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Title | Structural insights into how Prp5 proofreads the pre-mRNA branch site. |
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Journal, issue, pages | Nature, Vol. 596, Issue 7871, Page 296-300, Year 2021 |
Publish date | Aug 4, 2021 |
Authors | Zhenwei Zhang / Norbert Rigo / Olexandr Dybkov / Jean-Baptiste Fourmann / Cindy L Will / Vinay Kumar / Henning Urlaub / Holger Stark / Reinhard Lührmann / |
PubMed Abstract | During the splicing of introns from precursor messenger RNAs (pre-mRNAs), the U2 small nuclear ribonucleoprotein (snRNP) must undergo stable integration into the spliceosomal A complex-a poorly ...During the splicing of introns from precursor messenger RNAs (pre-mRNAs), the U2 small nuclear ribonucleoprotein (snRNP) must undergo stable integration into the spliceosomal A complex-a poorly understood, multistep process that is facilitated by the DEAD-box helicase Prp5 (refs. ). During this process, the U2 small nuclear RNA (snRNA) forms an RNA duplex with the pre-mRNA branch site (the U2-BS helix), which is proofread by Prp5 at this stage through an unclear mechanism. Here, by deleting the branch-site adenosine (BS-A) or mutating the branch-site sequence of an actin pre-mRNA, we stall the assembly of spliceosomes in extracts from the yeast Saccharomyces cerevisiae directly before the A complex is formed. We then determine the three-dimensional structure of this newly identified assembly intermediate by cryo-electron microscopy. Our structure indicates that the U2-BS helix has formed in this pre-A complex, but is not yet clamped by the HEAT domain of the Hsh155 protein (Hsh155), which exhibits an open conformation. The structure further reveals a large-scale remodelling/repositioning of the U1 and U2 snRNPs during the formation of the A complex that is required to allow subsequent binding of the U4/U6.U5 tri-snRNP, but that this repositioning is blocked in the pre-A complex by the presence of Prp5. Our data suggest that binding of Hsh155 to the bulged BS-A of the U2-BS helix triggers closure of Hsh155, which in turn destabilizes Prp5 binding. Thus, Prp5 proofreads the branch site indirectly, hindering spliceosome assembly if branch-site mutations prevent the remodelling of Hsh155. Our data provide structural insights into how a spliceosomal helicase enhances the fidelity of pre-mRNA splicing. |
External links | Nature / PubMed:34349264 / PubMed Central |
Methods | EM (single particle) |
Resolution | 4.1 - 13.0 Å |
Structure data | EMDB-13028, PDB-7oqb: EMDB-13029, PDB-7oqc: EMDB-13030: EMDB-13031: EMDB-13032: EMDB-13033, PDB-7oqe: |
Source |
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Keywords | SPLICING / S. cerevisiae / pre-A complex / Prp5 / U1 snRNP / U2 snRNP / prespliceosome |