Connecting Anxiety and Genomic Copy Number Variation: A Genome-Wide Analysis in CD-1 Mice.

Genomic copy number variants (CNVs) have been implicated in multiple psychiatric disorders, but not much is known about their influence on anxiety disorders specifically. Using next-generation sequencing (NGS) and two additional array-based genotyping approaches, we detected CNVs in a mouse model consisting of two inbred mouse lines showing high (HAB) and low (LAB) anxiety-related behavior, respectively. An influence of CNVs on gene expression in the central (CeA) and basolateral (BLA) amygdala, paraventricular nucleus (PVN), and cingulate cortex (Cg) was shown by a two-proportion Z-test (p = 1.6 x 10-31), with a positive correlation in the CeA (p = 0.0062), PVN (p = 0.0046) and Cg (p = 0.0114), indicating a contribution of CNVs to the genetic predisposition to trait anxiety in the specific context of HAB/LAB mice. In order to confirm anxiety-relevant CNVs and corresponding genes in a second mouse model, we further examined CD-1 outbred mice. We revealed the distribution of CNVs by genotyping 64 CD 1 individuals using a high-density genotyping array (Jackson Laboratory). 78 genes within those CNVs were identified to show nominally significant association (48 genes), or a statistical trend in their association (30 genes) with the time animals spent on the open arms of the elevated plus-maze (EPM). Fifteen of them were considered promising candidate genes of anxiety-related behavior as we could show a significant overlap (permutation test, p = 0.0051) with genes within HAB/LAB CNVs. Thus, here we provide what is to our knowledge the first extensive catalogue of CNVs in CD-1 mice and potential corresponding candidate genes linked to anxiety-related behavior in mice

Protein coding genes in genomic regions of CNVs detected in HAB/LAB and CD-1 mice.

<p>All genes listed overlap both CNVs in HAB/LAB mice detected with aCGH, JaxMDGA and NGS, and CNVs in CD-1 mice which were best associated with the time the animals spent on the open arm of the EPM (nominal <i>p</i>-value < 0.1).</p><p>Protein coding genes in genomic regions of CNVs detected in HAB/LAB and CD-1 mice.</p

Distribution of CNVs in CD-1 mice.

<p>Chromosomes are indicated by grey horizontal lines. Start points of CNVs are marked by dots and lines are drawn to the end points. Due to limitations in resolution, a small CNV might appear as dot only. CNVs highlighted in blue or red were associated with anxiety-related behavior (time on the open arm of the EPM) with a nominal <i>p</i>-value less than 0.1 or 0.05, respectively.</p

Genomic positions of CNVs on chromosome 3.

<p>The chromosome is indicated by a thick horizontal line (grey). Depending on the detection method, CNVs in HAB/LAB mice are depicted in orange (aCGH), dark red (JaxMDGA) and red (NGS), respectively. Data displayed above the grey line represent a copy number gain in HAB vs. LAB animals, data below a copy number loss. Data printed on the grey line show CNVs in 64 CD-1 mice, with those highlighted in color that could be associated with anxiety-related behavior (time on the open arm of EPM) with a nominal <i>p</i>-value less than 0.1 (light blue) or less than 0.05 (blue). Start points of CNVs are marked by dots and lines are drawn to the end points.</p

Association of copy number with anxiety-related behavior in CD-1 mice.

<p>Exemplarily, data of three associations resulting in nominal <i>p</i>-values reaching significance (<i>p</i> < 0.05), a trend (<i>p</i> < 0.1), and not reaching significance (<i>p</i> > 0.05), respectively, are shown. Each dot represents data of a single animal (N = 64). The relative copy number is represented by the mean normalized intensities of JaxMDGA probes within the respective CNV. <b>(A)</b> CNV no. 498; <i>P</i>_nom = 0.0009; regression line: y = 0.0091x + 9.4389. <b>(B)</b> CNV no. 164; <i>P</i>_nom = 0.0554; regression line: y = 0.0061x + 10.201. <b>(C)</b> CNV no. 453; <i>P</i>_nom = 0.9791; regression line: y = 0.0008x + 9.6225.</p