Metabolism of Cottonseed Microsomal N-Acylphosphatidylethanolamine

N-acylphosphatidylethanolamine (NAPE) was recently shown to be synthesized in vitro in cottonseed microsomes by the direct N-acylation of phosphatidylethanolamine (PE) with unesterified fatty acids (K. D. Chapman and T. S. Moore, 1993, Plant Physiol. 102, 761-769), Here we examine the relationship of the synthesis and turnover of NAPE in cottonseed microsomes to the O-acylation of other membrane phospholipids. PE was N-acylated in a time-dependent manner with [1- 14C]palmitic acid independent of exogenously supplied ATP, O-Acylation of PE and phosphatidylcholine (PC) with [1- 14 C]palmitic acid proceeded only in the presence of ATP. Further radiolabeling experiments with [1- 14C]palmitoylCoA and phosphatidyl( N-[1- 14C]palmitoyl)ethanolamine indicated that O-acylation of phospholipids occurred via an acylCoA intermediate and not via an NAPE intermediate, [1- 14C]palmitic acid was released from PC[1- 14C-dipalmitoyl] in cottonseed microsomes in a Ca 2+-dependent manner and this [ 14C]-FFA was incorporated into [ 14C]NAPE in a linear fashion. Cottonseed NAPE was selectively hydrolyzed to N-acylethanolamine (NAE) and N-acyl lysophosphatidylethanolamine (NAlysoPE) by Ca 2+-independent, membrane-bound phospholipase D and A activities, respectively. NAlysoPE was not hydrolyzed to NAE, indicating that the phospholipase D that was active to ward NAPE did not recognize NAlysoPE; instead NAlysoPE was converted to NAPE in the presence of Ca 2+. Collectively, our results indicate that NAPE synthesis and the O-acylation of other phospholipids occur by two separate pathways and that microsomal NAPE is selectively turned over by membrane-bound phospholipase activities. A pathway for the metabolism of cottonseed NAPE is outlined.