Alternative splicing and gene expression exhibit distinct patterns of transcriptomic differentiation between lake and stream ecotypes in threespine sticklebacks

The threespine stickleback (Gasterosteus aculeatus Linnaeus, 1758) has played an important role in addressing questions on parallel adaptive evolution, including through classic studies of phenotypic divergence between parapatric lake and stream ecotypes. While gene expression differences are documented between these ecotypes, the extent of alternative splicing differentiation between ecotypes remains elusive. We investigate both these regulatory mechanisms in four replicated lake–stream ecotype pairs from the Pacific and North Atlantic using previously published RNA-seq data of spleens and head kidneys. We identified a small set of genes exhibiting splicing divergence between ecotypes compared to widespread expression divergence. Differentially spliced genes were largely distinct from differentially expressed genes and had dissimilar functional compositions. Both expression and splicing exhibited strong tissue specificity and limited global parallelism. In contrast to patterns reported in marine–freshwater systems, differentially spliced and expressed genes were underrepresented in regions of genomic divergence. Finally, we found that inferences of transcriptional divergence were robust to reference genome choice, with high concordance in significantly differentiated genes. Together, our results indicate that gene expression and alternative splicing represent largely independent layers of transcriptomic differentiation between lake-stream stickleback ecotypes, highlighting the importance of integrating both regulatory processes to characterize transcriptional variation among ecotypes.