Turnover of inositol phospholipids in cultured murine keratinocytes: Possible involvement of inositol triphosphate in cellular differentiation

The relationship between the turnover of inositol phospholipids (Ptdlns) and the growth and differentiation of normal murine keratinocytes in culture was studied. Addition of myo-[U-¹⁴C] inositol to freshly plated cells resulted in a linear incorporation of radiolabel into the inositol phospholipids of the proliferating basal cells in culture during the initial 36 h, after which time the rate of radiolabel incorporation into the cells declined. The decrease in the incorporation of the radiolabel into the Ptdlns, particularly the more highly phosphorylated Ptdlns-4P and PtdIns4,5P₂, correlated with the marked hydrolysis of these polyphosphoinositides and the rapid hydrolytic release of the inositol phosphates (InsP₂ and InsP₃). The transient accumulation of the InsPs correlated with the onset of differentiation of these cells. To ascertain whether the above observations of keratinocytes that were undergoing normal proliferative and differentiating phases in culture are consistent with the more synchronized populations of proliferative and differentiating cells, we investigated the turnover of Ptdlns in a Ca²⁺-regulated system of homogenous populations of proliferating mouse keratinocytes in 0.09 mM Ca²⁺, and a differentiating population in 1.8 mM Ca²⁺. Our data from system revealed rapid hydrolysis of the Ptdlns in the prelabeled low-Ca²⁺ proliferating cells immediately after a switch from the low to normal extracellular Ca²⁺ medium. Associated with this hydrolysis was the rapid and transient accumulation of the InsPs (maximum of 60 sec). The hydrolysis of the Ptdlns and the accumulation of the InsP₃ were not observed when the prelabeled proliferating cells were switched from a low to a low extracellular Ca²⁺ medium. These results suggest that the rapid hydrolysis of the Ptdlns, particularly PtdIns4,5P₂, which was accompanied by the hydrolytic release of InsP₃, could be the initiating signal to program proliferating keratinocytes into differentiation.