Biomaterial Database

Elevated expression of M1 and M2 components and drug-induced posttranscriptional modulation of ribonucleotide reductase in a hydroxyurea-resistant mouse cell line. 1987

G A McClarty, and A K Chan, and Y Engstrom, and J A Wright, and L Thelander

Department of Biochemistry, Karolinska Institute, Stockholm, Sweden.

Ribonucleotide reductase, a rate-limiting enzyme in the synthesis of DNA, consists of two nonidentical subunits, proteins M1 and M2. Hydroxyurea, a specific inhibitor of DNA synthesis, acts by destroying the unique tyrosyl free radical of protein M2. In the past, we have described a mouse L cell line which exhibited a stable resistance to high concentrations of hydroxyurea [McClarty, G. A., Chan, A., & Wright, J.A. (1986) Somat. Cell Mol. Genet. 12, 121-131]. When this line was grown in the absence of hydroxyurea, the cells contained a modest but stable elevation in ribonucleotide reductase activity. However, the activity was further increased on the addition of drug to the culture medium. This was accompanied by an increase in protein M2 activity as shown by activity titration experiments. Likewise, removal of hydroxyurea resulted in a decrease in M2 activity. In the present study, we make use of recently isolated cDNAs and monoclonal antibodies for both the M1 and M2 proteins to further our understanding of the mechanism of hydroxyurea resistance at the molecular level in a subclone of this cell line. Our results indicated that protein M1 levels were elevated 2-3-fold and protein M2 levels were increased about 50-fold in the mutant cells when they were grown in the absence of hydroxyurea, compared to wild-type cells. These protein increases were accompanied by corresponding elevations in the levels of mRNAs for both subunits and increased rates of transcription of both genes. There was a 6-fold amplification in the gene copy number for protein M2.(ABSTRACT TRUNCATED AT 250 WORDS)

UI	MeSH Term	Description	Entries
D009154	Mutation	Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations.	Mutations
D009693	Nucleic Acid Hybridization	Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503)	Genomic Hybridization,Acid Hybridization, Nucleic,Acid Hybridizations, Nucleic,Genomic Hybridizations,Hybridization, Genomic,Hybridization, Nucleic Acid,Hybridizations, Genomic,Hybridizations, Nucleic Acid,Nucleic Acid Hybridizations
D002460	Cell Line	Established cell cultures that have the potential to propagate indefinitely.	Cell Lines,Line, Cell,Lines, Cell
D002467	Cell Nucleus	Within a eukaryotic cell, a membrane-limited body which contains chromosomes and one or more nucleoli (CELL NUCLEOLUS). The nuclear membrane consists of a double unit-type membrane which is perforated by a number of pores; the outermost membrane is continuous with the ENDOPLASMIC RETICULUM. A cell may contain more than one nucleus. (From Singleton & Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d ed)	Cell Nuclei,Nuclei, Cell,Nucleus, Cell
D004351	Drug Resistance	Diminished or failed response of an organism, disease or tissue to the intended effectiveness of a chemical or drug. It should be differentiated from DRUG TOLERANCE which is the progressive diminution of the susceptibility of a human or animal to the effects of a drug, as a result of continued administration.	Resistance, Drug
D004578	Electron Spin Resonance Spectroscopy	A technique applicable to the wide variety of substances which exhibit paramagnetism because of the magnetic moments of unpaired electrons. The spectra are useful for detection and identification, for determination of electron structure, for study of interactions between molecules, and for measurement of nuclear spins and moments. (From McGraw-Hill Encyclopedia of Science and Technology, 7th edition) Electron nuclear double resonance (ENDOR) spectroscopy is a variant of the technique which can give enhanced resolution. Electron spin resonance analysis can now be used in vivo, including imaging applications such as MAGNETIC RESONANCE IMAGING.	ENDOR,Electron Nuclear Double Resonance,Electron Paramagnetic Resonance,Paramagnetic Resonance,Electron Spin Resonance,Paramagnetic Resonance, Electron,Resonance, Electron Paramagnetic,Resonance, Electron Spin,Resonance, Paramagnetic
D005609	Free Radicals	Highly reactive molecules with an unsatisfied electron valence pair. Free radicals are produced in both normal and pathological processes. Free radicals include reactive oxygen and nitrogen species (RONS). They are proven or suspected agents of tissue damage in a wide variety of circumstances including radiation, damage from environment chemicals, and aging. Natural and pharmacological prevention of free radical damage is being actively investigated.	Free Radical
D006918	Hydroxyurea	An antineoplastic agent that inhibits DNA synthesis through the inhibition of ribonucleoside diphosphate reductase.	Hydroxycarbamid,Hydrea,Oncocarbide
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
D012264	Ribonucleotide Reductases		Ribonucleotide Reductase,Reductase, Ribonucleotide,Reductases, Ribonucleotide