Cell cycle regulation of an exogenous human poly(ADP-ribose) polymerase cDNA introduced into murine cells
Department of Cell Biology
Animals; Blotting, Northern; Cell Cycle; Cell Division; Cell Nucleus; DNA; *Gene Expression Regulation, Enzymologic; Hela Cells; Humans; Interphase; Mice; Plasmids; Poly(ADP-ribose) Polymerases; RNA; RNA, Messenger; Thymidine; *Transcription, Genetic; Transfection
We have evaluated the regulation of expression of the poly(ADP-ribose) polymerase gene during cell growth and replication. In a synchronized population of HeLa cells or in serum-stimulated WI-38 cells, steady-state levels of the polymerase mRNA were highest at late S and S-G2 phases and negligible in early S phase. Transcription did not solely account for the significant increase in the mRNA levels observed in late S phase by Northern analysis. The stability of the mRNA was dependent upon the percent proliferating cells in the culture. Accordingly, polymerase mRNA from cells in early exponential phase was significantly more stable than from cells in stationary phase of asynchronous growth. To clarify these observations, we utilized a novel heterologous expression system that involved murine 3T3 cells transfected with a human poly(ADP-ribose) polymerase cDNA under the control of a non-cell cycle-specific promoter. Cells were synchronized, and a comparison was made of the endogenous (murine) and exogenous (human) polymerase mRNA levels. Both the endogenous and the exogenous mRNA were specifically stabilized by the same mechanisms and only during late S phase; therefore, we concluded that mRNA pools for the polymerase are regulated at the post-transcriptional level. The heterologous expression system confirmed that the post-transcriptional regulation system in the mouse cells can recognize and faithfully regulate the human cDNA in response to the murine cell cycle signals. More importantly, the presence of extra copies (human) of the polymerase gene did not provide an increased amount of the total polymerase mRNA or protein and, in fact, the sum of the endogenous and exogenous mRNA in the transfected cells was approximately the same as the level of endogenous transcript in the control cells. This suggested that there might be a limit to the amount of polymerase protein accumulating in the cellular pool and thus levels of poly(ADP-ribose) polymerase may be autoregulated.
DOI of Published Version
J Cell Physiol. 1990 Aug;144(2):345-53. Link to article on publisher's site
Journal of cellular physiology
Bhatia K, Kang VH, Stein GS, Bustin M, Cherney BW, Notario V, Haque SJ, Huppi K, Smulson ME. (1990). Cell cycle regulation of an exogenous human poly(ADP-ribose) polymerase cDNA introduced into murine cells. Stein, Stein, Lian, vanWijnen Lab Publications. https://doi.org/10.1002/jcp.1041440221. Retrieved from https://escholarship.umassmed.edu/stein/123