Effect of antisense oligodeoxynucleotides directed to individual calmodulin gene transcripts on the proliferation and differentiation of PC12 cells.

Calmodulin (CaM) is encoded by three different genes that collectively give rise to five transcripts. In the present study, we used antisense oligodeoxynucleotides targeted to unique sequences in the transcripts from the individual CaM genes to selectively block the expression of the different genes and to investigate the roles these individual genes play in the proliferation and nerve growth factor (NGF)-induced differentiation of PC12 cells. Culturing PC12 cells in the presence of oligodeoxynucleotide antisense to the transcripts from CaM genes I and II caused a significant decrease in the proliferation and a significant delay in the NGF-induced differentiation of PC12 cells when compared with untreated cells and with cells treated with the corresponding randomized oligodeoxynucleotides. However, an oligodeoxynucleotide antisense to CaM gene III did not significantly alter the proliferation or the NGF-induced differentiation of PC12 cells. The inhibition of cell proliferation could be reversed by washing out the antisense oligodeoxynucleotides. The levels of CaM in cells treated with oligodeoxynucleotides antisense to CaM genes I or II were reduced 52% or 63%, respectively, of the levels found in the control cells. However, the levels of CaM were not significantly reduced in PC12 cells treated with CaM gene III antisense oligodeoxynucleotide. None of the randomized oligodeoxynucleotides had any effect on the levels of CaM in PC12 cells. The reduced levels of CaM in PC12 cells treated with an oligodeoxynucleotide antisense to CaM gene I were accompanied by a reduction in the levels of the CaM gene I mRNAs, supporting a true antisense mechanism of action for these oligodeoxynucleotides. These results suggest that altering the level of CaM by using antisense oligodeoxynucleotides targeted to the dominant CaM transcripts in a particular cell type will specifically inhibit their proliferation and, in the case of neuronal cells, alter the course of their differentiation.