Abstract
While decapping plays a major role in mRNA turnover in yeast, biochemical evidence for a similar activity in mammalian cells has been elusive. We have now identified a decapping activity in HeLa cytoplasmic extracts that releases (7me)GDP from capped transcripts. Decapping is activated in extracts by the addition of (7me)GpppG, which specifically sequesters cap-binding proteins such as eIF4E and the deadenylase DAN/PARN. Similar to in vivo observations, the presence of a poly(A) tail represses decapping of RNAs in vitro in a poly(A)-binding protein-dependent fashion. AU-rich elements (AREs), which act as regulators of mRNA stability in vivo, are potent stimulators of decapping in vitro. The stimulation of decapping by AREs requires sequence-specific ARE-binding proteins. These data suggest that cap recognition and decapping play key roles in mediating mRNA turnover in mammalian cells.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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AT Rich Sequence / genetics
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Binding, Competitive
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Cell Extracts
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Cytoplasm / metabolism*
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Dinucleoside Phosphates / metabolism
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Eukaryotic Initiation Factor-4E
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Guanosine Diphosphate / analogs & derivatives
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Guanosine Diphosphate / metabolism
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HeLa Cells
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Humans
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Models, Genetic
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Peptide Initiation Factors / metabolism
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Poly A / genetics
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Poly A / metabolism
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Poly(A)-Binding Proteins
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RNA Caps / genetics*
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RNA Stability / genetics
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RNA, Messenger / genetics*
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RNA, Messenger / metabolism*
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RNA-Binding Proteins / metabolism
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Regulatory Sequences, Nucleic Acid / genetics*
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Saccharomyces cerevisiae
Substances
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Cell Extracts
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Dinucleoside Phosphates
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Eukaryotic Initiation Factor-4E
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Peptide Initiation Factors
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Poly(A)-Binding Proteins
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RNA Caps
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RNA, Messenger
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RNA-Binding Proteins
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7-methylguanosine 5'-diphosphate
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Guanosine Diphosphate
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Poly A
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7-methyl-diguanosine triphosphate