High population frequencies of MICA copy number variations originate from independent recombination events.

MICA is a stress-induced ligand of the NKG2D receptor that stimulates NK and T cell responses and was identified as a key determinant of anti-tumor immunity. The MICA gene is located inside the MHC complex and is in strong linkage disequilibrium with HLA-B. While an HLA-B*48-linked MICA deletion-haplotype was previously described in Asian populations, little is known about other MICA copy number variations. Here, we report the genotyping of more than two million individuals revealing high frequencies of MICA duplications (1%) and MICA deletions (0.4%). Their prevalence differs between ethnic groups and can rise to 2.8% (Croatia) and 9.2% (Mexico), respectively. Targeted sequencing of more than 70 samples indicates that these copy number variations originate from independent nonallelic homologous recombination events between segmental duplications upstream of MICA and MICB. Overall, our data warrant further investigation of disease associations and consideration of MICA copy number data in oncological study protocols.

HLA-E diversity unfolded: Identification and characterization of 170 novel HLA-E alleles.

HLA-E is a member of the nonclassical HLA class Ib genes. Even though it is structurally highly similar to the classical HLA class Ia genes, it is less diverse and only 45 alleles and 12 proteins were known in December 2019 (IPD-IMGT/HLA, release 3.38.0). Since 2017, we have genotyped over 3 million voluntary stem cell donors for HLA-E by sequencing the most relevant allele-determining bases of exons 2 and 3. As expected, most donors harbor the two predominant alleles HLA-E*01:01 and/or HLA-E*01:03. However, in 1666 (0.05%) of our samples we detected 345 distinct novel HLA-E sequences. The most frequent one was identified in 162 samples and has by now been named HLA-E*01:114. To characterize these novel alleles in full-length, we used both short-read Illumina and long-read PacBio sequencing to obtain fully phased and highly accurate sequences. This resulted in 234 submissions to IPD-IMGT/HLA comprising 170 novel HLA-E alleles, which encode for 93 novel HLA-E proteins, as well as 64 confirmations or sequence extensions. Consequently, the number of HLA-E alleles in the database (release 3.42.0) has now increased to 256 HLA-E alleles and 110 HLA-E proteins.

High-Throughput MICA/B Genotyping of Over Two Million Samples: Workflow and Allele Frequencies.

MICA and MICB are ligands of the NKG2D receptor and thereby influence NK and T cell activity. MICA/B gene polymorphisms, expression levels and the amount of soluble MICA/B in the serum have been linked to autoimmune diseases, infections, and cancer. In hematopoietic stem cell transplantation, MICA matching between donor and patient has been correlated with reduced acute and chronic graft-vs.-host disease and improved survival. Hence, we developed an extremely cost-efficient high-throughput workflow for genotyping MICA/B for newly registered potential stem cell donors. Since mid-2017, we have genotyped over two million samples using NGS amplicon sequencing for MICA/B exons 2-5. In donors of German origin, MICA*008 is the most common MICA allele with a frequency of 42.3%. It is followed by MICA*002 (11.7%) and MICA*009 (8.8%). The three most common MICB alleles are MICB*005 (43.9%), MICB*004 (21.7%), and MICB*002 (18.9%). In general, MICB is less diverse than MICA and only 6 alleles, instead of 15, account for a cumulative allele frequency of 99.5%. In 0.5% of the samples we observed at least one allele of MICA or MICB which has so far not been reported to the IPD/IMGT-HLA database. By providing MICA/B typed voluntary donors, clinicians become empowered to include MICA/B into their donor selection process to further improve unrelated hematopoietic stem cell transplantation.