Reversible G1 arrest of a human lung epithelial cell line by staurosporine

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Abstract
Staurosporine, a microbial‐derived protein kinase inhibitor, reversibly blocked non‐synchronized, replicating cultures of the human lung epithelial cell line EKVX in the G1 phase of cell cycle and inhibited DNA synthesis and cell replication. The mechanism of this cell‐cycle arrest in EKVX cells by staurosporine was likely due to inhibition of protein kinase C (PKC) because: (1) dose‐dependent inhibition of DNA synthesis occurred at levels of staurosporine that inhibit phosphorylation of PKC substrate, (2) inhibition of DNA synthesis was also seen after treatment with another PKC inhibitor H7, but not by the chemically similar HA1004, which has a relative inhibitory specificity for cAMP‐dependent protein kinase, and (3) the DNA synthesis was not inhibited by specific tyrosine kinase inhibitors Genistein and Lavendustin A at concentrations that inhibit tyrosine kinase activity. Removal of staurosporine from cell culture media resulted in a rebound in PKC activity and synchronized DNA synthesis in EKVX cultures. The reversibility of the inhibition was noted even after 5 days of treatment with staurosporine, and DNA synthesis remained synchronized for at least two rounds of cell replication after removal of staurosporine. Flow cytometric analysis confirmed that more than 90% of the cell population was blocked in the G1 phase after cells were treated with staurosporine for 24 h. Agents such as staurosporine may be useful for synchronizing cell populations to study cell‐cycle specific biochemical events important for the regulation of cell replication in the EKVX cell line.

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