Sequence-specific Remodeling of a Topologically Complex RNP Substrate by Spb4

Overview

Journal Nat Struct Mol Biol

Specialties Cell Biology
Molecular Biology

Date 2022 Dec 9

PMID 36482249

Authors

Victor Emmanuel Cruz

Kamil Sekulski

Nagesh Peddada

Carolin Sailer

Sahana Balasubramanian

Christine S Weirich

Florian Stengel

Jan P Erzberger

Affiliations

Soon will be listed here.

Abstract

DEAD-box ATPases are ubiquitous enzymes essential in all aspects of RNA biology. However, the limited in vitro catalytic activities described for these enzymes are at odds with their complex cellular roles, most notably in driving large-scale RNA remodeling steps during the assembly of ribonucleoproteins (RNPs). We describe cryo-EM structures of 60S ribosomal biogenesis intermediates that reveal how context-specific RNA unwinding by the DEAD-box ATPase Spb4 results in extensive, sequence-specific remodeling of rRNA secondary structure. Multiple cis and trans interactions stabilize Spb4 in a post-catalytic, high-energy intermediate that drives the organization of the three-way junction at the base of rRNA domain IV. This mechanism explains how limited strand separation by DEAD-box ATPases is leveraged to provide non-equilibrium directionality and ensure efficient and accurate RNP assembly.

Citing Articles

Disassembly of unstable RNA structures by an E. coli DEAD-box chaperone accelerates ribosome assembly.

Sun Y, Woodson S Nucleic Acids Res. 2025; 53(4).

PMID: 39988318 PMC: 11840561. DOI: 10.1093/nar/gkaf104.

Intrinsically disordered RNA-binding motifs cooperate to catalyze RNA folding and drive phase separation.

Niedner-Boblenz A, Monecke T, Hennig J, Klostermann M, Hofweber M, Davydova E Nucleic Acids Res. 2024; 52(22):14205-14228.

PMID: 39558160 PMC: 11662656. DOI: 10.1093/nar/gkae1107.

Putting It All Together: The Roles of Ribosomal Proteins in Nucleolar Stages of 60S Ribosomal Assembly in the Yeast .

Ayers T, Woolford J Biomolecules. 2024; 14(8.

PMID: 39199362 PMC: 11353139. DOI: 10.3390/biom14080975.

The DEAD-box ATPase Dbp10/DDX54 initiates peptidyl transferase center formation during 60S ribosome biogenesis.

Cruz V, Weirich C, Peddada N, Erzberger J Nat Commun. 2024; 15(1):3296.

PMID: 38632236 PMC: 11024185. DOI: 10.1038/s41467-024-47616-7.

The RNA helicase Dbp10 coordinates assembly factor association with PTC maturation during ribosome biogenesis.

Mitterer V, Hamze H, Kunowska N, Stelzl U, Henras A, Hurt E Nucleic Acids Res. 2023; 52(4):1975-1987.

PMID: 38113283 PMC: 10899779. DOI: 10.1093/nar/gkad1206.

References

de la Cruz J, Kressler D, Rojo M, Tollervey D, Linder P . Spb4p, an essential putative RNA helicase, is required for a late step in the assembly of 60S ribosomal subunits in Saccharomyces cerevisiae. RNA. 1998; 4(10):1268-81. PMC: 1369699. DOI: 10.1017/s1355838298981158. View

Janke C, Magiera M, Rathfelder N, Taxis C, Reber S, Maekawa H . A versatile toolbox for PCR-based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes. Yeast. 2004; 21(11):947-62. DOI: 10.1002/yea.1142. View

McCann K, Charette J, Vincent N, Baserga S . A protein interaction map of the LSU processome. Genes Dev. 2015; 29(8):862-75. PMC: 4403261. DOI: 10.1101/gad.256370.114. View

Pettersen E, Goddard T, Huang C, Meng E, Couch G, Croll T . UCSF ChimeraX: Structure visualization for researchers, educators, and developers. Protein Sci. 2020; 30(1):70-82. PMC: 7737788. DOI: 10.1002/pro.3943. View

Garcia-Gomez J, Lebaron S, Froment C, Monsarrat B, Henry Y, de la Cruz J . Dynamics of the putative RNA helicase Spb4 during ribosome assembly in Saccharomyces cerevisiae. Mol Cell Biol. 2011; 31(20):4156-64. PMC: 3187284. DOI: 10.1128/MCB.05436-11. View

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

Martin R, Straub A, Doebele C, Bohnsack M . DExD/H-box RNA helicases in ribosome biogenesis. RNA Biol. 2012; 10(1):4-18. PMC: 3590236. DOI: 10.4161/rna.21879. View

Jankowsky E . RNA helicases at work: binding and rearranging. Trends Biochem Sci. 2010; 36(1):19-29. PMC: 3017212. DOI: 10.1016/j.tibs.2010.07.008. View

Bhaskaran H, Russell R . Kinetic redistribution of native and misfolded RNAs by a DEAD-box chaperone. Nature. 2007; 449(7165):1014-8. PMC: 2581903. DOI: 10.1038/nature06235. View

10.

Wu S, Tan D, Woolford Jr J, Dong M, Gao N . Atomic modeling of the ITS2 ribosome assembly subcomplex from cryo-EM together with mass spectrometry-identified protein-protein crosslinks. Protein Sci. 2016; 26(1):103-112. PMC: 5192979. DOI: 10.1002/pro.3045. View

11.

Kucukelbir A, Sigworth F, Tagare H . Quantifying the local resolution of cryo-EM density maps. Nat Methods. 2013; 11(1):63-5. PMC: 3903095. DOI: 10.1038/nmeth.2727. View

12.

Karbstein K . Quality control mechanisms during ribosome maturation. Trends Cell Biol. 2013; 23(5):242-50. PMC: 3640646. DOI: 10.1016/j.tcb.2013.01.004. View

13.

Kelley L, Mezulis S, Yates C, Wass M, Sternberg M . The Phyre2 web portal for protein modeling, prediction and analysis. Nat Protoc. 2015; 10(6):845-58. PMC: 5298202. DOI: 10.1038/nprot.2015.053. View

14.

Pettersen E, Goddard T, Huang C, Couch G, Greenblatt D, Meng E . UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem. 2004; 25(13):1605-12. DOI: 10.1002/jcc.20084. View

15.

Kressler D, Hurt E, Bassler J . A Puzzle of Life: Crafting Ribosomal Subunits. Trends Biochem Sci. 2017; 42(8):640-654. DOI: 10.1016/j.tibs.2017.05.005. View

16.

Bai X, Yan C, Yang G, Lu P, Ma D, Sun L . An atomic structure of human γ-secretase. Nature. 2015; 525(7568):212-217. PMC: 4568306. DOI: 10.1038/nature14892. View

17.

Zhang K . Gctf: Real-time CTF determination and correction. J Struct Biol. 2015; 193(1):1-12. PMC: 4711343. DOI: 10.1016/j.jsb.2015.11.003. View

18.

Yang Q, Jankowsky E . The DEAD-box protein Ded1 unwinds RNA duplexes by a mode distinct from translocating helicases. Nat Struct Mol Biol. 2006; 13(11):981-6. DOI: 10.1038/nsmb1165. View

19.

Bassler J, Kallas M, Pertschy B, Ulbrich C, Thoms M, Hurt E . The AAA-ATPase Rea1 drives removal of biogenesis factors during multiple stages of 60S ribosome assembly. Mol Cell. 2010; 38(5):712-21. PMC: 3372891. DOI: 10.1016/j.molcel.2010.05.024. View

20.

Daugeron M, Linder P . Dbp7p, a putative ATP-dependent RNA helicase from Saccharomyces cerevisiae, is required for 60S ribosomal subunit assembly. RNA. 1998; 4(5):566-81. PMC: 1369640. DOI: 10.1017/s1355838298980190. View