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. 1977 Sep;74(9):3705–3709. doi: 10.1073/pnas.74.9.3705

Secondary structure of heterogeneous nuclear RNA: Two classes of double-stranded RNA in native ribonucleoprotein*

James P Calvet 1, Thoru Pederson 1
PMCID: PMC431696  PMID: 410025

Abstract

Heterogeneous nuclear RNA (hnRNA) from HeLa cells contains intramolecular duplexes. Since hnRNA is associated with protein in vivo, it is possible that the double-stranded regions observed in deproteinized hnRNA form spontaneously upon the release of protein from single-stranded but potentially complementary sequences. We show here that this is not the case for a class of double-stranded sequences that is defined by resistance to RNases A + T1 at high ionic strength. Exposure of HeLa hnRNA·ribonucleoprotein (hnRNP) particles to Escherichia coli RNase III, a double-strand-specific endoribonuclease, destroys most of the sequences resistant to RNases A + T1. This effect is completely blocked when hnRNP is exposed to RNase III in the presence of an excess of purified double-stranded RNA. In addition, we show that there exist two classes of double-stranded RNA in hnRNP at a salt concentration of 0.13 M. These are distinguished by their relative resistance to RNases A + T1. The more stable double-stranded sequences, which are resistant to RNases A + T1 at 0.13 M, comprise 1.0-1.1% of the nucleotides in hnRNP. The less stable double-stranded sequences comprise an additional 1.5-2.0% of the nucleotides in hnRNP. These are sensitive to RNase III at 0.13 M, but are not resistant to RNases A + T1 unless the salt concentration is raised to 0.63 M. The demonstration that double-stranded sequences resistant to RNases A + T1 exist in native ribonucleoprotein and are not artifacts of deproteinization now makes it appropriate to seriously consider their possible functional role in hnRNA metabolism, perhaps as binding sites for regulatory proteins involved in mRNA processing.

Keywords: inverted-repeat DNA transcripts, Escherichia coli RNase III, RNases A + T1, mRNA processing

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Selected References

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  1. Angerer R. C., Davidson E. H., Britten R. J. DNA sequence organization in the mollusc Aplysia californica. Cell. 1975 Sep;6(1):29–39. doi: 10.1016/0092-8674(75)90070-7. [DOI] [PubMed] [Google Scholar]
  2. Augenlicht L. H., Lipkin M. Appearance of rapidly labeled, high molecular weight RNA in nuclear ribonucleoprotein. Release from chromatin and association with protein. J Biol Chem. 1976 May 10;251(9):2592–2599. [PubMed] [Google Scholar]
  3. Bishayee S., Maitra U. Specificity of cleavage by ribonuclease III. Biochem Biophys Res Commun. 1976 Nov 22;73(2):306–313. doi: 10.1016/0006-291x(76)90708-7. [DOI] [PubMed] [Google Scholar]
  4. Crouch R. J. Ribonuclease 3 does not degrade deoxyribonucleic acid-ribonucleic acid hybrids. J Biol Chem. 1974 Feb 25;249(4):1314–1316. [PubMed] [Google Scholar]
  5. Dunn J. J. RNase III cleavage of single-stranded RNA. Effect of ionic strength on the fideltiy of cleavage. J Biol Chem. 1976 Jun 25;251(12):3807–3814. [PubMed] [Google Scholar]
  6. Fedoroff N., Wellauer P. K., Wall R. Intermolecular duplexes in heterogeneous nuclear RNA from HeLa cells. Cell. 1977 Apr;10(4):597–610. doi: 10.1016/0092-8674(77)90092-7. [DOI] [PubMed] [Google Scholar]
  7. Firtel R. A., Pederson T. Ribonucleoprotein particles containing heterogeneous nuclear RNA in the cellular slime mold Dictyostelium discoideum. Proc Natl Acad Sci U S A. 1975 Jan;72(1):301–305. doi: 10.1073/pnas.72.1.301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Foe V. E., Wilkinson L. E., Laird C. D. Comparative organization of active transcription units in Oncopeltus fasciatus. Cell. 1976 Sep;9(1):131–146. doi: 10.1016/0092-8674(76)90059-3. [DOI] [PubMed] [Google Scholar]
  9. Hsu M. T., Jelinek W. R. Mapping of inverted repeated DNA sequences within the genome of simian virus 40. Proc Natl Acad Sci U S A. 1977 Apr;74(4):1631–1634. doi: 10.1073/pnas.74.4.1631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jelinek W., Darnell J. E. Double-stranded regions in heterogeneous nuclear RNA from Hela cells. Proc Natl Acad Sci U S A. 1972 Sep;69(9):2537–2541. doi: 10.1073/pnas.69.9.2537. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kish V. M., Pederson T. Heterogeneous nuclear RNA secondary structure: oligo (U) sequences base-paired with poly (A) and their possible role as binding sites for heterogeneous nuclear RNA-specific proteins. Proc Natl Acad Sci U S A. 1977 Apr;74(4):1426–1430. doi: 10.1073/pnas.74.4.1426. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kish V. M., Pederson T. Ribonucleoprotein organization of polyadenylate sequences in HeLa cell heterogeneous nuclear RNA. J Mol Biol. 1975 Jun 25;95(2):227–238. doi: 10.1016/0022-2836(75)90392-7. [DOI] [PubMed] [Google Scholar]
  13. Lukanidin E. M., Zalmanzon E. S., Komaromi L., Samarina O. P., Georgiev G. P. Structure and function of informofers. Nat New Biol. 1972 Aug 16;238(85):193–197. doi: 10.1038/newbio238193a0. [DOI] [PubMed] [Google Scholar]
  14. Malcolm D. B., Sommerville J. The structure of chromosome-derived ribonucleoprotein in oocytes of Triturus cristatus carnifex (Laurenti). Chromosoma. 1974;48(2):137–158. doi: 10.1007/BF00283960. [DOI] [PubMed] [Google Scholar]
  15. Miller O. L., Jr, Bakken A. H. Morphological studies of transcription. Acta Endocrinol Suppl (Copenh) 1972;168:155–177. doi: 10.1530/acta.0.071s155. [DOI] [PubMed] [Google Scholar]
  16. Monneron A., Bernhard W. Fine structural organization of the interphase nucleus in some mammalian cells. J Ultrastruct Res. 1969 May;27(3):266–288. doi: 10.1016/s0022-5320(69)80017-1. [DOI] [PubMed] [Google Scholar]
  17. Naora H., Whitelam J. M. Presence of sequences hybridisable to dsRNA in cytoplasmic mRNA molecules. Nature. 1975 Aug 28;256(5520):756–759. doi: 10.1038/256756a0. [DOI] [PubMed] [Google Scholar]
  18. Pederson T. Gene activation in eukaryotes: are nuclear acidic proteins the cause or the effect? Proc Natl Acad Sci U S A. 1974 Mar;71(3):617–621. doi: 10.1073/pnas.71.3.617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pederson T. Proteins associated with heterogeneous nuclear RNA in eukaryotic cells. J Mol Biol. 1974 Feb 25;83(2):163–183. doi: 10.1016/0022-2836(74)90386-6. [DOI] [PubMed] [Google Scholar]
  20. Perlman S., Phillips C., Bishop J. O. A study of foldback DNA. Cell. 1976 May;8(1):33–42. doi: 10.1016/0092-8674(76)90182-3. [DOI] [PubMed] [Google Scholar]
  21. Robertson H. D., Dickson E. RNA processing and the control of gene expression. Brookhaven Symp Biol. 1975 Jul;(26):240–266. [PubMed] [Google Scholar]
  22. Robertson H. D., Webster R. E., Zinder N. D. Purification and properties of ribonuclease III from Escherichia coli. J Biol Chem. 1968 Jan 10;243(1):82–91. [PubMed] [Google Scholar]
  23. Ryskov A. P., Farashyan V. R., Georgiev G. P. Ribonuclease-stable base sequences specific exclusively for giant dRNA. Biochim Biophys Acta. 1972 Apr 12;262(4):568–572. doi: 10.1016/0005-2787(72)90502-3. [DOI] [PubMed] [Google Scholar]
  24. Ryskov A. P., Kramerov D. A., Georgiev G. P. The structural organization of nuclear messenger RNA precursor. I. Reassociation and hybridization properties of double-stranded hairpin-like loops in messenger RNA precursor. Biochim Biophys Acta. 1976 Oct 4;447(2):214–229. doi: 10.1016/0005-2787(76)90344-0. [DOI] [PubMed] [Google Scholar]
  25. Ryskov A. P., Tokarskaya O. V., Georgiev G. P., Coutelle C., Thiele B. Globin mRNA contains a sequence complementary to double-stranded region of nuclear pre-mRNA. Nucleic Acids Res. 1976 Jun;3(6):1487–1498. doi: 10.1093/nar/3.6.1487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Ryskov A. P., Yenikolopov G. N., Limborska S. A. Complementary regions of the nuclear precursor of messenger RNA. FEBS Lett. 1974 Oct 1;47(1):98–102. doi: 10.1016/0014-5793(74)80434-5. [DOI] [PubMed] [Google Scholar]
  27. Shen C. K., Hearst J. E. Mapping of sequences with 2-fold symmetry on the simian virus 40 genome: a photochemical crosslinking approach. Proc Natl Acad Sci U S A. 1977 Apr;74(4):1363–1367. doi: 10.1073/pnas.74.4.1363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sommerville J. Ribonucleoprotein particles derived from the lampbrush chromosomes of newt oocytes. J Mol Biol. 1973 Aug 15;78(3):487–503. doi: 10.1016/0022-2836(73)90470-1. [DOI] [PubMed] [Google Scholar]
  29. Stevens B. J., Swift H. RNA transport from nucleus to cytoplasm in Chironomus salivary glands. J Cell Biol. 1966 Oct;31(1):55–77. doi: 10.1083/jcb.31.1.55. [DOI] [PMC free article] [PubMed] [Google Scholar]

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