysed the distribution of inhibitory (DL) and non-inhibitory (DS) killer immunoglobulin-like receptors (KIR). The frequency of individual receptor expression varied from 26% for 2DS2 to 68% for both 2DL1 and 2DL4. A comparison of the coexpression of DL and DS receptors showed a significant association in the expression of 2DL2 and 2DS2 (chi2=16.6; P<0.001) genes but no association between 2DL1 and 2DS1 or between 3DL1 and 3DS1 genes. Coexpression analysis of the 2DL1 and 2DL2 genes in 2DL4+ and 2DL4- cells showed a strong association in 2DL4+ but not in 2DL4- cells, suggesting a differential effect of the 2DL4 gene on the expression of 2DL1 and 2DL2 genes. Single-cell RT-PCR is a powerful tool to study multiple receptor gene expression ex vivo in individual NK cells and provides information about the expression pattern of KIR receptors that may suggest mechanisms of gene expression responsible for generation of the KIR repertoire.

Figure 1. NK receptor gene expression in single cells. RNA from two representative NK cells each from two subjects was subjected to gene-specific RT-PCR using primers for NK receptors: lane 1, KIR2DL1; lane 2, KIR3DL1; lane 3, KIR2DL2; lane 4, KIR2DL4; lane 5, KIR3DS1; lane 6, KIR2DS1; lane 7, KIR2DS2; lane 8, CD94; lane 9, NKG2A; lane 10, β-actin; and lane 11, intron.

Figure 2. Distribution of NK receptor transcripts in single NK cells. A and B are donors homozygous for HLA-Cw7 and C is heterozygous for HLA-Cw alleles. HLA typings of the donors is given in the Materials and Methods section. RNA from single cells was used to perform RT-PCR. Each row represents a single cell's repertoire of individual NK receptors given in labelled columns and is depicted by a shaded (expression) or blank (no expression) rectangle for individual receptor genes.