Identification of New Branch Points and Unconventional Introns in Saccharomyces Cerevisiae

Overview

Journal RNA

Specialty Molecular Biology

Date 2016 Jul 31

PMID 27473169

Citations 21

Authors

Genevieve M Gould

Joseph M Paggi

Yuchun Guo

David V Phizicky

Boris Zinshteyn

Eric T Wang

Wendy V Gilbert

David K Gifford

Christopher B Burge

Affiliations

Soon will be listed here.

Abstract

Spliced messages constitute one-fourth of expressed mRNAs in the yeast Saccharomyces cerevisiae, and most mRNAs in metazoans. Splicing requires 5' splice site (5'SS), branch point (BP), and 3' splice site (3'SS) elements, but the role of the BP in splicing control is poorly understood because BP identification remains difficult. We developed a high-throughput method, Branch-seq, to map BPs and 5'SSs of isolated RNA lariats. Applied to S. cerevisiae, Branch-seq detected 76% of expressed, annotated BPs and identified a comparable number of novel BPs. We performed RNA-seq to confirm associated 3'SS locations, identifying some 200 novel splice junctions, including an AT-AC intron. We show that several yeast introns use two or even three different BPs, with effects on 3'SS choice, protein coding potential, or RNA stability, and identify novel introns whose splicing changes during meiosis or in response to stress. Together, these findings show unanticipated complexity of splicing in yeast.

Citing Articles

Mechanisms and regulation of spliceosome-mediated pre-mRNA splicing in Saccharomyces cerevisiae.

Senn K, Hoskins A Wiley Interdiscip Rev RNA. 2024; 15(4):e1866.

PMID: 38972853 PMC: 11585973. DOI: 10.1002/wrna.1866.

Splicing factor Prp18p promotes genome-wide fidelity of consensus 3'-splice sites.

Roy K, Gabunilas J, Neutel D, Ai M, Yeh Z, Samson J Nucleic Acids Res. 2023; 51(22):12428-12442.

PMID: 37956322 PMC: 10711555. DOI: 10.1093/nar/gkad968.

A widespread inversion polymorphism conserved among Saccharomyces species is caused by recurrent homogenization of a sporulation gene family.

Salzberg L, Martos A, Lombardi L, Jermiin L, Blanco A, Byrne K PLoS Genet. 2022; 18(11):e1010525.

PMID: 36441813 PMC: 9731477. DOI: 10.1371/journal.pgen.1010525.

Reciprocal hemizygosity analysis reveals that the Saccharomyces cerevisiae CGI121 gene affects lag time duration in synthetic grape must.

Li R, Deed R G3 (Bethesda). 2021; 11(4).

PMID: 33681985 PMC: 8759811. DOI: 10.1093/g3journal/jkab061.

Saccharomyces cerevisiae RNA lariat debranching enzyme, Dbr1p, is required for completion of reverse transcription by the retrovirus-like element Ty1 and cleaves branched Ty1 RNAs.

Menees T Mol Genet Genomics. 2021; 296(2):409-422.

PMID: 33464395 DOI: 10.1007/s00438-020-01753-y.

References

Dumesic P, Natarajan P, Chen C, Drinnenberg I, Schiller B, Thompson J . Stalled spliceosomes are a signal for RNAi-mediated genome defense. Cell. 2013; 152(5):957-68. PMC: 3645481. DOI: 10.1016/j.cell.2013.01.046. View

Pleiss J, Whitworth G, Bergkessel M, Guthrie C . Rapid, transcript-specific changes in splicing in response to environmental stress. Mol Cell. 2007; 27(6):928-37. PMC: 2081968. DOI: 10.1016/j.molcel.2007.07.018. View

Friedman K, Brewer B . Analysis of replication intermediates by two-dimensional agarose gel electrophoresis. Methods Enzymol. 1995; 262:613-27. DOI: 10.1016/0076-6879(95)62048-6. View

Taggart A, DeSimone A, Shih J, Filloux M, Fairbrother W . Large-scale mapping of branchpoints in human pre-mRNA transcripts in vivo. Nat Struct Mol Biol. 2012; 19(7):719-21. PMC: 3465671. DOI: 10.1038/nsmb.2327. View

Spingola M, Grate L, Haussler D, Ares Jr M . Genome-wide bioinformatic and molecular analysis of introns in Saccharomyces cerevisiae. RNA. 1999; 5(2):221-34. PMC: 1369754. DOI: 10.1017/s1355838299981682. View

Graveley B, Brooks A, Carlson J, Duff M, Landolin J, Yang L . The developmental transcriptome of Drosophila melanogaster. Nature. 2010; 471(7339):473-9. PMC: 3075879. DOI: 10.1038/nature09715. View

Kent W . BLAT--the BLAST-like alignment tool. Genome Res. 2002; 12(4):656-64. PMC: 187518. DOI: 10.1101/gr.229202. View

Katz Y, Wang E, Airoldi E, Burge C . Analysis and design of RNA sequencing experiments for identifying isoform regulation. Nat Methods. 2010; 7(12):1009-15. PMC: 3037023. DOI: 10.1038/nmeth.1528. View

Awan A, Manfredo A, Pleiss J . Lariat sequencing in a unicellular yeast identifies regulated alternative splicing of exons that are evolutionarily conserved with humans. Proc Natl Acad Sci U S A. 2013; 110(31):12762-7. PMC: 3732940. DOI: 10.1073/pnas.1218353110. View

10.

Seraphin B . 3' splice site recognition in S. cerevisiae does not require base pairing with U1 snRNA. Cell. 1993; 73(4):803-12. DOI: 10.1016/0092-8674(93)90258-r. View

11.

Malone R, Bullard S, Hermiston M, Rieger R, Cool M, Galbraith A . Isolation of mutants defective in early steps of meiotic recombination in the yeast Saccharomyces cerevisiae. Genetics. 1991; 128(1):79-88. PMC: 1204456. DOI: 10.1093/genetics/128.1.79. View

12.

Kawashima T, Douglass S, Gabunilas J, Pellegrini M, Chanfreau G . Widespread use of non-productive alternative splice sites in Saccharomyces cerevisiae. PLoS Genet. 2014; 10(4):e1004249. PMC: 3983031. DOI: 10.1371/journal.pgen.1004249. View

13.

Burnette J, Miyamoto-Sato E, Schaub M, Conklin J, Lopez A . Subdivision of large introns in Drosophila by recursive splicing at nonexonic elements. Genetics. 2005; 170(2):661-74. PMC: 1450422. DOI: 10.1534/genetics.104.039701. View

14.

Kim D, Pertea G, Trapnell C, Pimentel H, Kelley R, Salzberg S . TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol. 2013; 14(4):R36. PMC: 4053844. DOI: 10.1186/gb-2013-14-4-r36. View

15.

Meyer M, Plass M, Perez-Valle J, Eyras E, Vilardell J . Deciphering 3'ss selection in the yeast genome reveals an RNA thermosensor that mediates alternative splicing. Mol Cell. 2011; 43(6):1033-9. DOI: 10.1016/j.molcel.2011.07.030. View

16.

Reich C, VanHoy R, PORTER G, Wise J . Mutations at the 3' splice site can be suppressed by compensatory base changes in U1 snRNA in fission yeast. Cell. 1992; 69(7):1159-69. DOI: 10.1016/0092-8674(92)90637-r. View

17.

Smith C, Nadal-Ginard B . Mutually exclusive splicing of alpha-tropomyosin exons enforced by an unusual lariat branch point location: implications for constitutive splicing. Cell. 1989; 56(5):749-58. DOI: 10.1016/0092-8674(89)90678-8. View

18.

Wahl M, Will C, Luhrmann R . The spliceosome: design principles of a dynamic RNP machine. Cell. 2009; 136(4):701-18. DOI: 10.1016/j.cell.2009.02.009. View

19.

Berchowitz L, Gajadhar A, van Werven F, De Rosa A, Samoylova M, Brar G . A developmentally regulated translational control pathway establishes the meiotic chromosome segregation pattern. Genes Dev. 2013; 27(19):2147-63. PMC: 3850098. DOI: 10.1101/gad.224253.113. View

20.

Stepankiw N, Raghavan M, Fogarty E, Grimson A, Pleiss J . Widespread alternative and aberrant splicing revealed by lariat sequencing. Nucleic Acids Res. 2015; 43(17):8488-501. PMC: 4787815. DOI: 10.1093/nar/gkv763. View