Lactate dehydrogenase A subunit messenger RNA stability is synergistically regulated via the protein kinase A and C signal transduction pathways.
We have identified and studied a posttranscriptional mechanism of lactate dehydrogenase A (LDH) subunit gene expression at the level of mRNA stability. Using the well differentiated rat C6 glioma cell line as a model system, the effects of activators of the protein kinase A and C pathways on the half-life of LDH A mRNA were measured by two independent methods: 1) by the RNA synthesis inhibitor-chase method using actinomycin D, and 2) by analysis of decay of LDH A [3H]mRNA in [3H]uridine-labeled cells. By each method, the half-life of relatively short-lived LDH A mRNA was increased 5- to 7-fold in 8- (4-chloro-phenylthio) cAMP or forskolin-treated and about 3-fold in 12-0-tetradecanoylphorbol-13- acetate (TPA) or dioctanoylglycerol-treated cells. Forskolin acted synergistically with TPA to prolong LDH A mRNA half-life from 55 min to more than 20 h. The relatively rapid basal decay rate of LDH A mRNA was also considerably slowed in the presence of the protein phosphatase inhibitor okadaic acid, suggesting a functional role for protein phosphorylation in the stabilization process. In glioma cells stably transformed with a protein kinase A catalytic subunit expression vector, overexpression of the catalytic subunit stabilized LDH mRNA to the degree seen in forskolin-treated cells. In cells transfected with a protein kinase A inhibitor-expression vector, cAMP-mediated stabilization of LDH A mRNA half-life was prevented. Furthermore, both staurosporin and 3- [1-(3-dimethylaminopropyl)-indol-3-yl]-3-(indol- 3-yl)- maleimide, inhibitors of protein kinase C, prevented the TPA-induced stabilization of LDH A mRNA. We conclude from the experimental data that the protein kinase A and C signal pathways play an active functional role in regulating LDH A mRNA stability and act cooperatively to achieve LDH A mRNA stability regulation.
D000818:Animals; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase / Q000235:genetics*; D019319:Okadaic Acid; D010703:Phorbol Esters / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D011493:Protein Kinase C / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D014158:Transcription, Genetic
D004791:Enzyme Inhibitors; D004988:Ethers, Cyclic; D007527:Isoenzymes; D010703:Phorbol Esters; D012333:RNA, Messenger; D005576:Colforsin; D019319:Okadaic Acid; D007770:L-Lactate Dehydrogenase; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases