Ribozyme-mediated RNA degradation in nuclei suspension.
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.
Heidenreich
O
O
Scripps Research Institute, Department of Neuropharmacology, La Jolla, CA 92037, USA.
Kang
S H
SH
Brown
D A
DA
Xu
X
X
Swiderski
P
P
Rossi
J J
JJ
Eckstein
F
F
Nerenberg
M
M
D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D002461:Cell Line, Transformed; D002467:Cell Nucleus / Q000378:metabolism*; D019286:Endopeptidase K; D005347:Fibroblasts / Q000648:ultrastructure; D016355:Genes, pX; D015368:Human T-lymphotropic virus 1 / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D016337:RNA, Catalytic / Q000737:chemistry / Q000378:metabolism*; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000494:pharmacology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013329:Structure-Activity Relationship
D016337:RNA, Catalytic; D012367:RNA, Viral; D012967:Sodium Dodecyl Sulfate; D012697:Serine Endopeptidases; D019286:Endopeptidase K