Mutational Analysis of the Yeast RNA Helicase Sub2p Reveals Conserved Domains Required for Growth, MRNA Export, and Genomic Stability

Overview

Journal RNA

Specialty Molecular Biology

Date 2013 Aug 22

PMID 23962665

Citations 12

Authors

Cyril Saguez

Fernando A Gonzales

Manfred Schmid

Andreas Boggild

Chrysa M Latrick

Francisco Malagon

Andrea Putnam

Lee Sanderson

Eckhard Jankowsky

Ditlev E Brodersen

Torben Heick Jensen

Affiliations

Soon will be listed here.

Abstract

Sub2p/UAP56 is a highly conserved DEAD-box RNA helicase involved in the packaging and nuclear export of mRNA/protein particles (mRNPs). In Saccharomyces cerevisiae, Sub2p is recruited to active chromatin by the pentameric THO complex and incorporated into the larger transcription-export (TREX) complex. Sub2p also plays a role in the maintenance of genome integrity as its inactivation causes severe transcription-dependent recombination of DNA. Despite the central role of Sub2p in early mRNP biology, little is known about its function. Here, we report the presence of an N-terminal motif (NTM) conserved specifically in the Sub2p branch of RNA helicases. Mutation of the NTM causes nuclear accumulation of poly(A)(+) RNA and impaired growth without affecting core helicase functions. Thus, the NTM functions as an autonomous unit. Moreover, two sub2 mutants, that are deficient in ATP binding, act in a trans-dominant negative fashion for growth and induce high recombination rates in vivo. Although wild-type Sub2p is prevented access to transcribed loci in such a background, this does not mechanistically explain the phenotype.

Citing Articles

Structures and mRNP remodeling mechanism of the TREX-2 complex.

Xie Y, Clarke B, Xie D, Mei M, Bhat P, Hill P Structure. 2025; 33(3):566-582.e6.

PMID: 39862860 PMC: 11890942. DOI: 10.1016/j.str.2024.12.019.

RNA biogenesis and RNA metabolism factors as R-loop suppressors: a hidden role in genome integrity.

Luna R, Gomez-Gonzalez B, Aguilera A Genes Dev. 2024; 38(11-12):504-527.

PMID: 38986581 PMC: 11293400. DOI: 10.1101/gad.351853.124.

The Hog1 MAPK substrate governs Candida glabrata-epithelial cell adhesion via the histone H2A variant.

Sahu M, Purushotham R, Kaur R PLoS Genet. 2024; 20(5):e1011281.

PMID: 38743788 PMC: 11125552. DOI: 10.1371/journal.pgen.1011281.

Critical Cellular Functions and Mechanisms of Action of the RNA Helicase UAP56.

Yellamaty R, Sharma S J Mol Biol. 2024; 436(12):168604.

PMID: 38729260 PMC: 11168752. DOI: 10.1016/j.jmb.2024.168604.

Structural differences between the closely related RNA helicases, UAP56 and URH49, fashion distinct functional apo-complexes.

Fujita K, Ito M, Irie M, Harada K, Fujiwara N, Ikeda Y Nat Commun. 2024; 15(1):455.

PMID: 38225262 PMC: 10789772. DOI: 10.1038/s41467-023-44217-8.

References

Abruzzi K, Lacadie S, Rosbash M . Biochemical analysis of TREX complex recruitment to intronless and intron-containing yeast genes. EMBO J. 2004; 23(13):2620-31. PMC: 449771. DOI: 10.1038/sj.emboj.7600261. View

Jensen T, Patricio K, McCarthy T, Rosbash M . A block to mRNA nuclear export in S. cerevisiae leads to hyperadenylation of transcripts that accumulate at the site of transcription. Mol Cell. 2001; 7(4):887-98. DOI: 10.1016/s1097-2765(01)00232-5. View

Schmid M, Rougemaille M, Libri D, Jensen T . Chapter 10. Estimating nuclear mRNA decay in Saccharomyces cerevisiae. Methods Enzymol. 2009; 449:205-19. DOI: 10.1016/S0076-6879(08)02410-5. View

Fan H, Merker R, Klein H . High-copy-number expression of Sub2p, a member of the RNA helicase superfamily, suppresses hpr1-mediated genomic instability. Mol Cell Biol. 2001; 21(16):5459-70. PMC: 87268. DOI: 10.1128/MCB.21.16.5459-5470.2001. View

Yang Q, Jankowsky E . ATP- and ADP-dependent modulation of RNA unwinding and strand annealing activities by the DEAD-box protein DED1. Biochemistry. 2005; 44(41):13591-601. DOI: 10.1021/bi0508946. View

Albuquerque C, Smolka M, Payne S, Bafna V, Eng J, Zhou H . A multidimensional chromatography technology for in-depth phosphoproteome analysis. Mol Cell Proteomics. 2008; 7(7):1389-96. PMC: 2493382. DOI: 10.1074/mcp.M700468-MCP200. View

Pause A, Methot N, Svitkin Y, Merrick W, Sonenberg N . Dominant negative mutants of mammalian translation initiation factor eIF-4A define a critical role for eIF-4F in cap-dependent and cap-independent initiation of translation. EMBO J. 1994; 13(5):1205-15. PMC: 394930. DOI: 10.1002/j.1460-2075.1994.tb06370.x. View

Gomez-Gonzalez B, Garcia-Rubio M, Bermejo R, Gaillard H, Shirahige K, Marin A . Genome-wide function of THO/TREX in active genes prevents R-loop-dependent replication obstacles. EMBO J. 2011; 30(15):3106-19. PMC: 3160181. DOI: 10.1038/emboj.2011.206. View

Shen J, Zhang L, Zhao R . Biochemical characterization of the ATPase and helicase activity of UAP56, an essential pre-mRNA splicing and mRNA export factor. J Biol Chem. 2007; 282(31):22544-50. DOI: 10.1074/jbc.M702304200. View

10.

Cordin O, Banroques J, Tanner N, Linder P . The DEAD-box protein family of RNA helicases. Gene. 2005; 367:17-37. DOI: 10.1016/j.gene.2005.10.019. View

11.

Jensen M, Christensen M, Bonven B, Jensen T . Requirements for chromatin reassembly during transcriptional downregulation of a heat shock gene in Saccharomyces cerevisiae. FEBS J. 2008; 275(11):2956-64. DOI: 10.1111/j.1742-4658.2008.06451.x. View

12.

Johnson S, Cubberley G, Bentley D . Cotranscriptional recruitment of the mRNA export factor Yra1 by direct interaction with the 3' end processing factor Pcf11. Mol Cell. 2008; 33(2):215-26. PMC: 2659397. DOI: 10.1016/j.molcel.2008.12.007. View

13.

Piruat J, Aguilera A . A novel yeast gene, THO2, is involved in RNA pol II transcription and provides new evidence for transcriptional elongation-associated recombination. EMBO J. 1998; 17(16):4859-72. PMC: 1170815. DOI: 10.1093/emboj/17.16.4859. View

14.

Fairman-Williams M, Guenther U, Jankowsky E . SF1 and SF2 helicases: family matters. Curr Opin Struct Biol. 2010; 20(3):313-24. PMC: 2916977. DOI: 10.1016/j.sbi.2010.03.011. View

15.

Kistler A, Guthrie C . Deletion of MUD2, the yeast homolog of U2AF65, can bypass the requirement for sub2, an essential spliceosomal ATPase. Genes Dev. 2001; 15(1):42-9. PMC: 312603. DOI: 10.1101/gad.851301. View

16.

Mohr G, Del Campo M, Mohr S, Yang Q, Jia H, Jankowsky E . Function of the C-terminal domain of the DEAD-box protein Mss116p analyzed in vivo and in vitro. J Mol Biol. 2007; 375(5):1344-64. PMC: 2242632. DOI: 10.1016/j.jmb.2007.11.041. View

17.

Tutucci E, Stutz F . Keeping mRNPs in check during assembly and nuclear export. Nat Rev Mol Cell Biol. 2011; 12(6):377-84. DOI: 10.1038/nrm3119. View

18.

Aguilera A, Klein H . Genetic control of intrachromosomal recombination in Saccharomyces cerevisiae. I. Isolation and genetic characterization of hyper-recombination mutations. Genetics. 1988; 119(4):779-90. PMC: 1203464. DOI: 10.1093/genetics/119.4.779. View

19.

Cariani L, Thomas L, Brito J, Del Castillo J . Bismuth citrate in the quantification of inorganic phosphate and its utility in the determination of membrane-bound phosphatases. Anal Biochem. 2003; 324(1):79-83. DOI: 10.1016/j.ab.2003.09.008. View

20.

Strasser K, Hurt E . Splicing factor Sub2p is required for nuclear mRNA export through its interaction with Yra1p. Nature. 2001; 413(6856):648-52. DOI: 10.1038/35098113. View