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Distinct RNP complexes of shuttling hnRNP proteins with pre-mRNA and mRNA: candidate intermediates in formation and
1Department of Biochemistry and Molecular Biology, Mount Sinai School of Medicine, New York, New York 10029-6574, USA.
Abstract:
Nascent pre-mRNAs associate with hnRNP proteins in hnRNP complexes, the natural substrates for mRNA processing. Several lines of evidence indicate that hnRNP complexes undergo substantial remodeling during mRNA formation and export. Here we report the isolation of three distinct types of pre-mRNP and mRNP complexes from HeLa cells associated with hnRNP A1, a shuttling hnRNP protein. Based on their RNA and protein compositions, these complexes are likely to represent distinct stages in the nucleocytoplasmic shuttling pathway of hnRNP A1 with its bound RNAs. In the cytoplasm, A1 is associated with its nuclear import receptor (transportin), the cytoplasmic poly(A)-binding protein, and mRNA. In the nucleus, A1 is found in two distinct types of complexes that are differently associated with nuclear structures. One class contains pre-mRNA and mRNA and is identical to previously described hnRNP complexes. The other class behaves as freely diffusible nuclear mRNPs (nmRNPs) at late nuclear stages of maturation and possibly associated with nuclear mRNA export. These nmRNPs differ from hnRNPs in that while they contain shuttling hnRNP proteins, the mRNA export factor REF, and mRNA, they do not contain nonshuttling hnRNP proteins or pre-mRNA. Importantly, nmRNPs also contain proteins not found in hnRNP complexes. These include the alternatively spliced isoforms D01 and D02 of the hnRNP D proteins, the E0 isoform of the hnRNP E proteins, and LRP130, a previously reported protein with unknown function that appears to have a novel type of RNA-binding domain. The characteristics of these complexes indicate that they result from RNP remodeling associated with mRNA maturation and delineate specific changes in RNP protein composition during formation and transport of mRNA in vivo.
Insights
Researchers identified distinct nuclear messenger ribonucleoprotein (mRNP) complexes involved in mRNA export. These complexes, containing specific hnRNP proteins and export factors, reveal dynamic RNP remodeling during mRNA maturation and transport.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Nascent pre-mRNAs form hnRNP complexes, undergoing remodeling during mRNA processing and export.
- hnRNP A1 is a shuttling protein involved in mRNA transport.
Purpose of the Study:
- To isolate and characterize distinct pre-messenger ribonucleoprotein (pre-mRNP) and messenger ribonucleoprotein (mRNP) complexes associated with hnRNP A1.
- To elucidate the stages of nucleocytoplasmic shuttling involving hnRNP A1 and its bound RNAs.
Main Methods:
- Isolation of three distinct pre-mRNP and mRNP complexes from HeLa cells.
- Analysis of RNA and protein compositions of isolated complexes.
- Characterization of nuclear structures associated with these complexes.
Main Results:
- Identified three types of hnRNP A1-associated complexes representing different stages of mRNA processing and export.
- Described cytoplasmic complexes with importin, PABP, and mRNA.
- Characterized nuclear complexes: one containing pre-mRNA/mRNA (hnRNPs), and another freely diffusible nmRNPs containing export factors (REF) and mRNA, but not pre-mRNA.
- Discovered novel proteins in nmRNPs, including hnRNP D isoforms (D01, D02), hnRNP E0, and LRP130.
Conclusions:
- The isolated complexes represent distinct stages in the nucleocytoplasmic shuttling pathway of hnRNP A1.
- nmRNPs are distinct from hnRNPs, indicating specific RNP remodeling during mRNA maturation and export.
- These findings delineate changes in RNP composition during in vivo mRNA formation and transport.