Phosphatidylcholine Synthesis in Castor Bean Endosperm

Abstract

Endosperm halves from 3-day-old castor bean (Ricinus communis var Hale) were incubated for 30 minutes with L-14Cjserine, after which label was observed in ethanolamine, choline, phosphatidylserine, phosphatidylethanolamine, phosphatidylcholine, ethanolaminephosphate, and CDPethanolamine, but not in cholinephosphate or CDPcholine. Only later did significant amounts of isotope become incorporated into cholinephosphate and CDPcholine. The choline kinase inhibitor hemicholinium-3 prevented the incorporation of label from serine into cholinephosphate and CDPcholine, reduced the incorporation of I'4Cicholine into phosphatidylcholine by 65%, but inhibited the incorporation of label into phosphatidylcholine from serine by only 15%. The inhibitor did not prevent the incorporation of labeled methyl groups from S-adenosyl-Lmethionine into phosphatidyldimethylethanolamine plus phosphatidylcholine. The amount of incorporation of label from the methyl donor was only 8% of that from choline into phosphatidylcholine. The implications of these results for the pathway and regulation of phosphatidylcholine synthesis from the water-soluble precursors are discussed.Phosphatidylcholine is the major phospholipid in most eukaryotic cell membranes, with phosphatidylethanolamine generally being second most abundant (1). Some evidence exists for regulation of the levels of PC2 independent ofthe other phospholipids in plants. For example, PC increases as a proportion of the total phospholipid over the 8-d postgermination period in castor bean endosperm (13,16). This regulation may be complex, however, since PC synthesis involves more than one pathway (10, 12). For instance, in an early study of the incorporation of ['4C]serine into intact tomato roots, Willemot and Boll (18) concluded that PE was synthesized by the decarboxylation of PS, and PC by the methylation of PE. This pathway also has been found in castor bean endosperm (10), but the activity appeared to be only a fraction ofthe activity ofthe enzymes of the nucleotide pathway, in which PC is synthesized from free choline. Thus, at least two pathways must be considered as sources for PC in plants.The source of the water-soluble precursors also is unclear. It seems likely that in many plant tissues the biosynthesis ofcholine involves the methylation of ethanolamine, and that ethanolamine is formed by the decarboxylation of serine (6). It is not firmly established, however, whether these reactions occur predominantly at the level of free bases, phosphoryl bases, or phosphatides. In other words, it is not clear if the syntheses involve 'Supported by National Science Foundation grant PCM-8402001 to T. S. M.2Abbreviations: PC, phosphatidylcholine; PE, phosphatidylethanolamine; PS, phosphatidylserine; SAM, S-adenosyl-L-methionine.only the water soluble substrates or a complex interrelationship between cytosolic and membrane-bound intermediates. Studies in some plant tissues, utilizing both precursor incorporation (8,14) and enzyme studies (9, 10), appear to indicate that the m...

Amino Acid Metabolism of Lemna minor L.

“…The ratios of glycine:serine and glutamate:aspartate increased dramatically (>10-fold) in response to AOA (Table II). These responses appear consistent with AOA inhibition of glycine decarboxylase (30,37) and glutamate:oxaloacetate aminotransferase (9 (4,18) and aspartate conversion to asparagine via asparaginesynthetase [20,23]). …”

Mitochondrial Phosphatidylserine Decarboxylase from Higher Plants. Functional Complementation in Yeast, Localization in Plants, and Overexpression in Arabidopsis

“…These results could therefore be due to artifactual decarboxylation of the type that occurs in soluble plant extracts with Ser and other amino acids (Birecka et al, 1985;Rontein et al, 2001). The results of radiotracer kinetic studies with plant tissues fed [U-14 C]Ser are consistent with the presence of PSD activity but do not unambiguously establish its existence (Marshall and Kates, 1974;Kinney and Moore, 1987a;Mudd and Datko, 1989).Plant membranes are known to contain small amounts of Ptd-Ser (Mudd, 1980), and two different plant enzymes that synthesize Ptd-Ser have been identified. Ptd-Ser synthase has been cloned from wheat (Triticum aestivum; Delhaize et al, 1999) providing strong evidence that the yeast-type pathway (Carman and Zeimetz, 1996) for Ptd-Ser synthesis exists in plant tissues.…”

“…These results could therefore be due to artifactual decarboxylation of the type that occurs in soluble plant extracts with Ser and other amino acids (Birecka et al, 1985;Rontein et al, 2001). The results of radiotracer kinetic studies with plant tissues fed [U-14 C]Ser are consistent with the presence of PSD activity but do not unambiguously establish its existence (Marshall and Kates, 1974;Kinney and Moore, 1987a;Mudd and Datko, 1989).…”

Radiotracer and Computer Modeling Evidence that Phospho-Base Methylation Is the Main Route of Choline Synthesis in Tobacco

“…It is therefore interesting that modeling of the [ 14 C]formate data indicated that the EA moiety of Ptd-EA came from a P-EA pool of low specific activity and from another source, also of low specific activity. This other source could be Ptd-Ser decarboxylation or a base exchange reaction between free EA and phospholipids (Kinney and Moore, 1987;Mudd and Datko, 1989c). Consistent with these possibilities, preliminary analyses showed that 14 C was present in products with the chromatographic properties of Ptd-Ser and free EA.…”