Specific splicing defects in S. pombe carrying a degron allele of the Survival of Motor Neuron gene

Abstract

Spinal muscular atrophy results from deletions or mutations in the survival of motor neuron (SMN1) gene. The SMN protein has an essential role in the biogenesis of spliceosomal snRNPs, but the link between a defect in this process and specific splicing inhibition of pre-mRNAs has not been established. In this study, we report the construction of a temperature-degron (td) allele of the Schizosaccharomyces pombe SMN protein and show that its depletion at 371C affects splicing and formation of U1, U2, U4 and U5 snRNPs, but not of U6 and U3 ribonucleoproteins. The function of the tdSMN allele in snRNP assembly is already perturbed at 251C, suggesting a deleterious effect of the tag at this temperature. Using a genome-wide approach, we report that introns react unequally to lower levels of snRNPs in tdSMN cells and that increasing the length of the polypyrimidine tract can improve the splicing efficiency of some, but not all, affected introns. Altogether, our results suggest that the defects observed in tdSMN fission yeast cells mimic splicing deficits observed in SMN-deficient metazoan cells.

Genomic mRNA Profiling Reveals Compensatory Mechanisms for the Requirement of the Essential Splicing Factor U2AF

“…In previous studies, no correlation between conventional Py-tract strength and U2AF 59 requirement for the splicing of specific introns was observed (46). However, the 5Ј end of the U2AF 59 -insensitive ulp2_c intron (54) contains an unusually long (30-nucleotide) uridine tract, which may facilitate splicing by involving either the putative residual activity of the prp2.1 mutant protein or the activity of factors such as p54, TIA-1-like, SMN, or Nam-8p-like protein (9,15,16,29,44). For the bpb1_a intron, which lacks a Py-tract within the 5Ј-most 30 nucleotides, different features ( Fig.…”

The many faces of SMN: deciphering the function critical to spinal muscular atrophy pathogenesis

“…Although protozoa, yeast and even models such as Drosophila, can be advantageous in studying biological processes such as snRNP assembly [44,59,60], results obtained with such model organisms should be interpreted with caution in regards to SMA pathogenesis. In these models, snRNP assembly may function by different mechanisms owing to ortholog divergence of proteins, which can be quite significant in certain models [44].…”

“…A caveat of this method is that it cannot detect intron-specific splicing defects [60]. This has prompted others to study splicing in SMA using a different approach to overcome this limitation.…”

“…This has prompted others to study splicing in SMA using a different approach to overcome this limitation. Using fission yeast cells expressing a temperature-sensitive degron of the SMN gene, Campion et al observed perturbed snRNP assembly and altered intron splicing at the degron-inducing temperature [60]. Of note, the authors also observed snRNP assembly defects at a temperature lower than that required to activate the temperature degron, suggesting that the fusion protein itself had deleterious effects [60].…”

“…Using fission yeast cells expressing a temperature-sensitive degron of the SMN gene, Campion et al observed perturbed snRNP assembly and altered intron splicing at the degron-inducing temperature [60]. Of note, the authors also observed snRNP assembly defects at a temperature lower than that required to activate the temperature degron, suggesting that the fusion protein itself had deleterious effects [60]. Furthermore, it remains to be seen if the changes in intron splicing occur prephenotype and if these splicing alterations are part of the primary mechanism affected in SMA pathogenesis.…”