No abstract

No abstract

SummaryPlant heterotrimeric G-proteins are involved in a variety of signaling pathways, though only one a and a few bc isoforms of their subunits exist. In isolated plasma membranes of California poppy (Eschscholzia californica), the plant-specific Ga subunit was isolated and identified immunologically and by homology of the cloned gene with that of several plants. In the same membrane, phospholipase A 2 (PLA 2 ) was activated by yeast elicitor only if GTPcS (an activator of Ga) was present. From the cholate-solubilized membrane proteins, PLA 2 was co-precipitated together with Ga by a polyclonal antiserum raised against the recombinant Ga. In this immunoprecipitate and in the plasma membrane (but not in the Ga-free supernatant) PLA 2 was stimulated by GTPcS. Plasma membranes and immunoprecipitates obtained from antisense transformants with a low Ga content allowed no such stimulation. An antiserum raised against the C-terminus (which in animal Gas is located near the target coupling site) precipitated Ga without any PLA 2 activity. Using non-denaturing PAGE, complexes of solubilized plasma membrane proteins were visualized that contained Ga plus PLA 2 activity and dissociated at pH 9.5. At this pH, PLA 2 was no longer stimulated by GTPcS. It is concluded that a distinct fraction of the plasma membrane-bound PLA 2 exists in a detergent-resistant complex with Ga that can be dissociated at pH 9.5. This complex allows the Ga-mediated activation of PLA 2 .

In cultured cells of California poppy (Eschscholzia californica), lysophosphatidylcholine (LPC) triggers a signal path that finally induces alkaloid biosynthesis. LPC is transiently generated by elicitor-activated phospholipase A(2) of the plasma membrane. Externally added LPC is rapidly acylated by a membrane-bound enzyme that shows the highest specific activity in the purified plasma membrane. The fatty acid incorporated into the sn-2 position of LPC is preferentially linoleic (18:2), which is the most abundant acyl component in the PC species of Eschscholzia cells, but a minor component of the pool of free fatty acids. The fatty acid at the sn-1 position of LPC is less important for substrate specificity. The capacity of LPC acylation by intact cells or isolated plasma membranes by far exceeds the rate of LPC generation by activated phospholipase A(2) and is not limited by the availability of acyl donors. Metabolites other than phosphatidylcholine (PC) were not significantly produced from labeled LPC within 20 min, indicating that lysophospholipases are not significantly contributing to the short-time metabolism of LPC. It is concluded that reacylation to PC is the dominating process in the detoxication of LPC and ensures the transient character of its steady state concentrations, even at maximum phospholipase A(2) activities.