Whole genome survey of copy number variation in the spontaneously hypertensive rat: relationship to quantitative trait loci, gene expression, and blood pressure

Abstract

Copy number variation has emerged recently as an important genetic mechanism leading to phenotypic heterogeneity. The aim of our study was to determine whether copy number variants (CNVs) exist between the spontaneously hypertensive rat (SHR) and its control strain, the Wistar-Kyoto rat, whether these map to quantitative trait loci in the rat and whether CNVs associate with gene expression or blood pressure differences between the 2 strains. We performed a comparative genomic hybridization assay between SHR and Wistar-Kyoto strains using a whole-genome array. In total, 16 CNVs were identified and validated (6 because of a relative loss of copy number in the SHR and 10 because of a relative gain). CNVs were present on rat autosomes 1, 3, 4, 6, 7, 10, 14, and 17 and varied in size from 10.0 kb to 1.6 Mb. Most of these CNVs mapped to chromosomal regions within previously identified quantitative trait loci, including those for blood pressure in the SHR. Transcriptomic experiments confirmed differences in the renal expression of several genes (including Ms4a6a, Ndrg3, Egln1, Cd36, Sema3a, Ugt2b, and Idi21) located in some of the CNVs between SHR and Wistar-Kyoto rats. In F(2) animals derived from an SHRxWistar-Kyoto cross, we also found a significant increase in blood pressure associated with an increase in copy number in the Egln1 gene. Our findings suggest that CNVs may play a role in the susceptibility to hypertension and related traits in the SHR.

Figure 1

A) Genomic locations of SHR CNVs. Location of 16 CNVs identified by aCGH experiments relative to the WKY strain. Green arrows and red arrows reflect gains and losses, respectively. B) Detection of CNVs in the SHR using oligonucleotide expression arrays. Log2 ratios are plotted with no moving average as a function of chromosomal position (Mb) for the DNA. Increased Log2 ratio is predictive of DNA duplication while decrease in Log2 ratio is indicative of DNA loss. An increase of Log2 ratio is evident for Chromosome 17 which are zoomed in the above boxes showing the Chromosome 17 region in more detail including gene content.

Figure 2

Representative validation of CNV regions using real-time PCR. Results are the average of triplicate reactions in SHR and WKY. * P<0.05 SHR vs WKY.

Figure 3

Expression of genes in regions of gene duplications or deletions in kidney, heart and spleen. The graph depicts the mean ± SD for data from three replicates for both SHR and WKY. *P<0.05; **P<0.01.

Figure 4

A representative figure of CNVs located on Chromosome 1 in relation to BP QTLs and our congenic strains in the region.

Figure 5

Impact of CNVs in the Egln1 gene (open bars) on systolic BP in the F2 population at 20 weeks of age. The numbers on the bars represent the numbers of animals in the group.