Phylotranscriptomics reveals the convergent evolution of aggression is associated with both shared and unique patterns of gene expression evolution in cavity nesting songbirds

Uncovering the genomic regulation of phenotypic adaptation is a major goal in biology, but this has been hard to achieve for complex behavioral traits. Here, we leverage the repeated evolution of obligate cavity-nesting in birds to test the hypothesis that the shared pressure to compete for a limited breeding resource drives convergent behavioral evolution via convergent gene regulatory changes in the brain. Using behavioral assays in the field, hormonal measures, and transcriptome-wide analyses in 10 focal species, we examined females and males in five avian families, each including one obligate cavity-nesting species and a related species with a more flexible nesting strategy. Results support the hypothesis of behavioral convergence, with high levels of territorial aggression in obligate cavity-nesters, particularly among females. Levels of testosterone were not associated with nesting strategy for either sex, but phylogenetic analyses of individual genes and co-regulated gene networks revealed some shared patterns of gene expression, with functional enrichment related to metabolic and mitochondrial processes. Though this gene regulatory convergence may contribute to behavioral convergence, concordance in gene expression did not scale across multiple families. These observations indicate that replicated evolutionary increases in aggression arise via largely lineage-specific changes in brain gene expression. To evaluate the role of nesting strategy in shaping avian behavior, hormones, and gene expression, we compared 10 species of birds with obligate versus more flexible nesting strategies. We conducted simulated territorial intrusion to measure aggression, conducted enzyme immunoassay to measure testosterone from blood plasma, and used RNA-seq to measure gene expression in the ventromedial telencephalon.