XAP5 CIRCADIAN TIMEKEEPER regulates RNA splicing and the circadian clock by genetically separable pathways

Regulators of post-transcriptional events, such as pre-mRNA splicing factors, are involved in controlling the pace of the circadian clock. However, in most cases the underlying mechanisms remain unclear. Previously, we have identified XAP5 CIRCADIAN TIMEKEEPER (XCT) as an Arabidopsis thaliana circadian clock regulator with uncharacterized molecular functions. To investigate a possible role for XCT in RNA splicing, we performed PacBio Isoform Sequencing (Iso-Seq) on wild-type Col-0, reduction-of-function allele xct-1, null allele xct-2, the complemented line xct-2 XCT and prl1-2 (a mutate with known defects in intron retention). We then performed differential splicing analysis on the Iso-Seq data using JunctionSeq R package. Our results indicate that xct mutants have transcriptome-wide pre-mRNA splicing defects, predominantly aberrant 3’ splice site selection. Expression of a genomic copy of XCT fully rescues those splicing defects. Differential splicing analysis of PacBio Iso-Seq data for XCT- and othe splicing-factor-related alleles.