Molecular genetics and population studies of the KIR and HLA gene complexes

Division Of Basic Sciences - Nci

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Abstract

The classical HLA class I molecules, HLA-A, -B and -C, present antigenic peptides to CD8+ T cells, eliciting an adaptive immune response. The genes encoding these molecules are highly polymorphic, resulting in extensive diversity of the peptide repertoire, both within individuals and at the population level. Peptide loading of HLA class I molecules takes place primarily in the endoplasmic reticulum within the peptide loading complex (PLC). Tapasin is a critical component of the PLC, which performs its peptide editing function by association with peptide-empty HLA class I, stabilizing its structure, and promoting dissociation of low affinity peptides. HLA class I allotypes vary in level of cell surface expression in the absence of tapasin. Some allotypes are expressed at very low levels on the surface of tapasin-deficient cells (tapasin-dependent allotypes), while others exhibit normal expression on these cells (tapasin-independent allotypes). The exact molecular determinants of tapasin dependence (TD) remain unknown, although amino acids in the peptide binding groove near the peptide C terminus appear to contribute most to this phenomenon. For example, a single amino acid change (D116Y) located in this region distinguishes the highly tapasin-dependent B4402 allotype from the tapasin-independent B4405 allotype. Tapasin function can be targeted by viruses as a means of downmodulating HLA class I and evading cytotoxic CD8+ T cell (CTL) responses. Similarly, loss of tapasin expression has been observed in various human cancers. Thus, tapasin-independent HLA class I allotypes may be advantageous in terms of eliciting CTL responses against virally infected cells or tumor cells when tapasin function has been diminished. Allotype-specific regulation of the peptide repertoire by tapasin may also affect the quality of CTL responses. We previously quantified the level of TD across all common HLA allotypes present in European and African Americans (N = 97) and tested the functional significance of differential HLA class I TD and its impact on disease. We have now completed TD values for an additional 153 alleles, which represent common alleles in worldwide populations. In addition, we are currently analyzing other components of the PLC including TAPBPR and calreticulin. Using CRISPR technology, we have now obtained knockouts of TAPBPR and calreticulin in 721.221 cells. These cell lines will be used in further downstream analyses. The development of methods for assessing the nature and extent of KIR genomic diversity has been limited by the complexity of the region. Members of this gene family share a high amount of sequence similarity. Also, there is remarkable diversity with respect to KIR gene content at the genomic level. Therefore, it is extremely challenging to determine the absence or presence of KIR genes based on short-read sequencing data. We have now developed a computational approach that allows predictions for KIR gene content based on either whole-genome or whole-exome sequencing data. This approach utilizes same-length sequence fragments (k-mers) that are unique to individual KIR genes. To validate our approach, we used 480 exome sequencing samples and 340 whole genome sequencing samples that were previously genotyped in our laboratory. We achieved 100% accuracy amongst whole genome samples and more than 99% accuracy amongst the whole exome sequencing samples. This method will allow us to interrogate publicly available datasets for disease association analyses. Human natural killer (NK) cells are essential for controlling infection, cancer, and fetal development. NK cell functions are modulated by interactions between polymorphic inhibitory killer cell immunoglobulin-like receptors (KIR) and polymorphic HLA-A, -B, and -C ligands expressed on tissue cells. All HLA-C alleles encode a KIR ligand and contribute to reproduction and immunity. In contrast, only some HLA-A and -B alleles encode KIR ligands and they focus on immunity. Two broad groups of KIR haplotypes are present in every human population. KIR A haplotypes carry all four of the HLA-class I-specific inhibitory receptors and are associated with resistance to infectious diseases. KIR B haplotypes are more variable in their gene number, carrying two or more genes for inhibitory receptors as well as various activating receptor genes, and favor fetal development. Additional to gene content variation, polymorphism of both receptors and ligands can directly affect NK cell activity. Thus, by varying the number, density, specificity, strength, or signal transduction properties of the receptor-ligand interaction, genetic variation of KIR and HLA class I can predetermine functional differences in NK cell repertoires between individuals. This genetic diversity is substantial among populations, but in most studies Asian populations are underrepresented. By high-resolution analysis of KIR and HLA-A, -B, and -C genes, we show that the Chinese Southern Han (CHS) are significantly enriched for interactions between inhibitory KIR and HLA-A and -B. This enrichment has had substantial input through population admixture with neighboring populations, who contributed HLA class I haplotypes expressing the KIR ligands B4601 and B58:01, which subsequently rose to high frequency by natural selection. Consequently, over 80% of Southern Han HLA haplotypes encode more than one KIR ligand. Complementing the high number of KIR ligands, the CHS KIR locus combines a high frequency of genes expressing potent inhibitory KIR, with a low frequency of those expressing activating KIR. The Southern Han centromeric KIR region encodes strong, conserved, inhibitory HLA-C-specific receptors, and the telomeric region provides a high number and diversity of inhibitory HLA-A and -B-specific receptors. In all these characteristics, the CHS represent other East Asians, whose NK cell repertoires are thus enhanced in quantity, diversity, and effector strength, likely augmenting resistance to endemic viral infections. In conclusion, our high-resolution analysis of KIR and HLA class I combinatorial diversity has uncovered a distinctive enhancement of the interactions between inhibitory KIR and HLA-A and -B in East Asians. These genetically determined distinctions likely underlie differences across human populations in their susceptibility to infections and immune- mediated diseases.