Differential expression and stability of poly(ADP-ribose)polymerase mRNA in human cells.
The regulation of the expression of the poly(ADP-ribose)polymerase gene was studied in HeLa cells and in quiescent and mitogen-stimulated human lymphocytes by quantitating the mRNA molecules with a new technique based on the polymerase chain reaction. Using plasmid constructs containing defined sequences of the poly(ADP-ribose)polymerase cDNA as internal standards in a competitive PCR reaction, precise measurements of reverse transcribed mRNA copies per microgram of total RNA were obtained. The value found for asynchronously growing HeLa cells (8.6 x 10(5) copies) was very close to that observed for proliferating lymphocytes (8.7 x 10(5)) whereas a 20-fold lower value (0.4 x 10(5)) was obtained for quiescent lymphocytes. The determination of the stability of the mRNA of the enzyme in G0 and stimulated lymphocytes, and in HeLa cells was performed by devising a new PCR amplification system, using non-competitive conditions and plasmid target sequences as internal standards. The half-life of mRNA for poly(ADP-ribose)polymerase was approx. 1 h in G0 lymphocytes and 4-5 h in stimulated lymphocytes and in HeLa cells. This observed difference in stability of the transcripts can partially account for the observed difference in mRNA levels between G0 and stimulated human lymphocytes.