A Novel Phosphatidylcholine-hydrolyzing Phospholipase C Induced by Phosphate Starvation in Arabidopsis

Abstract

During phosphate starvation, it is known that phospholipids are degraded, and conversely, a nonphosphorus galactolipid digalactosyldiacylglycerol accumulates in the root plasma membrane of plants. We report a novel phospholipase C that hydrolyzes phosphatidylcholine and is greatly induced in response to phosphate deprivation in Arabidopsis. Since phosphatidylcholinehydrolyzing activity by phospholipase C was highly upregulated in phosphate-deprived plants, gene expression of some phospholipase C was expected to be induced during phosphate starvation. Based on amino acid sequence similarity to a bacterial phosphatidylcholine-hydrolyzing phospholipase C, six putative phospholipase Cs were identified in the Arabidopsis genome, one of which, NPC4, showed significant transcriptional activation upon phosphate limitation. Molecular cloning and functional expression of NPC4 confirmed that the NPC4 gene encoded a functional phosphatidylcholinehydrolyzing phospholipase C that did not require Ca 2؉ for its activity. Subcellular localization analysis showed that NPC4 protein was highly enriched in the plasma membrane. Analyses of transferred DNA-tagged npc4 mutants revealed that disruption of NPC4 severely reduces the phosphatidylcholine-hydrolyzing phospholipase C activity in response to phosphate starvation. These results suggest that NPC4 plays an important role in the supply of both inorganic phosphate and diacylglycerol from membrane-localized phospholipids that would be used for phosphate supplementation and the replacement of polar lipids in the root plasma membrane during phosphate deprivation.Phosphorus is an essential element for plant growth, development, and reproduction. It plays decisive roles not only in regulation of various enzymes but also in constitutive components such as membrane phospholipids and nucleic acids. In most soils, despite its abundance, phosphorus is not freely available for assimilation by roots (1). Therefore, plants have developed distinct systems to cope with phosphate deficiency.When plants suffer from phosphate limitation, highly integrated systems are activated both for assimilation of P i and supplementation of P i from innermost phosphorus storage. The former action is represented by a morphological modification of root architecture upon P i starvation, which presumably facilitates P i uptake by enlargement of absorptive root surface areas (2). On the other hand, the latter has been described by the dynamic evolution of metabolism that is altered toward the supply of free P i . During P i starvation, overall phospholipid content that corresponds to 30% of total P i storage is decreased, and conversely, contents of nonphosphorus galactolipid, digalactosyldiacylglycerol (DGDG), 1 increase significantly (3). Galactolipids such as monogalactosyldiacylglycerol (MGDG) and DGDG are ubiquitous in plants, but are typically found only in plastids, especially in photosynthetic membranes (4). These galactolipids are synthesized by the galactosylation of diacylglycerol (DAG) by MGDG synthase an...

The Phosphate Fast-Responsive Genes PECP1 and PPsPase1 Affect Phosphocholine and Phosphoethanolamine Content

“…In response to a lack of Pi, we observed that NPC4, PLDz2, and GDPD1, 2, 3, 5, and 6 were induced in wild-type plants as previously reported (Supplemental Fig. 5; Nakamura et al, 2005;Li et al, 2006;Cheng et al, 2011), even if GDPD2 and 5 were only moderately induced. PLDz1 and PAH2 did not respond to Pi deficiency, and PAH1 was moderately (yet consistently) induced in the absence of Pi.…”

“…To verify if altered levels of PCho and PEth could lead to the transcriptomic activation or inhibition of specific lipid metabolism pathways, we selected several genes representative of membrane lipid recycling pathways triggered during Pi starvation (Nakamura, 2013). These genes included NPC4 (representative of the nonspecific PLC pathway; Nakamura et al, 2005), PLDz1 and 2, PAH1 and 2 (for the PLD/PAP pathway; Qin and Wang, 2002;Li et al, 2006;Nakamura et al, 2009;Eastmond et al, 2010), and GDPD1, 2, 3, 5, and 6 (for the LAH/GDPD pathway; Cheng et al, 2011). In response to a lack of Pi, we observed that NPC4, PLDz2, and GDPD1, 2, 3, 5, and 6 were induced in wild-type plants as previously reported (Supplemental Fig.…”

“…Many enzymes involved in this process are well known, and independent pathways are concomitantly triggered to ensure a fast and sizable lipid remobilization (Nakamura, 2013;Pant et al, 2015). For instance, SQD1 and SQD2 are involved in sulfolipid biosynthesis, whereas MGD2 and MGD3 are strongly induced to increase galactolipid content (Yu et al, 2002;Jouhet et al, 2004;Nakamura et al, 2005;Kobayashi et al, 2009). To achieve these substitutions, phospholipids are degraded by different types of lipases, resulting in the release of lipid subcomponents that can reenter the metabolism or be converted into other lipid forms (Nakamura, 2013).…”

Quantitative Profiling of Arabidopsis Polar Glycerolipids in Response to Phosphorus Starvation. Roles of Phospholipases Dζ1 and Dζ2 in Phosphatidylcholine Hydrolysis and Digalactosyldiacylglycerol Accumulation in Phosphorus-Starved Plants

“…Phospholipid-hydrolyzing enzymes, such as phospholipase D (PLD), and phospholipase C (PLC) have been proposed to be involved in the decrease in phospholipid content. The expression of one PC-hydrolyzing PLC was induced greatly in phosphorus-limited Arabidopsis (Arabidopsis thaliana), but the gene knockout of that PLC has no apparent effect on the lipid composition under normal and phosphorus-starved growth conditions (Nakamura et al, 2005). Of 12 Arabidopsis PLDs, PLDa(3), b(2), g(3), d, e, and z(2), the expression of the two PLDzs and particularly PLDz2 was induced most in phosphorus-deficient conditions (Misson et al, 2005;Li et al, 2006).…”

“…To date, changes in the major head-group classes, PC, PE, PI, PG, MGDG, and DGDG, during phosphorus starvation have been described (Benning and Ohta, 2005;Nakamura et al, 2005), but detailed analysis of lipid alterations of the molecular species has not been reported. In addition, alterations of the minor lipid classes, such as PA, PS, lysophosphatidylcholine (lysoPC), lysophosphatidylethanolamine (lysoPE), and lysophosphatidylglycerol (lysoPG), have not been investigated.…”