Results:
First, we evaluated whether there were 2-way interactions between gene and tissue and between gene and country to determine whether subsequent analyses should be carried out separately for each gene. There was a significant interaction between gene and tissue (F4,660= 5.69, p < 0.001). Inclusion of individual ID and country as random effects also revealed significant repeatability of among-individual (r = 0.26, 95% CI = 0.23, 0.28) and among-country (r = 0.42, 95% CI = 0.28, 0.59) variation in gene expression.
Next, we were interested in determining which covariates best explained the among-country variation observed. Our model selection results (Table 2) revealed that the same set of factors were consistent predictors of expression for all three genes: tissue, population type, temperature predictability, genetic group, and latitude; urbanization was a significant predictor for TET2 only. Sex, body mass, altitude, and precipitation predictability were excluded from all models. Moreover, the best-fit models were quite effective at explaining variation in expression of all three genes (conditional R2 range: 0.36 – 0.48; Table 2)
Finally, given that our first analysis revealed significant among individual and among country repeatability, and our model selection revealed that the same set of factors explained variation in gene expression for all three genes, we evaluated correlations in gene expression among-countries, among-individuals, and within-individuals. We found strong support for correlations in gene expression among countries and within individuals, but not among individuals (see Figure 3). Controlling for tissue type, in countries where house sparrows exhibited high average expression of DNMT1, they also had had average expression of DNMT3 (r = 0.15, 95% CI = 0.11, 0.35) and TET2 (r = 0.16, 95% CI = 0.11, 0.36), and similarly high expression of DNMT3 was associated with high expression of TET2 (r = 0.18, 95% CI = 0.11, 0.36). However, when controlling for these among country correlations, there was no correlation between an individual’s average expression of DNMT1 and DNMT3 (r = 0.000, 95% CI = -0.018, 0.059), between DNMT1 and TET2 (r = 0.00, -0.024, 0.031), or between DNMT3 and TET2 (r = 0.00, 95% CI = -0.02, 0.06). There were significant within individual correlations in gene expression. Higher residual expression of DNMT1 was positively correlated with expression of DNMT3 (r = 0.23, 95% CI = 0.20, 0.26) and TET2 (r = 0.16, 95% CI = 0.13, 0.19), and higher expression of DNMT3 was correlated with higher expression of TET2 (r = 0.20, 95% CI = 0.17, 0.23), controlling for tissue type.