Significance of the myxovirus resistance A (MxA) gene -123C>a single-nucleotide polymorphism in suppressed interferon beta induction of severe acute respiratory syndrome coronavirus infection

Abstract

Myxovirus resistance A (MxA) is an antiviral protein induced by interferon alpha and beta (IFN-alpha, IFN-beta) that can inhibit viral replication. The minor alleles of the -88G>T and -123C>A MxA promoter single-nucleotide polymorphisms (SNPs) are associated with increased promoter activity and altered response to IFN-alpha and IFN-beta treatment. Here, we demonstrate that the -123A minor allele provided stronger binding affinity to nuclear proteins extracted from IFN-beta-untreated cells than did the wild-type allele, whereas the -88T allele showed preferential binding after IFN-beta stimulation. Endogenous IFN-alpha and IFN-beta induction can be suppressed in severe acute respiratory syndrome (SARS) coronavirus infection. In support of our in vitro findings, a large case-control genetic-association study for SARS coronavirus infection confirmed that the -123A minor-allele carriers were significantly associated with lower risk of SARS coronavirus infection, whereas the -88T minor-allele carriers were insignificant after adjustment for confounding effects. This suggests that -123C>A plays a more important role in modulating basal MxA expression, thus contributing more significantly to innate immune response against viral infections that suppress endogenous IFN-alpha and IFN-beta induction such as SARS coronavirus.

Figure 1.

Electrophoretic mobility shift assay (EMSA) for the myxovirus resistance A (MxA) -88G>T and -123C>A single-nucleotide polymorphisms (SNPs). A , The sequence of the probes used in EMSA. S, sense; A, anti-sense. The SNPs are underlined in each oligonucleotide for the MxA SNPs. B , EMSA for the -88G>T SNP, using nuclear extracts from untreated 293 cells. Comparison of lanes 3 and 4 and lanes 5 and 6 showed that the -88T probe bound slightly stronger than did the -88G probe. Comparison between lanes 9 and 10 and lanes 11 and 12 also showed that the -88T probe bound slightly stronger (also shown in the bar chart). However, the intensity of the upshifted band was low, and the competition by cold probes was weak. C , EMSA for the -123C>A SNP, using nuclear extracts from untreated 293 cells. The presence of 2 upshifted bands suggested that there might be 11 transcription factor complexes binding to the probes. The stronger band intensity of the -123A probes and the stronger competition by the cold -123A probe suggest that the binding affinity of -123A allele is stronger than that of -123C. D , EMSA for -88G>T SNP, using nuclear extracts from IFN-β-treated 293 cells. There was an upshifted band of the -88T probe, which could be competed by the cold probe. However, the -88G probe was not upshifted. E , EMSA for the -123C>A SNP, using nuclear extracts from IFN-β-treated 293 cells. The upshift pattern of the probes is similar to that with untreated 293 cells nuclear extract, indicating that the binding affinity of the probe is not significantly affected by IFNb treatment. In the figures, the arrows indicate the upshifted bands being competed away by the cold (unlabeled) probes. The relative intensities of the upshifted bands were quantified, and the results were plotted as bar charts. Asterisks indicate the intensity of the upshifted band used to normalize that of other bands in the respective EMSA assays.

Table 1.

Demographic Feature and Clinical Profile of Patients with Severe Acute Respiratory Syndrome and Control Subjects Who Were Successfully Genotyped

Figure 2.

Electrophoresis of the digested polymerase chain reaction (PCR) products of PCR restriction fragment length polymorphism for the myxovirus resistance A (MxA) -88G>T single-nucleotide polymorphism (SNP) and the MxA -123C>A SNP.

Table 2.

List of Clinical Outcome Measures of Severe Acute Respiratory Syndrome (SARS) Patients

Figure 3.

Expression of Myxovirus resistance-A (MxA) and induction of MxA messenger RNA (mRNA) by IFN-β in the 293 cell line.

Table 3.

Numbers of Household Subjects (Genetically Related or Unrelated) and Independent Subjects

Table 4.

Risk Association Analyses of the Myxovirus Resistance A Promoter -88G>T and -123C>A Single-Nucleotide Polymorphisms

Table 5.

Risk Association Analyses of Myxovirus Resistance A (MxA) Promoter -88G>T and -123C>A Single-Nucleotide Polymorphisms Adjusted for Age and Sex in Genetically Unrelated Subjects

Table 6.

Risk Association Analyses of OAS-1 +1295A>G and OAS-1 +590A>G Single-Nucleotide Polymorphism (SNP) Using X² Test and Logistic Regression Adjusted for Genetic

Table 7.

Risk Association Analyses of Myxovirus Resistance A (MxA) Promoter -88G>T and -123C>A Single-Nucleotide Polymorphisms (SNPs) and the OAS-1 SNPs Adjusted for Age and Sex in Genetically Unrelated Subjects

Table 8.

Haplotype Risk Association Analyses of Genetically Unrelated Subjects