The Prp8 RNase H-like domain inhibits Brr2-mediated U4/U6 snRNA unwinding by blocking Brr2 loading onto the U4 snRNA.
Genes Dev
; 26(21): 2422-34, 2012 Nov 01.
Article
em En
| MEDLINE
| ID: mdl-23124066
ABSTRACT
The spliceosomal RNA helicase Brr2 catalyzes unwinding of the U4/U6 snRNA duplex, an essential step for spliceosome catalytic activation. Brr2 is regulated in part by the spliceosomal Prp8 protein by an unknown mechanism. We demonstrate that the RNase H (RH) domain of yeast Prp8 binds U4/U6 small nuclear RNA (snRNA) with the single-stranded regions of U4 and U6 preceding U4/U6 stem I, contributing to its binding. Via cross-linking coupled with mass spectrometry, we identify RH domain residues that contact the U4/U6 snRNA. We further demonstrate that the same single-stranded region of U4 preceding U4/U6 stem I is recognized by Brr2, indicating that it translocates along U4 and first unwinds stem I of the U4/U6 duplex. Finally, we show that the RH domain of Prp8 interferes with U4/U6 unwinding by blocking Brr2's interaction with the U4 snRNA. Our data reveal a novel mechanism whereby Prp8 negatively regulates Brr2 and potentially prevents premature U4/U6 unwinding during splicing. They also support the idea that the RH domain acts as a platform for the exchange of U6 snRNA for U1 at the 5' splice site. Our results provide insights into the mechanism whereby Brr2 unwinds U4/U6 and show how this activity is potentially regulated prior to spliceosome activation.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Saccharomyces cerevisiae
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RNA Nuclear Pequeno
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Ribonucleoproteína Nuclear Pequena U5
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Ribonucleoproteína Nuclear Pequena U4-U6
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RNA Helicases
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Proteínas de Saccharomyces cerevisiae
Idioma:
En
Revista:
Genes Dev
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2012
Tipo de documento:
Article
País de afiliação:
Alemanha