Genetic differentiation of hypothalamus parentally biased transcripts in populations of the house mouse implicate the Prader-Willi syndrome imprinted region as a possible source of behavioral divergence

Abstract

Parentally biased expression of transcripts (genomic imprinting) in adult tissues, including the brain, can influence and possibly drive the evolution of behavioral traits. We have previously found that paternally determined cues are involved in population-specific mate choice decisions between two populations of the Western house mouse (Mus musculus domesticus). Here, we ask whether this could be mediated by genomically imprinted transcripts that are subject to fast differentiation between these populations. We focus on three organs that are of special relevance for mate choice and behavior: The vomeronasal organ (VNO), the hypothalamus, and the liver. To first identify candidate transcripts at a genome-wide scale, we used reciprocal crosses between M. m. domesticus and M. m. musculus inbred strains and RNA sequencing of the respective tissues. Using a false discovery cutoff derived from mock reciprocal cross comparisons, we find a total of 66 imprinted transcripts, 13 of which have previously not been described as imprinted. The largest number of imprinted transcripts were found in the hypothalamus; fewer were found in the VNO, and the least were found in the liver. To assess molecular differentiation and imprinting in the wild-derived M. m. domesticus populations, we sequenced the RNA of the hypothalamus from individuals of these populations. This confirmed the presence of the above identified transcripts also in wild populations and allowed us to search for those that show a high genetic differentiation between these populations. Our results identify the Ube3a-Snrpn imprinted region on chromosome 7 as a region that encompasses the largest number of previously not described transcripts with paternal expression bias, several of which are at the same time highly differentiated. For four of these, we confirmed their imprinting status via single nucleotide polymorphism-specific pyrosequencing assays with RNA from reciprocal crosses. In addition, we find the paternally expressed Peg13 transcript within the Trappc9 gene region on chromosome 15 to be highly differentiated. Interestingly, both regions have been implicated in Prader-Willi nervous system disorder phenotypes in humans. We suggest that these genomically imprinted regions are candidates for influencing the population-specific mate-choice in mice.

Keywords: Mus domesticus; Mus musculus; RNASeq; genetic differentiation; hypothalamus; imprinting; liver; vomeronasal organ.

Fig. 1.

Determination of FDR cutoff for the three tissues. To find a chi-square test cutoff corresponding to an FDR of 5%, reads from the same cross direction were split randomly into two groups and analyzed as if they originated from reciprocal crosses. Then the ratio of SNPs from such mock comparisons to the number of SNPs identified in reciprocal crosses was calculated. Values are plotted as a function of different P value cutoffs. The acceptable level of false positives was set to 5% (dashed lines), and corresponding cutoff values were applied for the data analysis. The relative variability in the LIV data is due to the small number of actually imprinted transcripts.

Fig. 2.

Pyrograms confirming the imprinting status at four loci in six groups of mice. The loci tested were (A) A230073K19Rik, (B/) AK038761, (C) AK139082, and (D) D7Ertd715e. Reciprocal crosses of WSB and PWD inbred strains as well as pure outbred French (MC), pure outbred German (CB) individuals, and reciprocal crosses of these strains were used with three samples per group (female is always listed first in the cross designations). The percent contribution of the one or other allele is listed in each field. Note that not all SNPs are completely diagnostic, but the data are consistent for the respective crosses. In cases where two peaks show the same height but the percent contribution confirms the skew of allelic expression, this is due to the amplicon sequence given for pyro-sequencing. If a SNP is preceded by a nucleotide being the same as one of the two SNP states, three scenarios can occur: 1) equal expression results in the one peak being twice as high as the other with normal height (not present in our data set), 2) unique expression of the allele preceded by the same nucleotide results in a peak twice as high as a normal peak and no peak for the second allele (e.g., individual 7), and 3) if just the second allele is expressed, this results in the first peak from the detected SNP-preceding nucleotide and the second normal height peak resulting from the expressed allele. Amplicon sequences are listed in supplementary table S3, Supplementary Material online.