Differential microRNA regulation of HLA-C expression and its association with HIV control - PubMed (original) (raw)

. 2011 Apr 28;472(7344):495-8.

doi: 10.1038/nature09914. Epub 2011 Apr 17.

Ram Savan, Ying Qi, Xiaojiang Gao, Yuko Yuki, Sara E Bass, Maureen P Martin, Peter Hunt, Steven G Deeks, Amalio Telenti, Florencia Pereyra, David Goldstein, Steven Wolinsky, Bruce Walker, Howard A Young, Mary Carrington

Affiliations

Differential microRNA regulation of HLA-C expression and its association with HIV control

Smita Kulkarni et al. Nature. 2011.

Abstract

The HLA-C locus is distinct relative to the other classical HLA class I loci in that it has relatively limited polymorphism, lower expression on the cell surface, and more extensive ligand-receptor interactions with killer-cell immunoglobulin-like receptors. A single nucleotide polymorphism (SNP) 35 kb upstream of HLA-C (rs9264942; termed -35) associates with control of HIV, and with levels of HLA-C messenger RNA transcripts and cell-surface expression, but the mechanism underlying its varied expression is unknown. We proposed that the -35 SNP is not the causal variant for differential HLA-C expression, but rather is marking another polymorphism that directly affects levels of HLA-C. Here we show that variation within the 3' untranslated region (UTR) of HLA-C regulates binding of the microRNA hsa-miR-148 to its target site, resulting in relatively low surface expression of alleles that bind this microRNA and high expression of HLA-C alleles that escape post-transcriptional regulation. The 3' UTR variant associates strongly with control of HIV, potentially adding to the effects of genetic variation encoding the peptide-binding region of the HLA class I loci. Variation in HLA-C expression adds another layer of diversity to this highly polymorphic locus that must be considered when deciphering the function of these molecules in health and disease.

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Conflict of interest statement

The authors declare no competing financial interests.

Figures

Figure 1

Figure 1. Variation in the HLA-C 3′UTR differentially affects the expression of a reporter gene

Full length 3′UTRs of various HLA-C alleles cloned into luciferase reporter constructs were transfected into B721.221 cell lines and the stability of the mRNA was estimated by dual luciferase reporter assays. The normalized luciferase activity is presented as fold change of relative light units (RLU). The data represent six replicates in each experimental group, the Mean ±SE are depicted as horizontal and vertical bars for each group, respectively, and one of three comparable experiments performed is shown. Non-parametric Wilcoxon-Mann-Whitney tests were used for statistical comparisons and two tailed p values are indicated. ns = not significant. a. Schematic representations of the luciferase reporter constructs used in this study. b. Fold change in luciferase activity of 3′UTRs of HLA-C alleles as compared to that of Cw*0602.

Figure 2

Figure 2. Disruption of miR-148a target site rescues suppression

a. Partial sequence of mutated 3′UTRs of Cw*0602 and Cw*0702 (06mut and 07mut, respectively) are aligned to 3′UTR sequences of native Cw*0602 and Cw*0702. Identical nucleotides are shown as dots, altered nucleotides are underlined, and deletions are indicated by hyphens (−) for optimal alignment. b. Fold change in luciferase activity of the modified 3′UTR (06mut and 07mut). c. Fold change in luciferase activity of reporters containing wild type Cw*0602 or Cw*0702 3′UTR sequences upon introduction of miR-148a mimic and inhibitor. d. Fold change in luciferase activity of reporters containing 06mut and 07mut 3′UTR sequences upon introduction of miR-148a mimic and inhibitor. Presence (+) or absence (−) of each variable, including a negative control (NC) miRNA, a mimic of miR-148a, or an inhibitor of miR-148a is shown. The data represent six replicates in each experimental group, the Mean ±SE are depicted as horizontal and vertical bars for each group, respectively, and one of three comparable experiments performed is shown. Non-parametric Wilcoxon-Mann-Whitney tests were used for statistical comparisons and two tailed p values are indicated.

Figure 3

Figure 3. miR-148a affects cell surface expression of HLA-C

Histograms of HLA-C cell surface expression on HLA-C homozygous BLCL using flow cytometry are illustrated. In each plot, a NC miRNA that does not bind to the 3′UTR of HLA-C was included. a–b. HLA-Cw*0602 homozygous cells (BLCL-Cw*0602Hom) transfected with either a mimic or an inhibitor of miR-148a (a) or miR-148b (b). c–d. HLA-Cw*0702 homozygous cells (BLCL-Cw*0702Hom) transfected with either a mimic or an inhibitor of miR-148a (c) or miR-148b (d).

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