Developmentally-regulated splice remodeling shapes functional CACNA1C channel diversity during human neurodevelopment

Abstract

CACNA1C encodes the pore-forming subunit of the CaV1.2 L-type voltage-gated calcium channel, a major regulator of neuronal calcium signaling and one of the strongest genetic risk loci for bipolar disorder and other psychiatric disorders. Although extensive alternative splicing of CACNA1C has been described in the adult human brain, how alternative splicing of CACNA1C is regulated during early human neurodevelopment remains largely unknown. Here, we used long-read RNA sequencing to characterize full-length CACNA1C transcripts during differentiation of human induced pluripotent stem cells, from healthy controls and patients with bipolar disorder homozygous for either the A or G allele at rs1006737, into cortical glutamatergic neurons. We identified 60 putative isoforms, including ten annotated transcripts and 23 previously reported in adult human brain. We also demonstrate that CACNA1C undergoes developmentally-regulated remodeling of isoform composition during neuronal differentiation. This remodeling was characterized by an exon-switching program involving mutually exclusive exon pairs and alternative splice site usage, including prominent developmental transitions between exon pairs 8/8a, 21/22, and 31/32. Neither rs1006737 genotype, diagnosis, sex nor age significantly affected splicing profiles. The developmental splice trajectories were recapitulated in dorsal forebrain organoids, showing that CACNA1C splice remodeling is conserved across independent human neural differentiation models. Electrophysiological characterization of the two most abundant neuronal isoforms found that exon 31/32 switching alters channel inactivation properties. Together, our findings demonstrate that developmentally-regulated alternative splicing of CACNA1C transcripts contributes to the functional diversity of CaV1.2 channels expressed during human neurodevelopment.