Nuclear proteins that bind the Pre-mRNA 3′ splice site sequence r(UUAG/G) and the human telomeric DNA sequence d(TTAGGG)n

Fuyuki Ishikawa, Michael J Matunis, Gideon Dreyfuss, Thomas R. Cech

Research output: Contribution to journalArticle

Abstract

HeLa cell nuclear proteins that bind to single-stranded d(TTAGGG)n, the human telomeric DNA repeat, were identified and purified by a gel retardation assay. Immunological data and peptide sequencing experiments indicated that the purified proteins were identical or closely related to the heterogeneous nuclear ribonucleoproteins (hnRNPs) A1, A2-B1, D, and E and to nucleolin. These proteins bound to RNA oligonucleotides having r(UUAGGG) repeats more tightly than to DNA of the same sequence. The binding was sequence specific, as point mutation of any of the first 4 bases [r(UUAG)] abolished it. The fraction containing D and E hnRNPs was shown to bind specifically to a synthetic oligoribonucleotide having the 3′ splice site sequence of the human β-globin intervening sequence 1, which includes the sequence UUAGG. Proteins in this fraction were further identified by two-dimensional gel electrophoresis as D01, D02, D1*, and E0; intriguingly, these members of the hnRNP D and E groups are nuclear proteins that are not stably associated with hnRNP complexes. These studies establish the binding specificities of these D and E hnRNPs. Furthermore, they suggest the possibility that these hnRNPs could perhaps bind to chromosome telomeres, in addition to having a role in pre-mRNA metabolism.

Original languageEnglish (US)
Pages (from-to)4301-4310
Number of pages10
JournalMolecular and Cellular Biology
Volume13
Issue number7
StatePublished - Jul 1993
Externally publishedYes

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Heterogeneous-Nuclear Ribonucleoproteins
Heterogeneous-Nuclear Ribonucleoprotein D
RNA Splice Sites
RNA Precursors
Nuclear Proteins
Heterogeneous-Nuclear Ribonucleoprotein Group A-B
Oligoribonucleotides
Proteins
Globins
Telomere
Electrophoresis, Gel, Two-Dimensional
Electrophoretic Mobility Shift Assay
HeLa Cells
Point Mutation
Oligonucleotides
Introns
Chromosomes
RNA
Peptides
DNA

ASJC Scopus subject areas

  • Cell Biology
  • Genetics
  • Molecular Biology

Cite this

Nuclear proteins that bind the Pre-mRNA 3′ splice site sequence r(UUAG/G) and the human telomeric DNA sequence d(TTAGGG)n. / Ishikawa, Fuyuki; Matunis, Michael J; Dreyfuss, Gideon; Cech, Thomas R.

In: Molecular and Cellular Biology, Vol. 13, No. 7, 07.1993, p. 4301-4310.

Research output: Contribution to journalArticle

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abstract = "HeLa cell nuclear proteins that bind to single-stranded d(TTAGGG)n, the human telomeric DNA repeat, were identified and purified by a gel retardation assay. Immunological data and peptide sequencing experiments indicated that the purified proteins were identical or closely related to the heterogeneous nuclear ribonucleoproteins (hnRNPs) A1, A2-B1, D, and E and to nucleolin. These proteins bound to RNA oligonucleotides having r(UUAGGG) repeats more tightly than to DNA of the same sequence. The binding was sequence specific, as point mutation of any of the first 4 bases [r(UUAG)] abolished it. The fraction containing D and E hnRNPs was shown to bind specifically to a synthetic oligoribonucleotide having the 3′ splice site sequence of the human β-globin intervening sequence 1, which includes the sequence UUAGG. Proteins in this fraction were further identified by two-dimensional gel electrophoresis as D01, D02, D1*, and E0; intriguingly, these members of the hnRNP D and E groups are nuclear proteins that are not stably associated with hnRNP complexes. These studies establish the binding specificities of these D and E hnRNPs. Furthermore, they suggest the possibility that these hnRNPs could perhaps bind to chromosome telomeres, in addition to having a role in pre-mRNA metabolism.",
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