Targeted neutralization of calmodulin in the nucleus blocks DNA synthesis and cell cycle progression

Calmodulin (CaM) is a major intracellular calcium binding protein which has been implicated in the regulation of cell proliferation. Previous studies using chemically synthesized CaM antagonists and anti-sense RNA indicated that CaM is important for initiation of DNA synthesis and cell cycle progression. However, these methods reduce total intracellular CaM and globally interfering with all the CaM-dependent processes. In order to explore the function of nuclear CaM during the cell cycle, a CaM inhibitor peptide was targeted to the nucleus of intact mammalian cells. Cell progression through S-phase was assessed by incorporation of the thymidine analogue, BrdU. Cells were transfected for 48 h with either the CaM inhibitor peptide gene or the control plasmid prior to analysis. Approx. 70% of the control cells incorporated BrdU. In striking contrast, double immunofluorescent labeling demonstrated that none of the cells expressing the CaM inhibitor peptide entered S-phase. This result indicates that neutralization of nuclear CaM by targeted expression of a CaM inhibitor peptide blocks DNA synthesis and cell cycle progression.