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During the early hours after exposure to differentiation inducing agents, Friend erythroleukaemia cells undergo alterations which commit them to cessation of growth and development of the characteristics of differentiation. Our current experiments have compared the expression and activity of phosphoinositide 3-kinase (PI 3-kinase) in control cells with cells undergoing differentiation which has been induced by dimethyl sulfoxide (DMSO). When the cultures were initiated with stationary phase cells and DMSO was added at the time of seeding, PI 3-kinase activity was stimulated in both treated and control cells during the first 3 h from seeding. This event appears to be a rate limiting step in commitment since pretreatment of cells with 10 microM LY294002 or down-regulation of p85 expression prior to adding DMSO completely prevents commitment to erythropoiesis. Accordingly, PI 3-kinase inhibition during the commitment period prevents DNA-binding of the transcription factor GATA-1, essential for erythroid differentiation. However, once cells are committed to differentiate, PI 3-kinase activity and expression dramatically decreases along with the differentiation programme, to become barely detectable after 96 h. Remarkably, LY294002 treatment leads to accumulation of cell in G1 phase and prevents DMSO-dependent cyclin D3 induction. Based on these data, we suggest that PI 3-kinase is rate limiting for the completion of the first round cycle of cell division required for initiation of erythrocytic differentiation. On the other hand, the late decrease of PI 3-kinase associated with the differentiation process seems to be part of the programmed shut off of genes not needed in mature erythrocytes.

Inhibition of phosphoinositide 3-kinase impairs pre-commitment cell cycle traverse and prevents differentiation in erythroleukaemia cells