Characterization and In Vivo Functional Analysis of the Schizosaccharomyces pombe ICLN Gene

Abstract

During the early steps of snRNP biogenesis, the survival motor neuron (SMN) complex acts together with the methylosome, an entity formed by the pICln protein, WD45, and the PRMT5 methyltransferase. To expand our understanding of the functional relationship between pICln and SMN in vivo, we performed a genetic analysis of an uncharacterized Schizosaccharomyces pombe pICln homolog. Although not essential, the S. pombe ICln (SpICln) protein is important for optimal yeast cell growth. The human ICLN gene complements the Δicln slow-growth phenotype, demonstrating that the identified SpICln sequence is the bona fide human homolog. Consistent with the role of human pICln inferred from in vitro experiments, we found that the SpICln protein is required for optimal production of the spliceosomal snRNPs and for efficient splicing in vivo. Genetic interaction approaches further demonstrate that modulation of ICln activity is unable to compensate for growth defects of SMN-deficient cells. Using a genome-wide approach and reverse transcription (RT)-PCR validation tests, we also show that splicing is differentially altered in Δicln cells. Our data are consistent with the notion that splice site selection and spliceosome kinetics are highly dependent on the concentration of core spliceosomal components.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.01407-13.

ACKNOWLEDGMENTS

We thank Florence Rage and Marta Radman-Livaja for discussions and critical reading of the manuscript. We gratefully acknowledge members of the fission yeast community for sharing strains and plasmids. We also thank T. Nakamura (YGRC, Osaka, Japan) for the pTN-RC5 library. Affymetrix microarrays were processed in the Microarray Core Facility of the Institute of Research on Biotherapy, CHRU-INSERM-UM1 Montpellier, France.

This work was supported by grants from SMA Europe to R.B. and the CNRS.