Characterization of Inositol Phosphates in Carrot (Daucus carota L.) Cells 1

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We have shown previously that inositol-1,4,5-trisphosphate (IP3) stimulates an efflux of 45Ca2+ from fusogenic carrot protoplasts (M Rincón, WF Boss [1987] Plant Physiol 83: 395-398). In light of these results, we suggested that IP3 might serve as a second messenger for the mobilization of intracellular Ca2+ in higher plant cells. To determine whether or not IP3 and other inositol phosphates were present in the carrot cells, the cells were labeled with myo-[2-3H]inositol for 18 hours and extracted with ice-cold 10% trichloroacetic acid. The inositol metabolites were separated by anion exchange chromatography and by paper electrophoresis. We found that [3H]inositol metabolites coeluted with inositol bisphosphate (IP2) and IP3 when separated by anion exchange chromatography. However, we could not detect IP2 or IP3 when the inositol metabolites were analyzed by paper electrophoresis even though the polyphosphoinositides, which are the source of IP2 and IP3, were present in these cells. Thus, [3H] inositol metabolites other than IP2 and IP3 had coeluted on the anion exchange columns. The data indicate that either IP3 is rapidly metabolized or that it is not present at a detectable level in the carrot cells.

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