DNA precursor compartmentation in mammalian cells: metabolic and antimetabolic studies of nuclear and mitochondrial DNA synthesis
HeLa cells were used for the quantitation of cellular and mitochondrial deoxyribonucleoside triphosphate (dNTP) and ribonucleoside triphosphate (rNTP) pools and of changes in pools in response to treatment with the antimetabolites methotrexate (mtx) and 5-fluorodeoxyuridine (FUdR). Use of an enzymatic assay of dNTPs and of improved nucleotide extraction methods allowed quantitation of mitochondrial dNTP pools. All four mitochondrial dNTP pools expand following treatment with mtx or FUdR whereas cellular dTTP and dGTP pools are depleted. Mitochrondrial rNTP pools were also found to expand in response to these antimetabolites. Mouse L-cells were used to determine the relative contributions of an exogenously supplied precursor to nuclear and mitochrondrial DNA replication. Cells were labeled to near steady state specific activities with /sup 32/P-orthophosphate and subsequently labeled with (/sup 3/H)uridine, a general pyrimidine precursor, in the continuing presence of /sup 32/P. Deoxyribonucleoside monophosphates derived from these DNAs were separated by HPLC and the /sup 3/H//sup 32/P ratio in each pyrimidine determined. The dCMP residues in mitochondrial DNA (mtDNA) were found to be derived exclusively from the exogenous supplied uridine. The dTMP residues from nuclear and mtDNA and the dCMP residues from nuclear DNA were seen to be synthesized partly from exogenous sources and partly from other sources, presumably de novo pyrimidine synthesis.
- Research Organization:
- Oregon State Univ., Corvallis (USA)
- OSTI ID:
- 5741974
- Resource Relation:
- Other Information: Thesis (Ph. D.)
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
HELA CELLS
DNA REPLICATION
L CELLS
BIOASSAY
ENZYMES
EXPERIMENTAL DATA
FUDR
LABELLED COMPOUNDS
LIQUID COLUMN CHROMATOGRAPHY
METHOTREXATE
MICE
MITOCHONDRIA
PHOSPHORUS 32
PYRIMIDINES
TRACER TECHNIQUES
TRITIUM COMPOUNDS
URIDINE
ANIMALS
ANTI-INFECTIVE AGENTS
ANTIMETABOLITES
ANTIMICROBIAL AGENTS
AZINES
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CELL CONSTITUENTS
CHROMATOGRAPHY
DATA
DAYS LIVING RADIOISOTOPES
DRUGS
FLUOROURACILS
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
INFORMATION
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MAMMALS
NUCLEI
NUCLEIC ACID REPLICATION
NUCLEOSIDES
NUCLEOTIDES
NUMERICAL DATA
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC FLUORINE COMPOUNDS
ORGANIC HALOGEN COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANOIDS
PHOSPHORUS ISOTOPES
RADIOISOTOPES
RADIOSENSITIZERS
RIBOSIDES
RODENTS
SEPARATION PROCESSES
URACILS
VERTEBRATES
550501* - Metabolism- Tracer Techniques
560301 - Chemicals Metabolism & Toxicology- Cells- (-1987)