Allele‐specific cis‐regulatory methylation of the gene for vasoactive intestinal peptide in white‐throated sparrows

White-throated sparrows (Zonotrichia albicollis) offer a unique
opportunity to connect genotype with behavioral phenotype. In this
species, a rearrangement of the second chromosome is linked with
territorial aggression; birds with a copy of this “supergene”
rearrangement are more aggressive than those without it. The supergene has
captured the gene VIP, which encodes vasoactive intestinal
peptide, a neuromodulator that causes aggression in other songbirds. In
white-throated sparrows, VIP expression is higher in the
anterior hypothalamus of birds with the supergene than those without it,
and expression of VIP in this region predicts the level
of territorial aggression regardless of genotype. Here, we aimed to
identify epigenetic mechanisms that could contribute to differential
expression of VIP both in breeding adults, which exhibit
morph differences in territorial aggression, and in nestlings, before
territorial behavior develops. We extracted and bisulfite-converted DNA
from samples of the hypothalamus in wild-caught adults and nestlings and
used high-throughput sequencing to measure DNA methylation of a region
upstream of the VIP start site. We found that the allele inside
the supergene was less methylated than the alternative allele in both
adults and nestlings. The differential methylation was attributed
primarily to CpG sites that were shared between the alleles, not to
polymorphic sites, which suggests that epigenetic regulation is occurring
independently of the genetic differentiation within the supergene. This
work represents an initial step toward understanding how epigenetic
differentiation inside chromosomal inversions leads to the development of
alternative behavioral phenotypes.