Factors and mechanism of intron-specific splicing of pre- mRNAs with branchpoint-distant exons

Abstract

Gene expression is a highly regulated process and a key regulation happens at the level of RNA splicing. Coding region of pre-mRNAs are interspaced by non-coding sequences called introns. They are marked by splicing signals such as 5’splice site (5’ss), branch point (BP) and 3’ splice site (3’ss), which are recognized by spliceosome formed after sequential rearrangements. The removal of introns is essential for the formation of mature transcripts and requires spliceosome. Spliceosome is comprised of multiple snRNPs like U1, U2, U4, U5 and U6 and hundreds of proteins which helps in the removal of introns from pre-mRNAs (Wilkinson et al., 2020). RNA component of different snRNPs contribute significantly to the recognition of introns and in the catalysis of exon splicing reaction. Efficiency of the splicing depends on the optimal recognition of these splice signals (Will & Lührmann, 2011). Longer spacing between BP and 3’ss makes the recognition of 3’ss inefficient, since this excess RNA needs to be accommodated in the multi-megadalton RNA-protein complex, spliceosome, for the proper docking of 3’ss into the catalytic centre. Optimum distance between BP and 3’ss in introns in humans is 19 to 23 nt, suggesting any deviations from this distance might make the splicing inefficient. Recognitions of BP-distant 3’ss introns might require additional factors which can be modulated by binding of different splicing regulators like RNA-binding proteins (RBP), protein-modifying enzymes and ubiquitin-like proteins (UBL). These regulators can bind at cis-regulatory elements or to different complex of spliceosome and regulate splicing. UBLs are dispensable for constitutive pre-mRNA splicing in the budding yeast. Hub1, an unconventional UBL, was shown to modify spliceosome and is required for the alternative splicing of Src1/HEH1 (Chanarat & Mishra, 2018; Mishra et al., 2011). Another such example of UBL is the ubiquitin-fold-activated splicing regulator Sde2 that is conserved from Schizosaccharomyces pombe to humans. Cells lacking Sde2 show temperature sensitivity and defective telomeric silencing in S. pombe. The Sde2 precursor has a N-terminal UBL domain which gets processed by the deubiquitinating enzymes (DUB), Ubp5 and Ubp15. This processing generates Sde2-C which is incorporated into the spliceosome for efficient splicing of a subset of introns (Thakran et al., 2018).