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Substrate preference of stress‐activated phospholipase D in Chlamydomonas and its contribution to PA formation
Author(s) -
Arisz Steven A.,
Valianpour Fredoen,
Van Gennip Albert H.,
Munnik Teun
Publication year - 2003
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.2003.01750.x
Subject(s) - phosphatidic acid , phospholipase d , diacylglycerol kinase , phosphatidylethanolamine , chemistry , substrate (aquarium) , phospholipase , biochemistry , phospholipase c , fatty acid , second messenger system , osmotic shock , phospholipid , biophysics , intracellular , enzyme , phosphatidylcholine , biology , membrane , protein kinase c , ecology , gene
Summary In response to various environmental stress conditions, plants rapidly form the intracellular lipid second messenger phosphatidic acid (PA). It can be generated by two independent signalling pathways via phospholipase D (PLD) and via phospholipase C (PLC) in combination with diacylglycerol kinase (DGK). In the green alga Chlamydomonas , the phospholipid substrates for these pathways are characterized by specific fatty acid compositions. This allowed us to establish: (i) PLD's in vivo substrate preference; and (ii) PLD's contribution to PA formation during stress signalling. Accordingly, G‐protein activation (1 µ m mastoparan), hyperosmotic stress (150 m m NaCl) and membrane depolarization (50 m m KCl) were used to stimulate PLD, as monitored by the accumulation in 5 min of its unique transphosphatidylation product phosphatidylbutanol (PBut). In each case, PBut's fatty acid composition specifically matched that of phosphatidylethanolamine (PE), identifying this lipid as PLD's favoured substrate. This conclusion was substantiated by analysing the molecular species by electrospray ionization‐mass spectrometry (ESI‐MS/MS), which revealed that PE and NaCl‐induced PBut share a unique (18 : 1) 2 ‐structure. The fatty acid composition of PA was much more complex, reflecting the different contributions from the PLC/DGK and PLD pathways. During KCl‐induced stress, the PA rise was largely accounted for by PLD activity. In contrast, PLD's contribution to hyperosmotic stress‐induced PA was less, being approximately 63% of the total increase. This was because the PLC/DGK pathway was activated as well, resulting in phosphoinositide‐specific fatty acids and molecular species in PA.