Effects of TPA on mitosis of V79 Chinese hamster cells. Reduced precision of chromosome distribution and modification of entrance into and exit out of mitosis.

Abstract

Protein kinase C (PKC) was activated in V79 Chinese hamster cells with 10 nM 12-O-tetradecanoylphorbol-13-acetate (TPA). Within 30 min soluble activity decreased concomitant with a 10-fold increase of particulate activity. The latter was still elevated after 3 h but was back to control levels after 24 h of treatment; by then soluble activity was lost. The frequency of mitotic cells with signs of abnormal spindle function increased within 15 min and reached a plateau after 45-60 min which lasted throughout the 4 h treatment. The c-mitotic effect was delayed and significantly lower when 10 nM TPA was combined with 50 microM of the protein kinase inhibitor H7. The frequency of disturbed mitotic cells decreased after 24 h of treatment but remained significantly higher than in non-treated cells. Change of medium and addition of new TPA caused a slight but significant further increase. It is suggested that PKC takes part in eliciting the c-mitotic effect of TPA. However, the sustained effect coincident with down-regulation points to significant alterations of the level or the activity of an as yet unidentified ultimate elicitor. TPA also caused a transient block in the G2 phase which was ameliorated by H7 and which could not be detected at all in TPA-pretreated cells (24 h) given new TPA. This suggests that PKC takes part in eliciting the G2/M block as well but the mechanism is different from the one(s) behind the c-mitotic effect. V79 cells were found to exit from mitosis in the presence of 0.2 microgram/ml of colcemid but TPA-pretreated cells showed a decreased exit rate. There was no sign of hampered exit among cells going into mitosis soon after the G2 block was reversed, which implies that the spontaneous reversal of the block does not involve rapid down-regulation of PKC.