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A complex gene regulatory mechanism that operates at the nexus of multiple RNA processing decisions

Abstract

Expression of crs1 pre-mRNA, encoding a meiotic cyclin, is blocked in actively growing fission yeast cells by a multifaceted mechanism. The most striking feature is that in vegetative cells, crs1 transcripts are continuously synthesized but are targeted for degradation rather than splicing and polyadenylation. Turnover of crs1 RNA requires the exosome, as do previously described nuclear surveillance and silencing mechanisms, but does not involve a noncanonical poly(A) polymerase. Instead, crs1 transcripts are targeted for destruction by a factor previously implicated in turnover of meiotic RNAs in growing cells. Like exosome mutants, mmi1 mutants splice and polyadenylate vegetative crs1 transcripts. Two regulatory elements are located at the 3′ end of the crs1 gene, consistent with the increased accumulation of spliced RNA in polyadenylation factor mutants. This highly integrated regulatory strategy may ensure a rapid response to adverse conditions, thereby guaranteeing survival.

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Figure 1: Transcription of the crs1 gene does not determine accumulation of the RNA.
Figure 2: Polyadenylation of crs1 is activated concurrently with splicing during meiosis.
Figure 3: The mechanism that blocks crs1 processing in vegetative cells requires the nuclear exosome in conjunction with a factor implicated in turnover of meiotic transcripts in proliferating cells.
Figure 4: The pfs2-11 mutation leads to splicing and increased accumulation of crs1 RNA in proliferating cells.
Figure 5: The proximal (noncanonical) polyadenylation signal regulates splicing and accumulation of crs1 RNA.
Figure 6: The proximal (noncanonical) polyadenylation signal also regulates 3′ processing of crs1 RNA.
Figure 7: An element within the fifth exon coordinately inhibits crs1 splicing and polyadenylation.
Figure 8: Effect of reversing the order of the polyadenylation signals on crs1 RNA processing.
Figure 9: Model to explain the different processing fates of the crs1 RNA during vegetative growth and meiotic differentiation of wild-type cells and in the mmi1-ts mutants.

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Acknowledgements

The authors would like to thank M. Yamamoto (University of Tokyo), C. Norbury (Oxford University) and R. Maraia (US National Institutes of Health (NIH)) for strains (Supplementary Table 3). We are grateful to our colleagues K. Baker, J. Gott, H. Lou, T. Nilsen, C. Patterson, H. Salz and S. Sanders for helpful discussions and critical reading of the manuscript. This work was funded by NIH grants R01-GM073217, awarded jointly to J.A.W. and J.L., R01-GM064682, awarded to D.S.M., and R01-GM38070, awarded to J.A.W.

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Contributions

D.S.M. performed the TRO experiments, designed the real-time PCR assay and participated in development of the model as well as writing of the manuscript; N.C. constructed and analyzed the chimeric and mutant alleles of crs1 to define the regulatory element, performed the RNA analyses on trans-acting factor mutants and qPCR, and expertly proofread the manuscript; S.S. analyzed processing of crs1 RNA over a meiotic time course and conducted initial experiments to map the crs1 regulatory element; N.A. mapped the crs1 RNA termini by RACE and constructed the crs1 deletion strain. H.-M.C. and J.L. discovered the splicing defect in the pfs2-11 mutant; J.A.W. wrote the manuscript and contributed to the design and interpretation of all experiments.

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Correspondence to Jo Ann Wise.

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McPheeters, D., Cremona, N., Sunder, S. et al. A complex gene regulatory mechanism that operates at the nexus of multiple RNA processing decisions. Nat Struct Mol Biol 16, 255–264 (2009). https://doi.org/10.1038/nsmb.1556

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