Biomaterial Database

Actinomycin D stimulates the transcription of rRNA minigenes transfected into mouse cells. Implications for the in vivo hypersensitivity of rRNA gene transcription. 1995

K V Hadjiolova, and A A Hadjiolov, and J P Bachellerie

Laboratoire de Biologie Moléculaire Eukaryote, Université Paul Sabatier, Toulouse, France.

The in vivo hypersensitivity of eukaryotic rRNA gene transcription to actinomycin D has long been known, but this effect could not be reproduced in model systems and its molecular mechanisms remain uncertain. We studied the action of actinomycin D using mouse rRNA minigenes (with RNA polymerase I promoter and terminator signals), carrying truncated mouse or human rDNA inserts, which are faithfully transcribed upon transient transfection into mouse cells. Low concentrations (0.01-0.08 micrograms/ml) of actinomycin D caused within 1-2 h a 2-7-fold stimulation of the transcription of rRNA minigenes which is inversely related to the size of the rDNA transcript. With transcripts longer than 3 kb the effect was reversed and at 4 kb a practically complete inhibition of the formation of full-length transcripts was observed, accompanied, however, by an enhanced accumulation of unfinished rDNA transcripts. The dependence of actinomycin D action on transcript length was also observed with lacZ gene segments of different size inserted into the mouse rRNA minigenes. The transcription initiation of endogenous rRNA genes was also stimulated by the low doses of actinomycin D as indicated by the enhanced synthesis of unfinished rDNA transcripts (spanning mainly the 5' external transcribed spacer), whereas the synthesis of full-length transcripts was abolished. Removal of actinomycin D from the medium caused within 8-24 h a dramatic increase of the transcription from all rRNA minigenes tested. This stimulation was also inversely related to the size of the transcripts and varied from twofold to fivefold for the 3-4-kb transcripts to about 50-80-fold for the basic minigene transcript (395 nucleotides). The amount of endogenous aborted rDNA transcripts was also markedly increased, but the synthesis of full-length transcripts was not restored even 24 h after removal of the drug. The present results reproduce in a model cellular system the in vivo hypersensitivity of rRNA gene transcription to actinomycin D and reveal that the major factor involved is the size of the rRNA gene transcript. This effect requires only the basic rRNA gene promoter and terminator signals and does not depend on the G + C content of the RNA polymerase I transcripts. We suggest that at low concentrations, the intercalation of actinomycin D changes the conformation of DNA in the promoter region in a manner that stimulates the transcription of both endogenous and transfected rRNA genes.(ABSTRACT TRUNCATED AT 400 WORDS)

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D009838	Oligodeoxyribonucleotides	A group of deoxyribonucleotides (up to 12) in which the phosphate residues of each deoxyribonucleotide act as bridges in forming diester linkages between the deoxyribose moieties.	Oligodeoxynucleotide,Oligodeoxyribonucleotide,Oligodeoxynucleotides
D003609	Dactinomycin	A compound composed of a two CYCLIC PEPTIDES attached to a phenoxazine that is derived from STREPTOMYCES parvullus. It binds to DNA and inhibits RNA synthesis (transcription), with chain elongation more sensitive than initiation, termination, or release. As a result of impaired mRNA production, protein synthesis also declines after dactinomycin therapy. (From AMA Drug Evaluations Annual, 1993, p2015)	Actinomycin,Actinomycin D,Meractinomycin,Cosmegen,Cosmegen Lyovac,Lyovac-Cosmegen,Lyovac Cosmegen,Lyovac, Cosmegen,LyovacCosmegen
D004275	DNA, Ribosomal	DNA sequences encoding RIBOSOMAL RNA and the segments of DNA separating the individual ribosomal RNA genes, referred to as RIBOSOMAL SPACER DNA.	Ribosomal DNA,rDNA
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D012335	RNA, Ribosomal	The most abundant form of RNA. Together with proteins, it forms the ribosomes, playing a structural role and also a role in ribosomal binding of mRNA and tRNAs. Individual chains are conventionally designated by their sedimentation coefficients. In eukaryotes, four large chains exist, synthesized in the nucleolus and constituting about 50% of the ribosome. (Dorland, 28th ed)	Ribosomal RNA,15S RNA,RNA, 15S
D014158	Transcription, Genetic	The biosynthesis of RNA carried out on a template of DNA. The biosynthesis of DNA from an RNA template is called REVERSE TRANSCRIPTION.	Genetic Transcription
D014162	Transfection	The uptake of naked or purified DNA by CELLS, usually meaning the process as it occurs in eukaryotic cells. It is analogous to bacterial transformation (TRANSFORMATION, BACTERIAL) and both are routinely employed in GENE TRANSFER TECHNIQUES.	Transfections