A B2 SINE insertion in the Comt1 gene (Comt1(B2i)) results in an overexpressing, behavior modifying allele present in classical inbred mouse strains

Abstract

Catechol-O-methyltransferase (COMT) is a key enzyme for dopamine catabolism and COMT is a candidate gene for human psychiatric disorders. In mouse it is located on chromosome 16 in a large genomic region of extremely low variation among the classical inbred strains, with no confirmed single nucleotide polymorphisms (SNPs) between strains C57BL/6J and DBA/2J within a 600-kB window. We found a B2 SINE in the 3' untranslated region (UTR) of Comt1 which is present in C57BL/6J (Comt1(B2i)) and other strains including 129 (multiple sublines), but is not found in DBA/2J (Comt1(+)) and many other strains including wild-derived Mus domesticus, M. musculus, M. molossinus, M.castaneus and M. spretus. Comt1(B2i) is absent in strains closely related to C57BL/6, such as C57L and C57BR, indicating that it was polymorphic in the cross that gave rise to these strains. The strain distribution of Comt1(B2i) indicates a likely origin of the allele in the parental Lathrop stock. A stringent association test, using 670 highly outbred mice (Boulder Heterogeneous Stock), indicates that this insertion allele may be responsible for a difference in behavior related to exploration. Gene expression differences at the mRNA and enzyme activity level (1.7-fold relative to wild type) indicate a mechanism for this behavioral effect. Taken together, these findings show that Comt1(B2i) (a B2 SINE insertion) results in a relatively modest difference in Comt1 expression and enzyme activity (comparable to the human Val-Met polymorphism) which has a demonstrable behavioral phenotype across a variety of outbred genetic backgrounds.

Figure 1

Annotated sequence for the C57BL/6J strain Comt1 gene. Sequence is a composite of RefSeq cDNA sequence NM_001111062 and sequence of the B2 insertion found in this study. Sequencing of additional strains shows identical sequence in those with the insertion, except for some differences in the length of the poly-A run at the end of the B2 element. Primers used for the sequencing and genotyping are displayed in color, as is the position of the B2 SINE insertion in the Comt1^B2i allele.

Figure 2

Strain distribution. Presence of Comt1^B2i across inbred strains. Boxes group related strains, asterisks denote HS stock progenitors (or in the case of Is/CamEi and RIIIS/J, the probable nearest surviving relative).

Figure 3

Exon array Comt1 gene expression data. Genotype group means of standardized array signal intensities are plotted by genomic position. Comt1^+/+ expression means are plotted in dark blue, Comt1^+/B2i in teal and Comt1^B2i/B2i in green. The structure of the gene is shown at the bottom, in genomic orientation (the 3′ end is at the left). Probe set positions are marked in light blue. The position of the B2 SINE insertion is shown in yellow. Probe sets flank, but do not span, the insertion site. Data presented are based on NCBI36/mm8.

Figure 4

Comt enzyme activity. Means ± standard error of the mean for specific Comt enzyme activity. Enzyme activity is around 1.7-fold higher in C57BL/6J (Comt1^B2i) compared to DBA/2J (Comt1⁺) (t = 3.43, df = 14, P = 8 × 10⁻³).

Figure 5

Behavior. Means ± standard error of the mean for measures from the novel object task (NO) and the open field task (OF) in the HS mice grouped by Comt1^B2i genotype. Duration and frequency of exploration in the novel object are greater in Comt1⁺ than the other genotype classes, and least in the Comt1⁺/Comt1^B2i genotype (Duration: F_2,572 = 8.7, P = 1 × 10⁻⁴; Frequency: F_2,572 = 6.1, P = 2 × 10⁻³). No significant differences are seen in the OF.