Transmembrane signaling and phosphoinositides

This chapter illustrates that inositol phospholipid metabolism can be monitored in plants but it must be done carefully to be able to interpret the results. 32Pi and myo[2-3H]inositol are the most common radioisotopes used to label the inositol phospholipids. 32Pi labels phospholipids, sugar phosphates, nucleotides, and, other cellular metabolites that can interfere with the analysis of the inositol phospholipids and inositol phosphates. myo[2-3H]inositol is a more selective means of labeling the inositol phospholipids and inositol phosphates in vivo than 32Pi. 32P incorporation into phosphatidylinositol bisphosphate (PIP2) saturates after approximately 10 min with the carrot cells. The major drawback to using short-term labeling is that the stimulus applied may affect [32P] Pi uptake or the levels of [32P]ATP within the cell and thereby affect the recovery of [32P]PIP or PIP2 without affecting phospholipid turnover. In addition, differences in the uptake of the label among tissue samples prior to applying the stimulus are more pronounced with short-term labeling. © 1995, Academic Press Inc.