Ribozyme-mediated RNA degradation in nuclei suspension

O Heidenreich; S H Kang; D A Brown; X Xu; P Swiderski; J J Rossi; F Eckstein; M Nerenberg

doi:10.1093/nar/23.12.2223

Ribozyme-mediated RNA degradation in nuclei suspension

O Heidenreich
, S H Kang
, D A Brown
, X Xu
, P Swiderski
, J J Rossi
, F Eckstein
, M Nerenberg

Research output: Contribution to journal › Article › peer-review

Abstract

Ribozymes containing 2'-fluoro- and 2'-amino-modified pyrimidine nucleosides in combination with terminal phosphorothioate linkages were targeted against HTLV-I tax RNA. In order to examine the activity of such chemically modified ribozymes in the nuclear environment, they were incubated with nuclei of a Tax-transformed mouse fibroblast cell line. Ribozyme cleavage of tax RNA was analyzed by the RNase protection assay. Comparison of the cleavage of tax RNA isolated nuclei with that of tax RNA present in nuclei suspension revealed a 30 times more efficient cleavage of the latter one. Pre-treatment with proteinase K and SDS abolished the enhancement of the ribozyme-mediated RNA cleavage. Catalytically inactive ribozymes did not yield any cleavage products. These results demonstrate an augmenting effect of nuclear proteins on the ribozyme-mediated RNA cleavage.

Original language	English
Pages (from-to)	2223-8
Number of pages	6
Journal	Nucleic acids research
Volume	23
Issue number	12
DOIs	https://doi.org/10.1093/nar/23.12.2223
Publication status	Published - 25 Jun 1995
Externally published	Yes

Keywords

Animals
Base Sequence
Catalysis
Cell Line, Transformed
Cell Nucleus/metabolism
Endopeptidase K
Fibroblasts/ultrastructure
Genes, pX
Human T-lymphotropic virus 1/genetics
Mice
Molecular Sequence Data
RNA, Catalytic/chemistry
RNA, Viral/chemistry
Serine Endopeptidases/pharmacology
Sodium Dodecyl Sulfate/pharmacology
Structure-Activity Relationship

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10.1093/nar/23.12.2223

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title = "Ribozyme-mediated RNA degradation in nuclei suspension",

abstract = "Ribozymes containing 2'-fluoro- and 2'-amino-modified pyrimidine nucleosides in combination with terminal phosphorothioate linkages were targeted against HTLV-I tax RNA. In order to examine the activity of such chemically modified ribozymes in the nuclear environment, they were incubated with nuclei of a Tax-transformed mouse fibroblast cell line. Ribozyme cleavage of tax RNA was analyzed by the RNase protection assay. Comparison of the cleavage of tax RNA isolated nuclei with that of tax RNA present in nuclei suspension revealed a 30 times more efficient cleavage of the latter one. Pre-treatment with proteinase K and SDS abolished the enhancement of the ribozyme-mediated RNA cleavage. Catalytically inactive ribozymes did not yield any cleavage products. These results demonstrate an augmenting effect of nuclear proteins on the ribozyme-mediated RNA cleavage.",

keywords = "Animals, Base Sequence, Catalysis, Cell Line, Transformed, Cell Nucleus/metabolism, Endopeptidase K, Fibroblasts/ultrastructure, Genes, pX, Human T-lymphotropic virus 1/genetics, Mice, Molecular Sequence Data, RNA, Catalytic/chemistry, RNA, Viral/chemistry, Serine Endopeptidases/pharmacology, Sodium Dodecyl Sulfate/pharmacology, Structure-Activity Relationship",

author = "O Heidenreich and Kang, \{S H\} and Brown, \{D A\} and X Xu and P Swiderski and Rossi, \{J J\} and F Eckstein and M Nerenberg",

year = "1995",

month = jun,

day = "25",

doi = "10.1093/nar/23.12.2223",

language = "English",

volume = "23",

pages = "2223--8",

journal = "Nucleic acids research",

issn = "0305-1048",

publisher = "Oxford University Press",

number = "12",

}

TY - JOUR

T1 - Ribozyme-mediated RNA degradation in nuclei suspension

AU - Heidenreich, O

AU - Kang, S H

AU - Brown, D A

AU - Xu, X

AU - Swiderski, P

AU - Rossi, J J

AU - Eckstein, F

AU - Nerenberg, M

PY - 1995/6/25

Y1 - 1995/6/25

N2 - Ribozymes containing 2'-fluoro- and 2'-amino-modified pyrimidine nucleosides in combination with terminal phosphorothioate linkages were targeted against HTLV-I tax RNA. In order to examine the activity of such chemically modified ribozymes in the nuclear environment, they were incubated with nuclei of a Tax-transformed mouse fibroblast cell line. Ribozyme cleavage of tax RNA was analyzed by the RNase protection assay. Comparison of the cleavage of tax RNA isolated nuclei with that of tax RNA present in nuclei suspension revealed a 30 times more efficient cleavage of the latter one. Pre-treatment with proteinase K and SDS abolished the enhancement of the ribozyme-mediated RNA cleavage. Catalytically inactive ribozymes did not yield any cleavage products. These results demonstrate an augmenting effect of nuclear proteins on the ribozyme-mediated RNA cleavage.

AB - Ribozymes containing 2'-fluoro- and 2'-amino-modified pyrimidine nucleosides in combination with terminal phosphorothioate linkages were targeted against HTLV-I tax RNA. In order to examine the activity of such chemically modified ribozymes in the nuclear environment, they were incubated with nuclei of a Tax-transformed mouse fibroblast cell line. Ribozyme cleavage of tax RNA was analyzed by the RNase protection assay. Comparison of the cleavage of tax RNA isolated nuclei with that of tax RNA present in nuclei suspension revealed a 30 times more efficient cleavage of the latter one. Pre-treatment with proteinase K and SDS abolished the enhancement of the ribozyme-mediated RNA cleavage. Catalytically inactive ribozymes did not yield any cleavage products. These results demonstrate an augmenting effect of nuclear proteins on the ribozyme-mediated RNA cleavage.

KW - Animals

KW - Base Sequence

KW - Catalysis

KW - Cell Line, Transformed

KW - Cell Nucleus/metabolism

KW - Endopeptidase K

KW - Fibroblasts/ultrastructure

KW - Genes, pX

KW - Human T-lymphotropic virus 1/genetics

KW - Mice

KW - Molecular Sequence Data

KW - RNA, Catalytic/chemistry

KW - RNA, Viral/chemistry

KW - Serine Endopeptidases/pharmacology

KW - Sodium Dodecyl Sulfate/pharmacology

KW - Structure-Activity Relationship

U2 - 10.1093/nar/23.12.2223

DO - 10.1093/nar/23.12.2223

M3 - Article

C2 - 7610051

SN - 0305-1048

VL - 23

SP - 2223

EP - 2228

JO - Nucleic acids research

JF - Nucleic acids research

IS - 12

ER -

Ribozyme-mediated RNA degradation in nuclei suspension

Abstract

Keywords

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