Branch point evolution controls species-specific alternative splicing and regulates long term potentiation

Regulation and functionality of species-specific alternative splicing has remained enigmatic for many years. Calcium/calmodulin-dependent protein kinase IIß (CaMKIIß) is expressed in several splice variants and plays a key role in learning and memory. Here, we identify and characterize several primate-specific CAMK2B splice isoforms, which show altered kinetic properties and changes in substrate specificity. Furthermore, we demonstrate that primate-specific Camk2ß alternative splicing is achieved through branch point weakening during evolution. We show that reducing branch point and splice site strength during evolution globally renders constitutive exons alternative, thus providing a paradigm for cis-directed species-specific alternative splicing regulation. Using CRISPR/Cas9 we introduced the weaker human branch point into the mouse genome, resulting in human-like CAMK2B splicing in the brain of mutant mice. We observe a strong impairment of long-term potentiation in CA3-CA1 synapses of mutant mice, thus connecting branch point-controlled, species-specific alternative splicing with a fundamental function in learning and memory. Overall design: A mouse model of humanized Camk2b alternative splicing was generated. Cerebellum tissue was extracted from the wild type strain (C57Bl/6) and heterozygous and homozygous mice of the mouse model. Total RNA was extracted using RNATri (Bio&Sell).