Abstract
Splicing, a key step in the eukaryotic gene-expression pathway, converts precursor messenger RNA (pre-mRNA) into mRNA by excising introns and ligating exons. This task is accomplished by the spliceosome, a macromolecular machine that must undergo sequential conformational changes to establish its active site. Each of these major changes requires a dedicated DExD/H-box ATPase, but how these enzymes are activated remain obscure. Here we show that Prp28, a yeast DEAD-box ATPase, transiently interacts with the conserved 5' splice-site (5'SS) GU dinucleotide and makes splicing-dependent contacts with the U1 snRNP protein U1C, and U4/U6.U5 tri-snRNP proteins, Prp8, Brr2, and Snu114. We further show that Prp28's ATPase activity is potentiated by the phosphorylated Npl3, but not the unphosphorylated Npl3, thus suggesting a strategy for regulating DExD/H-box ATPases. We propose that Npl3 is a functional counterpart of the metazoan-specific Prp28 N-terminal region, which can be phosphorylated and serves as an anchor to human spliceosome.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphate / metabolism
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DEAD-box RNA Helicases / genetics
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DEAD-box RNA Helicases / metabolism*
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Humans
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Multiprotein Complexes / genetics
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Multiprotein Complexes / metabolism
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Mutation
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Phosphorylation
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Protein Binding
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RNA Helicases / genetics
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RNA Helicases / metabolism
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RNA Precursors / genetics
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RNA Precursors / metabolism
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RNA Splicing*
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RNA-Binding Proteins / genetics
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RNA-Binding Proteins / metabolism*
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Ribonuclease H / metabolism
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Ribonucleoproteins, Small Nuclear / metabolism
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism*
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Spliceosomes / genetics
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Spliceosomes / metabolism*
Substances
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Multiprotein Complexes
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NPL3 protein, S cerevisiae
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Nuclear Proteins
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RNA Precursors
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RNA-Binding Proteins
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Ribonucleoproteins, Small Nuclear
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Saccharomyces cerevisiae Proteins
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Adenosine Triphosphate
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Ribonuclease H
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BRR2 protein, S cerevisiae
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PRP28 protein, S cerevisiae
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DEAD-box RNA Helicases
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RNA Helicases