The Wise lab studies the mechanism and regulation of pre-mRNA splicing in S.pombe.
Pre-messenger RNA splicing is the process whereby non-coding segments (introns) are removed from primary transcripts within the nucleus, with concomitant joining of exons for export to the cytoplasm. Because splicing defects lie at the root of a significant fraction of human genetic disorders, it is critically important to understand precisely how this process contributes to the proper read-out of a cell’s genomic sequence. Such insights can be difficult to gain in higher eukaryotes due to their complexity and the paucity of in vivo approaches. Thus, our laboratory is exploiting the facile genetic strategies available in the fission yeast Schizosaccharomyces pombe, a unicellular eukaryote in which both cis-acting splicing signals and trans-acting splicing factors closely resemble their human counterparts, to investigate the mechanism and regulation of pre-mRNA splicing.
A key tool at our disposal, which has become even more powerful with the advent of a complete genome sequence for S. pombe, is "reverse" genetics. We are using this approach to elucidate the roles of three factors that function in the earliest recognition of intron/exon boundaries: the U1 snRNP, an RNA-protein complex that binds to the 5’ splice site; U2AF, a heterodimeric protein that contacts the 3’ splice site, and Srp2, a serine/arginine-rich (SR) protein that interacts with exonic enhancer elements. Our goal is to understand how these factors function, both individually and collaboratively, to promote splicing of diverse pre-mRNAs. Current work emphasizes genome-wide strategies such as microarray analysis to understand the interface between splicing and biology.
In a more recently established project, we are investigating the role of regulated splicing in the control of meiotic differentiation in collaboration with Dr. Janet Leatherwood’s laboratory (SUNY Stony Brook). We have discovered an extensive network of genes for which regulated splicing is used to turn on protein production during meiosis. More detailed investigation of two pre-mRNAs that encode cyclins reveals that splicing is restricted to meiosis by a novel inhibitory mechanism that requires non-intronic elements located outside the coding regions, a considerable distance from the target introns. This shared and highly unusual feature suggests communication between the splicing machinery and macromolecular complexes that mediate the preceding and subsequent steps in gene expression (see Figure). Thus, this research provides an unprecedented opportunity to explore the function of the mRNA production factory in a native biological context.
McPheeters, D.S., Cremona, N., Sunder, S., Averbeck, N., Chen, H.-M., Leatherwood, J. and Wise, J.A. 2009. A Complex Gene Regulatory Mechanism that Operates at the Nexus of Multiple RNA Processing Decisions. Nature Structural and Molecular Biology 16, 255-264.
Averbeck, N. A., Sunder, S., Sample, N., Wise, J. A. and Leatherwood, J. K. (2005) Negative Control Contributes to an Extensive Program of Meiotic Splicing in Fission Yeast. Mol. Cell 18, 491-498.
Webb, C. J., Romfo, C. M., van Heeckeren, W. J., and Wise, J. A. (2005) Exonic Splicing Enhancers in Fission Yeast: Functional Conservation Demonstrates an Early Evolutionary Origin. Genes Dev. 19, 242-254.
Webb, C. J. Romfo, C. M., Reddy, S.-L. and Wise, J. A. (2005) Analysis of Mutant Phenotypes and Splicing Defects Demonstrates Functional Collaboration between the Large and Small Subunits of the Essential Splicing Factor U2AF. Mol. Biol. Cell 16, 584-596.
Webb, C. J. and Wise, J. A. (2004) The Splicing Factor U2AF Small Subunit is Functionally Conserved between Fission Yeast and Humans. Mol. Cell. Biol. 21, 7955-7970.
Alvarez. C. J. and Wise, J. A. (2001) Activation of a cryptic 5' splice site by U1 snRNA. RNA 7, 342-350.