The recent advance in genetic association study has uncovered more and more susceptibility genes for complex diseases such as SLE. In populations of European ancestry, rs3733197 and rs17266594 in BANK1
were found to be associated with SLE (OR=1.23, P
=4.67 × 10−5
; OR=1.42, P
=4.74 × 10−11
In our study, the contributions of rs3733197 and rs17266594 were verified (OR=1.19, P
=0.021; OR=1.65, P
=4.67 × 10−9
), with a larger effect size for rs17266594 in Chinese (OR=1.65, CI=1.38–1.96) than that in the Caucasians (OR=1.42, CI=1.28–1.58).
Concerning the independent effects of SNPs, Kozyrev et al.4
reported that none of the SNPs studied in BANK1
(rs3733197, rs17266594 and rs10516487) were independent of each other in conditional logistic regression analysis as a result of the LD between them. In our study, the effects of rs3733197 and rs17266594 did not explain each other. This population difference could be a result of the larger effect size observed in Chinese, and thus a more readily detectable independence between the SNPs by statistical methods. In addition, the difference in LD patterns may also play a critical role. In Chinese, LD between rs3733197 and rs17266594 is much weaker than that in Caucasian (HapMap-HCB: r2
=0.08; HapMap-CEU: r2
=0.36, ). In addition, rs3733197 also has a weak LD with the rest of the SNPs () and therefore it is likely to confer SLE risk independently in the BANK1 region.
In addition to statistical analyzes of their independent effects, possible functional implications of the SNPs were also considered to better justify their roles in SLE. SNP rs3733197 causes a substitution in the ankyrin domain (A383T). The alanine residue at this position is well conserved in all the mammals examined, including Monodelphis domestica
(Gray short-tailed opossum), suggesting a functional constraint on this position during evolutionary courses. The mutations in ankyrin motifs have also been shown to alter interactions with IP3
R, thus affecting cytoplasmic calcium mobilization in cardiac arrhythmia and sudden cardiac death.13
These data supports the possible functional importance of rs3733197 and its independent contribution to the risk of SLE. On the other hand, although rs17266594 (the branch point-site polymorphism) confers a strong association with SLE, it is hard to distinguish a direct association at the functional level from an indirect association caused by its strong LD with rs10516487 (r2
=1 in HapMap-HCB). The non-synonymous substitution because of rs10516487 (R61H) is at a highly variable position across species. On the same position in its orthologs, it is a histidine in dog, leucine in horse, proline in mouse and rat, cysteine in cow, and serine in Opossum. Variations at the position may indicate a relaxation of selection pressure on this amino acid residue, although it can also be explained by functional diversities among the orthologs. On the other hand, it has been shown that the expression of two isoforms of BANK1
transcripts are associated with the genotypes of rs17266594, the branch point-site SNP,4
which thus seems to be a more likely candidate responsible for the association of SLE.
It is noted that besides rs3733197 and rs17266594, rs4522865 also contributed to SLE susceptibility, which was not reported in the earlier study. SNP rs4522865 had the most significant P-value (P=8.49 × 10−4, ) in our GWAS and was successfully replicated by further genotyping non-overlapping samples (P=2.93 × 10−3, ). Furthermore, it was found that rs4522865 had a weak LD with rs10516487 (r2=0.25, HapMap-HCB, ) and thus the effect of rs4522865 was unlikely to be dependent on rs10516487. This was consistent with the result of conditional analysis, in which it was found that after controlling the effect of rs4522865, the effects of most of the other SNPs in the BANK1 region were gone except rs10516487. Therefore, rs4522865 is likely to be the other independent contributor to SLE susceptibility besides rs3733197 and rs17266594. Locating at the first intron of BANK1, rs4522865 may have a role in the expression of BANK1. It is interesting to find out several candidates having independent contribution to SLE association within the same locus, however, further functional studies and deep sequencing are required to find out the genuine functional variants in this region.
In the case of TNFSF4
, the minor alleles of rs844648 and rs2205960 tagged an over-transmitted upstream haplotype in SLE cases in Caucasians (MAF−case
=7.0 × 10−3
In addition, rs844644, which has a high LD with rs844648 in our data and the Caucasians (r2
=0.92 and 0.70, ), had been shown to be the single genotyped variant that showed a marginal P
-value after conditional analysis (P
=0.015, Graham et al.
,5 Supplementary Table 6
). In our study, the association of rs844648 and rs2205960 were confirmed (MAF−case
=2.47 × 10−3
=2.41 × 10−4
, respectively). However, after controlling the effect of rs2205960, the effect of rs844648 became insignificant in both of our GWAS and the replication data. The moderate independent effect of rs844644 in Caucasian study was also not observed by GWAS in our population. It is difficult to tell whether this is a genuine difference between populations or only inconsistent statistical results because of the moderate effect sizes of rs844644 and rs844648, which would reduce the power of the conditional analyzes. SNP rs2205960 is located 38.6
kb upstream of TNFSF4
and probably involved in a cis
-acting factor of transcription regulation. Together with the results of conditional analysis, rs2205960 is therefore likely to be a major variant contributing to disease susceptibility in this locus.
In conclusion, this study evaluated SNPs from BANK1 and TNFSF4 to further define their roles in SLE risk. Their associations with SLE are confirmed in Chinese; nevertheless differences with the earlier studies are also showed, especially in terms of individual effects of SNPs. This suggests that multiple variants are probably present and affect the genes by different mechanisms. Further studies are needed to confirm or dissect out the genuine functional variants and to understand how these polymorphisms affect SLE.