Association between a variation in the phosphodiesterase 4D gene and bone mineral density

Abstract

Background: Fragility fractures caused by osteoporosis are a major cause of morbidity and mortality in aging populations. Bone mineral density (BMD) is a useful surrogate marker for risk of fracture and is a highly heritable trait. The genetic variants underlying this genetic contribution are largely unknown.

Methods: We performed a large-scale association study investigating more than 25,000 single nucleotide polymorphisms (SNPs) located within 16,000 genes. Allele frequencies were estimated in contrasting DNA pools from white females selected for low (<0.87 g/cm2, n = 319) and high (> 1.11 g/cm2, n = 321) BMD at the lumbar spine. Significant findings were verified in two additional sample collections.

Results: Based on allele frequency differences between DNA pools and subsequent individual genotyping, one of the candidate loci indicated was the phosphodiesterase 4D (PDE4D) gene region on chromosome 5q12. We subsequently tested the marker SNP, rs1498608, in a second sample of 138 white females with low (<0.91 g/cm2) and 138 females with high (>1.04 g/cm2) lumbar spine BMD. Odds ratios were 1.5 (P = 0.035) in the original sample and 2.1 (P = 0.018) in the replication sample. Association fine mapping with 80 SNPs located within 50 kilobases of the marker SNP identified a 20 kilobase region of association containing exon 6 of PDE4D. In a second, family-based replication sample with a preponderance of females with low BMD, rs1498608 showed an opposite relationship with BMD at different sites (p = 0.00044-0.09). We also replicated the previously reported association of the Ser37Ala polymorphism in BMP2, known to interact biologically with PDE4D, with BMD.

Conclusion: This study indicates that variants in the gene encoding PDE4D account for some of the genetic contribution to bone mineral density variation in humans. The contrasting results from different samples indicate that the effect may be context-dependent. PDE4 inhibitors have been shown to increase bone mass in normal and osteopenic mice, but up until now there have been no reports implicating any member of the PDE4 gene family in human osteoporosis.

Figure 1

Overview of DNA pool-based large-scale association strategy. Phases 1 and 2 are conducted using DNA pools yielding allele frequencies, all subsequent steps involve genotyping of individual samples. hME, homogeneous MassEXTEND; MS, mass spectrometry. See text for more details.

Figure 2

Association fine mapping of PDE4D region. Eighty public domain SNPs in a 100-kb window around the initial marker SNP (indicated as a bold symbol at the center of the plot) were compared between the UK low and high BMD pools. Ten of 80 SNPs were significant at P = 0.05 (horizontal dashed line). The x-axis corresponds to the chromosomal position of each SNP, the y-axis to the test P-values (--log₁₀ scale). The continuous dark line presents the results of a goodness-of-fit test for an excess of significant associations (at a 5% significance level) in a 10-kb sliding window assessed at 1-kb increments. The continuous light gray line is the result of a nonlinear smoothing function showing a weighted average of the P-values across the region. The color of each point corresponds to the minor allele frequency of each SNP in the control sample (see legend below graph). Vertical dashed lines are placed every 20 kb. The LocusLink gene annotation for NCBI genome build 34 shows the location of exon six. All investigated SNPs are located within introns.