Real-time Imaging of Cotranscriptional Splicing Reveals a Kinetic Model That Reduces Noise: Implications for Alternative Splicing Regulation

Overview

Journal J Cell Biol

Specialty Cell Biology

Date 2011 Jun 1

PMID 21624952

Citations 64

Authors

Ute Schmidt

Eugenia Basyuk

Marie-Cecile Robert

Minoru Yoshida

Jean-Philippe Villemin

Didier Auboeuf

Stuart Aitken

Edouard Bertrand

Affiliations

Soon will be listed here.

Abstract

Splicing is a key process that expands the coding capacity of genomes. Its kinetics remain poorly characterized, and the distribution of splicing time caused by the stochasticity of single splicing events is expected to affect regulation efficiency. We conducted a small-scale survey on 40 introns in human cells and observed that most were spliced cotranscriptionally. Consequently, we constructed a reporter system that splices cotranscriptionally and can be monitored in live cells and in real time through the use of MS2-GFP. All small nuclear ribonucleoproteins (snRNPs) are loaded on nascent pre-mRNAs, and spliceostatin A inhibits splicing but not snRNP recruitment. Intron removal occurs in minutes and is best described by a model where several successive steps are rate limiting. Each pre-mRNA molecule is predicted to require a similar time to splice, reducing kinetic noise and improving the regulation of alternative splicing. This model is relevant to other kinetically controlled processes acting on few molecules.

Citing Articles

Live Single-Cell Transcriptional Dynamics in Plant Cells.

Hani S, Mercier C, David P, Bertrand E, Desnos T, Nussaume L Methods Mol Biol. 2024; 2875:37-58.

PMID: 39535638 DOI: 10.1007/978-1-0716-4248-1_4.

Spatial organization of translation and translational repression in two phases of germ granules.

Ramat A, Haidar A, Garret C, Simonelig M Nat Commun. 2024; 15(1):8020.

PMID: 39271704 PMC: 11399267. DOI: 10.1038/s41467-024-52346-x.

Co-transcriptional splicing facilitates transcription of gigantic genes.

Fingerhut J, Lannes R, Whitfield T, Thiru P, Yamashita Y PLoS Genet. 2024; 20(6):e1011241.

PMID: 38870220 PMC: 11207136. DOI: 10.1371/journal.pgen.1011241.

Co-transcriptional gene regulation in eukaryotes and prokaryotes.

Shine M, Gordon J, Scharfen L, Zigackova D, Herzel L, Neugebauer K Nat Rev Mol Cell Biol. 2024; 25(7):534-554.

PMID: 38509203 PMC: 11199108. DOI: 10.1038/s41580-024-00706-2.

Shell protein composition specified by the lncRNA NEAT1 domains dictates the formation of paraspeckles as distinct membraneless organelles.

Takakuwa H, Yamazaki T, Souquere S, Adachi S, Yoshino H, Fujiwara N Nat Cell Biol. 2023; 25(11):1664-1675.

PMID: 37932453 DOI: 10.1038/s41556-023-01254-1.

References

Pandya-Jones A, Black D . Co-transcriptional splicing of constitutive and alternative exons. RNA. 2009; 15(10):1896-908. PMC: 2743041. DOI: 10.1261/rna.1714509. View

Sultan M, Schulz M, Richard H, Magen A, Klingenhoff A, Scherf M . A global view of gene activity and alternative splicing by deep sequencing of the human transcriptome. Science. 2008; 321(5891):956-60. DOI: 10.1126/science.1160342. View

Wada Y, Ohta Y, Xu M, Tsutsumi S, Minami T, Inoue K . A wave of nascent transcription on activated human genes. Proc Natl Acad Sci U S A. 2009; 106(43):18357-61. PMC: 2761237. DOI: 10.1073/pnas.0902573106. View

Roybal G, Jurica M . Spliceostatin A inhibits spliceosome assembly subsequent to prespliceosome formation. Nucleic Acids Res. 2010; 38(19):6664-72. PMC: 2965229. DOI: 10.1093/nar/gkq494. View

Kiseleva E, Wurtz T, Visa N, Daneholt B . Assembly and disassembly of spliceosomes along a specific pre-messenger RNP fiber. EMBO J. 1994; 13(24):6052-61. PMC: 395583. DOI: 10.1002/j.1460-2075.1994.tb06952.x. View

Wang E, Sandberg R, Luo S, Khrebtukova I, Zhang L, Mayr C . Alternative isoform regulation in human tissue transcriptomes. Nature. 2008; 456(7221):470-6. PMC: 2593745. DOI: 10.1038/nature07509. View

Zhang G, TANEJA K, Singer R, Green M . Localization of pre-mRNA splicing in mammalian nuclei. Nature. 1994; 372(6508):809-12. DOI: 10.1038/372809a0. View

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

Huranova M, Ivani I, Benda A, Poser I, Brody Y, Hof M . The differential interaction of snRNPs with pre-mRNA reveals splicing kinetics in living cells. J Cell Biol. 2010; 191(1):75-86. PMC: 2953428. DOI: 10.1083/jcb.201004030. View

10.

McCracken S, Fong N, Yankulov K, Ballantyne S, Pan G, GREENBLATT J . The C-terminal domain of RNA polymerase II couples mRNA processing to transcription. Nature. 1997; 385(6614):357-61. DOI: 10.1038/385357a0. View

11.

Mata M, Alonso C, Kadener S, Fededa J, Blaustein M, Pelisch F . A slow RNA polymerase II affects alternative splicing in vivo. Mol Cell. 2003; 12(2):525-32. DOI: 10.1016/j.molcel.2003.08.001. View

12.

Tardiff D, Lacadie S, Rosbash M . A genome-wide analysis indicates that yeast pre-mRNA splicing is predominantly posttranscriptional. Mol Cell. 2006; 24(6):917-29. PMC: 1828117. DOI: 10.1016/j.molcel.2006.12.002. View

13.

Oesterreich F, Preibisch S, Neugebauer K . Global analysis of nascent RNA reveals transcriptional pausing in terminal exons. Mol Cell. 2010; 40(4):571-81. DOI: 10.1016/j.molcel.2010.11.004. View

14.

Fusco D, Accornero N, Lavoie B, Shenoy S, Blanchard J, Singer R . Single mRNA molecules demonstrate probabilistic movement in living mammalian cells. Curr Biol. 2003; 13(2):161-167. PMC: 4764064. DOI: 10.1016/s0960-9822(02)01436-7. View

15.

Blencowe B . Alternative splicing: new insights from global analyses. Cell. 2006; 126(1):37-47. DOI: 10.1016/j.cell.2006.06.023. View

16.

Wetterberg I, Zhao J, Masich S, Wieslander L, Skoglund U . In situ transcription and splicing in the Balbiani ring 3 gene. EMBO J. 2001; 20(10):2564-74. PMC: 125468. DOI: 10.1093/emboj/20.10.2564. View

17.

Molle D, Segura-Morales C, Camus G, Berlioz-Torrent C, Kjems J, Basyuk E . Endosomal trafficking of HIV-1 gag and genomic RNAs regulates viral egress. J Biol Chem. 2009; 284(29):19727-43. PMC: 2740597. DOI: 10.1074/jbc.M109.019844. View

18.

Pandit S, Wang D, Fu X . Functional integration of transcriptional and RNA processing machineries. Curr Opin Cell Biol. 2008; 20(3):260-5. PMC: 2701685. DOI: 10.1016/j.ceb.2008.03.001. View

19.

Singh J, Padgett R . Rates of in situ transcription and splicing in large human genes. Nat Struct Mol Biol. 2009; 16(11):1128-33. PMC: 2783620. DOI: 10.1038/nsmb.1666. View

20.

Zillmann M, Zapp M, Berget S . Gel electrophoretic isolation of splicing complexes containing U1 small nuclear ribonucleoprotein particles. Mol Cell Biol. 1988; 8(2):814-21. PMC: 363208. DOI: 10.1128/mcb.8.2.814-821.1988. View