No abstract

NAPE is a widespread, albeit minor, membrane phospholipid in animal and plant tissues (Schmid et al., 1990; Chapman and Moore, 1993). Its unusual structural features (a third fatty acid moiety linked to the amino head group of PE) impart stabilizing properties to membrane bilayers (Domingo et al., 1994; LaFrance et al., 1997). NAPE and its hydrolysis products, NAEs, are known to accumulate in vertebrate tissues under pathological conditions (for review, see Schmid et al., 1990). Recently, there has been renewed interest in NAEs because of the contention that anandamide (N-arachidonylethanolamine) is an endogenous ligand for the cannabinoid receptor in mammalian brain (Devane et al., 1992; Fontana et al., 1995;Schmid et al., 1996). The likely route for NAE formation in neural and nonneural tissues, although the matter of some debate, is via the signal-mediated hydrolysis of NAPE (DiMarzo et al., 1994;Schmid et al., 1996;Sugiura, et al., 1996).In plants little is known regarding the catabolism of NAPE. In cottonseed microsomes NAPE was metabolized to NAE or NAlysoPE by PLD-or PLA-type activities, respectively (Chapman et al., 1995b). However, the metabolic fate of NAPE in vivo and the factors that regulate NAPE hydrolysis remain largely unknown. We previously noted that the biosynthesis of NAPE was increased in elicitortreated cell suspensions of tobacco (Nicotiana tabacum L.). Here we extend our investigations with this model system to examine NAPE catabolism by plant cells in vivo. NAE was released from NAPE, and it accumulated extracellularly. We identified by GC-MS these tobacco NAEs as N-lauroylethanolamine and N-myristoylethanolamine. These NAEs were increased in elicitor-treated cell suspensions. Furthermore, we detected the enzymatic machinery capable of the release and the degradation of NAEs in tobacco cells. To our knowledge this represents the first identification of the NAE molecular species in plant cells. It is tempting to speculate that NAPE hydrolysis in elicitortreated plant cells may be involved in a signaling pathway analogous to that found in mammalian cells. MATERIALS AND METHODS Cell Cultures, Elicitor Treatment, and Lipid ExtractionsTobacco (Nicotiana tabacum L. cv KY-14) cell suspensions were subcultured every 7 d (Chapman et al., 1995a); cell suspensions in log phase were treated with elicitor (xylanase, 1 g/mL) as previously described (Chapman et al., 1995a). Control and experimental treatments were carried out on aliquots of the same population of cells. Culture supernatants were separated from cells by filtration. Cells were quick frozen in liquid N 2 , powdered in a mortar, and

While cannabinoids are secondary metabolites synthesized by just a few plant species, N-acylethanolamines (NAEs) are distributed widely in the plant kingdom, and are recovered in measurable, bioactive quantities in many plant-derived products. NAEs in higher plants are ethanolamides of fatty acids with acyl-chain lenghts of C12-C(18) and zero to three C=C bonds. Generally, the most-abundant NAEs found in plants and vertebrates are similar, including NAE 16 : 0, 18 : 1, 18 : 2, and 18 : 3. Like in animal systems, NAEs are formed in plants from N-acylphosphatidylethanolamines (NAPEs), and they are hydrolyzed by an amidase to yield ethanolamine and free fatty acids (FFA). Recently, a homologue of the mammalian fatty acid amide hydrolase (FAAH-1) was identified in Arabidopsis thaliana and several other plant species. Overexpression of Arabidopsis FAAH (AtFAAH) resulted in plants that grew faster, but were more sensitive to biotic and abiotic insults, suggesting that the metabolism of NAEs in plants resides at the balance between growth and responses to environmental stresses. Similar to animal systems, exogenously applied NAEs have potent and varied effects on plant cells. Recent pharmacological approaches combined with molecular-genetic experiments revealed that NAEs may act in certain plant tissues via specific membrane-associated proteins or by interacting with phospholipase D-alpha, although other, direct targets for NAE action in plants are likely to be discovered. Polyunsaturated NAEs can be oxidized via the lipoxygenase pathway in plants, producing an array of oxylipin products that have received little attention so far. Overall, the conservation of NAE occurrence and metabolic machinery in plants, coupled with the profound physiological effects of elevating NAE content or perturbing endogenous NAE metabolism, suggest that an NAE-mediated regulatory pathway, sharing similarities with the mammalian endocannabinoid pathway, indeed exists.

While cannabinoids are secondary metabolites synthesized by just a few plant species, Nacylethanolamines (NAEs) are distributed widely in the plant kingdom, and are recovered in measurable, bioactive quantities in many plant-derived products. NAEs in higher plants are ethanolamides of fatty acids with acyl-chain lenghts of C 12 ÀC 18 and zero to three C¼C bonds. Generally, the most-abundant NAEs found in plants and vertebrates are similar, including NAE 16 : 0, 18 : 1, 18 : 2, and 18 : 3. Like in animal systems, NAEs are formed in plants from N-acylphosphatidylethanolamines (NAPEs), and they are hydrolyzed by an amidase to yield ethanolamine and free fatty acids (FFA). Recently, a homologue of the mammalian fatty acid amide hydrolase (FAAH-1) was identified in Arabidopsis thaliana and several other plant species. Overexpression of Arabidopsis FAAH (AtFAAH) resulted in plants that grew faster, but were more sensitive to biotic and abiotic insults, suggesting that the metabolism of NAEs in plants resides at the balance between growth and responses to environmental stresses. Similar to animal systems, exogenously applied NAEs have potent and varied effects on plant cells. Recent pharmacological approaches combined with molecular-genetic experiments revealed that NAEs may act in certain plant tissues via specific membrane-associated proteins or by interacting with phospholipase D-a, although other, direct targets for NAE action in plants are likely to be discovered. Polyunsaturated NAEs can be oxidized via the lipoxygenase pathway in plants, producing an array of oxylipin products that have received little attention so far. Overall, the conservation of NAE occurrence and metabolic machinery in plants, coupled with the profound physiological effects of elevating NAE content or perturbing endogenous NAE metabolism, suggest that an NAE-mediated regulatory pathway, sharing similarities with the mammalian endocannabinoid pathway, indeed exists.