Pre-mRNA and mRNA binding of human nuclear DNA helicase II (RNA helicase A).
Nuclear DNA helicase II (NDH II), alternatively named RNA helicase A, seems to function as a pre-mRNA and mRNA binding protein in human cells. Immunofluorescence studies of NDH II gave a highly diffused nucleoplasmic staining that was similar to that of hnRNP A1 but differed from the localization of the RNA splicing factor Sc-35. Upon transcriptional inhibition, NDH II migrated from the nucleus into the cytoplasm. During mitosis, NDH II was released into the cytoplasm during pro- to metaphase, and was gradually recruited back into telophase nuclei. The timing of nuclear import of NDH II at telophase was found to be later than that of hnRNP A1 but paralleled that of Sc-35. At the ultrastructural level, both NDH II and hnRNP A1 were identified within perichromatin ribonucleoparticle fibrils. However, the subnuclear distributions of NDH II and hnRNP A1 were not overlapping. NDH II could be extracted together with poly(A)-containing mRNA from HeLa cell nuclei and, to a much lesser extent, from the cytoplasm. Following transcriptional inhibition, NDH II was preferentially associated with mRNA from the cytosol, which biochemically confirmed the microscopic observations. Although NDH II is mainly a nuclear enzyme, it is apparently not associated with the nuclear matrix, since it could be extracted with 2 M NaCl from DNase I-treated nuclei. Our cellular and biochemical observations strongly suggest that NDH II is a pre-mRNA and mRNA binding protein. Its significant affinity for ssDNA, but not for dsDNA, points to a transient role in DNA binding during the process of transcript formation. According to our model, single-stranded DNA might be necessary to retain NDH II in the nuclear compartment.
Zhang
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Department of Biochemistry and Department of Electron Microscopy and Molecular Cytology, Institute for Molecular Biotechnology, Beutenbergstrasse 11, D-07745 Jena, Germany.
Herrmann
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C
Grosse
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F
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D004265:DNA Helicases; D003609:Dactinomycin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D008854:Microscopy, Electron; D015530:Nuclear Matrix / Q000378:metabolism; D012322:RNA Precursors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D011994:Recombinant Proteins; D012259:Ribonuclease, Pancreatic / Q000494:pharmacology; D012261:Ribonucleoproteins / Q000378:metabolism; D012965:Sodium Chloride / Q000378:metabolism
D012322:RNA Precursors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D003609:Dactinomycin; D012965:Sodium Chloride; D012259:Ribonuclease, Pancreatic; D000251:Adenosine Triphosphatases; D004265:DNA Helicases