Dataset Information

Tuning of NK-Specific HLA-C Expression by Alternative mRNA Splicing.

ABSTRACT: A complex system regulating HLA-C expression in NK cells, driven by an NK-specific promoter that produces alternatively spliced variants of the 5'-UTR has been recently identified. Exon content of the NK-specific 5'-UTR varies strikingly across HLA-C alleles, with some exons being allele specific. In order to investigate the possibility that allelic variation in the 5'-UTR modulates HLA-C expression levels, cDNAs containing several distinct classes of 5'-UTR were compared. Subtle changes in 5'-UTR content had a significant effect on the expression of HLA-C * 03 and HLA-C * 12 cDNA clones, suggesting that alternative splicing can fine-tune the level of protein expression. The HLA-C * 06 allele was found to be highly expressed in relation to the other alleles studied. However, its increased expression was primarily associated with differences in the peptide-binding groove. Although the impact of allele-specific alternative splicing of NK-Pro transcripts on protein levels can be modest when compared with the effect of changes in peptide-loading, alternative splicing may represent an additional regulatory mechanism to fine-tune HLA-C levels within NK cells in distinct tissue environments or at different stages of maturation in order to achieve optimal levels of missing-self recognition.

SUBMITTER: Goodson-Gregg FJ

PROVIDER: S-EPMC6966967 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Tuning of NK-Specific HLA-C Expression by Alternative mRNA Splicing.

Goodson-Gregg Frederick J FJ Rothbard Brian B Zhang Amy A Wright Paul W PW Li Hongchuan H Walker-Sperling Victoria E VE Carrington Mary M Anderson Stephen K SK

Frontiers in immunology 20200110

A complex system regulating HLA-C expression in NK cells, driven by an NK-specific promoter that produces alternatively spliced variants of the 5'-UTR has been recently identified. Exon content of the NK-specific 5'-UTR varies strikingly across <i>HLA-C</i> alleles, with some exons being allele specific. In order to investigate the possibility that allelic variation in the 5'-UTR modulates HLA-C expression levels, cDNAs containing several distinct classes of 5'-UTR were compared. Subtle changes ...[more]

PMID: 31998314

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Project description:Splice isoform structure and abundance can be affected by either noncoding or masquerading coding variants that alter the structure or abundance of transcripts. When these variants are common in the population, these nonconstitutive transcripts are sufficiently frequent so as to resemble naturally occurring, alternative mRNA splicing. Prediction of the effects of such variants has been shown to be accurate using information theory-based methods. Single nucleotide polymorphisms (SNPs) predicted to significantly alter natural and/or cryptic splice site strength were shown to affect gene expression. Splicing changes for known SNP genotypes were confirmed in HapMap lymphoblastoid cell lines with gene expression microarrays and custom designed q-RT-PCR or TaqMan assays. The majority of these SNPs (15 of 22) as well as an independent set of 24 variants were then subjected to RNAseq analysis using the ValidSpliceMut web beacon (http://validsplicemut.cytognomix.com), which is based on data from the Cancer Genome Atlas and International Cancer Genome Consortium. SNPs from different genes analyzed with gene expression microarray and q-RT-PCR exhibited significant changes in affected splice site use. Thirteen SNPs directly affected exon inclusion and 10 altered cryptic site use. Homozygous SNP genotypes resulting in stronger splice sites exhibited higher levels of processed mRNA than alleles associated with weaker sites. Four SNPs exhibited variable expression among individuals with the same genotypes, masking statistically significant expression differences between alleles. Genome-wide information theory and expression analyses (RNAseq) in tumor exomes and genomes confirmed splicing effects for 7 of the HapMap SNP and 14 SNPs identified from tumor genomes. q-RT-PCR resolved rare splice isoforms with read abundance too low for statistical significance in ValidSpliceMut. Nevertheless, the web-beacon provides evidence of unanticipated splicing outcomes, for example, intron retention due to compromised recognition of constitutive splice sites. Thus, ValidSpliceMut and q-RT-PCR represent complementary resources for identification of allele-specific, alternative splicing.

Dataset Information

Tuning of NK-Specific HLA-C Expression by Alternative mRNA Splicing.

Publications

Tuning of NK-Specific HLA-C Expression by Alternative mRNA Splicing.

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