Mammalian splicing divergence is shaped by drift, buffering in trans, and a scaling law

Alternative splicing is ubiquitous, but the mechanisms underlying its pattern of evolutionary divergence across mammalian tissues are still underexplored. Here, we investigated the cis-regulatory divergences and their relationship with tissue-dependent trans-regulation in multiple tissues of an F1 hybrid between two mouse species. Large splicing changes between tissues are highly conserved and likely reflect functional tissue-dependent regulation. In particular, micro-exons frequently exhibit this pattern with high inclusion levels in the brain. Cis-divergence of splicing appears to be largely non-adaptive. Although divergence is in general associated with higher densities of sequence variants in regulatory regions, events with high usage of the dominant isoform apparently tolerate more mutations, explaining why their exon sequences are highly conserved but their intronic splicing site flanking regions are not. Moreover, we demonstrate that non-adaptive mutations are often masked in tissues where accurate splicing likely is more important, and experimentally attribute such buffering effect to trans-regulatory splicing efficiency.