Fluctuations and nuclear translocation of p53, c-MYC, CDC-2 and PCNA proteins during growth and commitment to differentiation of AGF cells

Transcription regulatory proteins such as c-myc and p53 play an important role in the regulation of cell growth, differentiation and apoptosis. Similarly, cell cycle regulatory proteins such as CDC-2, cyclin A and the proliferation cell nuclear antigen (PCNA), play an important role in regulation of cell growth. Yet, there are contradictory reports as for the exact mechanism by which these proteins affect antagonistic processes like growth, differentiation and apoptosis. In the present study, we report, for the first time a detailed analysis of the steady- state level and the nuclear/cytoplasmic distribution of oncoproteins and cell cycle regulatory proteins, in the early phase of PMA induced differentiation of AGF cells. Particular emphasis was put on the first 6 h of commitment to differentiation as well as on the committed/differentiated cells (24 h post induction). Using Western blots of protein extracted from the cytosolic, membranal and nuclear fraction we document an early intranuclear influx of p53 with concomitant extranuclear efflux of c-myc as early as 1-6 h post induction. Similarly, intranuclear sequestration of CDC-2 occurs throughout the commitment phase. General down regulation of p53 and partial down regulation of CDC-2, PCNA and c-myc occur in committed/differentiated cells (24 h post induction). The results obtained by Western blots were further supported by immunolocalization using confocal microscopy.