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Aluminum ions (Al) have been recognized as a major toxic factor for crop production in acidic soils. The first indication of the Al toxicity in plants is the cessation of root growth, but the mechanism of root growth inhibition is largely unknown. Here we examined the impact of Al on the expression, activity and function of the non-specific phospholipase C4 (NPC4), a plasma membrane-bound isoform of NPC, a member of the plant phospholipase family, in Arabidopsis thaliana.We observed a lower expression of NPC4 using GUS assay and a decreased formation of labeled diacylglycerol, product of NPC activity, using fluorescently labeled phosphatidylcholine as a phospholipase substrate in Arabidopsis WT seedlings treated with AlCl3 for 2 h. The effect on in situ NPC activity persisted for longer Al treatment periods (8, 14 h). Interestingly, in seedlings overexpressing NPC4, the Al-mediated NPC-inhibiting effect was alleviated at 14 h. However, in vitro activity and localization of NPC4 were not affected by Al, thus excluding direct inhibition by Al ions or possible translocation of NPC4 as the mechanisms involved in NPC-inhibiting effect. Furthermore, the growth of tobacco pollen tubes rapidly arrested by Al was partially rescued by the overexpression of AtNPC4 while Arabidopsis npc4 knockout lines were found to be more sensitive to Al stress during long-term exposure of Al at low phosphate conditions.Our observations suggest that NPC4 plays a role in both early and long-term responses to Al stress.

The actin cytoskeleton plays a key role in the plant morphogenesis and is involved in polar cell growth, movement of subcellular organelles, cell division, and plant defense. Organization of actin cytoskeleton undergoes dynamic remodeling in response to internal developmental cues and diverse environmental signals. This dynamic behavior is regulated by numerous actin-binding proteins that integrate various signaling pathways. Production of the signaling lipids phosphatidylinositol 4,5-bisphosphate and phosphatidic acid affects the activity and subcellular distribution of several actin-binding proteins, and typically correlates with increased actin polymerization. Here we review current knowledge of the inter-regulatory dynamics between signaling phospholipids and the actin cytoskeleton in plant cells.

Phosphatidic acid (PA) is a substantial intermediate in membrane lipid metabolism and also acts as an important component of the signalling pathways, regulating the spatio-temporal dynamics of the endomembrane system and cytoskeleton. Using tobacco pollen tubes as a model, we addressed signalling properties of PA in the single plant cell probing functions of three most relevant enzyme activities regulating production/localization and degradatin of PA – phospholipases D (PLD), diacylglycerol kinases (DGK) and lipid phosphate phosphatases (LPP). Phylogenetic analysis indicated a very dynamic evolution of all these three PA-modifying enzymes in land plants with many clade-specific duplications or losses. An in silico transcriptomic survey suggested enhanced recruitment of all three PA regulating activities into the pollen development – trend, surprisingly the most apparent in the case of DGKs. Using established specific inhibitors we could distinguish the contributions of PLDs, DGKs and LPPs into the PA regulated processes. LPPs inhibitors corroborated our previous data implying enhanced PA in the enhanced pollen tube growth – all LPPs inhibitors tested significantly stimulated pollen tube growth. The same effect was achieved by suppression of LPPs expression using antisense knock-down. In contrast, reduction of PA levels compromised membrane trafficking and pollen tube growth but not early endocytosis. Also the tip localised deposition of cell wall material (especially pectins) was disrupted after the inhibition of PA-producing enzymes. In contrast to PLD inhibition, DGK inhibition induced intracellular accumulation of pectins. While inhibition of DGK-produced PA changed vacuolar dynamics and morphology of pollen tubes, actin polymerization was positively regulated by PLD-derived PA. Our results clearly demonstrate strikingly different roles of PA produced by PLDs and DGKs in pollen tube growth.