Phospholipases (PLs) are lipid-hydrolyzing enzymes known to have diverse signaling roles during plant abiotic and biotic stress responses. They catalyze lipid remodeling, which is required to generate rapid responses of plants to environmental cues. Moreover, they produce second messenger molecules, such as phosphatidic acid (PA) and thus trigger or modulate signaling cascades that lead to changes in gene expression. The roles of phospholipases in plant abiotic and biotic stress responses have been intensively studied. Nevertheless, emerging evidence suggests that they also make significant contributions to plants' cellular and developmental processes. In this mini review, we summarized recent advances in the study of the cellular and developmental roles of phospholipases in plants.

• Klíčová slova
• cellular functions, phosphatidic acid, phospholipase A, phospholipase C, phospholipase D, phospholipases, phytohormones, plant development,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

BACKGROUND AND AIMS: The non-specific phospholipase C (NPC) is a new member of the plant phospholipase family that reacts to abiotic environmental stresses, such as phosphate deficiency, high salinity, heat and aluminium toxicity, and is involved in root development, silicon distribution and brassinolide signalling. Six NPC genes (NPC1-NPC6) are found in the Arabidopsis genome. The NPC2 isoform has not been experimentally characterized so far. METHODS: The Arabidopsis NPC2 isoform was cloned and heterologously expressed in Escherichia coli. NPC2 enzyme activity was determined using fluorescent phosphatidylcholine as a substrate. Tissue expression and subcellular localization were analysed using GUS- and GFP-tagged NPC2. The expression patterns of NPC2 were analysed via quantitative real-time PCR. Independent homozygous transgenic plant lines overexpressing NPC2 under the control of a 35S promoter were generated, and reactive oxygen species were measured using a luminol-based assay. KEY RESULTS: The heterologously expressed protein possessed phospholipase C activity, being able to hydrolyse phosphatidylcholine to diacylglycerol. NPC2 tagged with GFP was predominantly localized to the Golgi apparatus in Arabidopsis roots. The level of NPC2 transcript is rapidly altered during plant immune responses and correlates with the activation of multiple layers of the plant defence system. Transcription of NPC2 decreased substantially after plant infiltration with Pseudomonas syringae, flagellin peptide flg22 and salicylic acid treatments and expression of the effector molecule AvrRpm1. The decrease in NPC2 transcript levels correlated with a decrease in NPC2 enzyme activity. NPC2-overexpressing mutants showed higher reactive oxygen species production triggered by flg22. CONCLUSIONS: This first experimental characterization of NPC2 provides new insights into the role of the non-specific phospholipase C protein family. The results suggest that NPC2 is involved in the response of Arabidopsis to P. syringae attack.

• Klíčová slova
• Arabidopsis thaliana, MAMP-triggered immunity, Pseudomonas syringae, effector-triggered immunity, flagellin, non-specific phospholipase C, phosphatidylcholine-specific phospholipase C, reactive oxygen species,
• MeSH
• Arabidopsis enzymologie   imunologie   mikrobiologie   MeSH
• fosfatidylcholiny metabolismus   MeSH
• fosfolipasy typu C genetika   fyziologie   MeSH
• Golgiho aparát enzymologie   MeSH
• imunita rostlin fyziologie   MeSH
• klonování DNA MeSH
• konfokální mikroskopie MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• nemoci rostlin imunologie   mikrobiologie   MeSH
• proteiny huseníčku genetika   fyziologie   MeSH
• protoplasty enzymologie   MeSH
• Pseudomonas syringae * MeSH
• reaktivní formy kyslíku MeSH
• regulace genové exprese u rostlin MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfatidylcholiny MeSH
• fosfolipasy typu C MeSH
• NPC2 protein, Arabidopsis MeSH Prohlížeč
• proteiny huseníčku MeSH
• reaktivní formy kyslíku MeSH

The Arabidopsis non-specific phospholipase C (NPC) protein family is encoded by the genes NPC1 - NPC6. It has been shown that NPC4 and NPC5 possess phospholipase C activity; NPC3 has lysophosphatidic acid phosphatase activity. NPC3, 4 and 5 play roles in the responses to hormones and abiotic stresses. NPC1, 2 and 6 has not been studied functionally yet. We found that Arabidopsis NPC1 expressed in Escherichia coli possesses phospholipase C activity in vitro. This protein was able to hydrolyse phosphatidylcholine to diacylglycerol. NPC1-green fluorescent protein was localized to secretory pathway compartments in Arabidopsis roots. In the knock out T-DNA insertion line NPC1 (npc1) basal thermotolerance was impaired compared with wild-type (WT); npc1 exhibited significant decreases in survival rate and chlorophyll content at the seventh day after heat stress (HS). Conversely, plants overexpressing NPC1 (NPC1-OE) were more resistant to HS compared with WT. These findings suggest that NPC1 is involved in the plant response to heat.

• Klíčová slova
• Arabidopsis thaliana, diacylglycerol, heat stress, non-specific phospholipase C, phospholipids,
• Publikační typ
• časopisecké články MeSH

The first indication of the aluminum (Al) toxicity in plants growing in acidic soils is the cessation of root growth, but the detailed mechanism of Al effect is unknown. Here we examined the impact of Al stress on the activity of non-specific phospholipase C (NPC) in the connection with the processes related to the plasma membrane using fluorescently labeled phosphatidylcholine. We observed a rapid and significant decrease of labeled diacylglycerol (DAG), product of NPC activity, in Arabidopsis seedlings treated with AlCl₃. Interestingly, an application of the membrane fluidizer, benzyl alcohol, restored the level of DAG during Al treatment. Our observations suggest that the activity of NPC is affected by Al-induced changes in plasma membrane physical properties.

• Klíčová slova
• Arabidopsis thaliana, BA, benzyl alcohol, BODIPY, BODIPY, 4, 4-difluoro-4-bora-3a, 4a-diaza-s-indacene, BY-2, Bright Yellow 2, DAG, diacylglycerol, HP-TLC, high-performance thin-layer chromatography, MS, Murashige-Skoog, NPC, non-specific phospholipase C, PA, phosphatidic acid, PC, phosphatidylcholine, PC-PLC, phosphatidylcholine-specific phospholipase C, PI-PLC, phosphatidylinositol-specific phospholipase C, PIP2, phosphatidylinositol 4, 5-bisphosphate, PLD, phospholipase D, PM, plasma membrane., aluminum toxicity, benzyl alcohol, diacylglycerol, membrane fluidity, non-specific phospholipase C,
• MeSH
• Arabidopsis účinky léků   enzymologie   MeSH
• benzylalkohol farmakologie   MeSH
• buněčná membrána účinky léků   metabolismus   MeSH
• diglyceridy metabolismus   MeSH
• fosfolipasy typu C metabolismus   MeSH
• hliník farmakologie   MeSH
• ionty MeSH
• kořeny rostlin účinky léků   metabolismus   MeSH
• semenáček účinky léků   metabolismus   MeSH
• sloučeniny boru metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 1,2-diacylglycerol MeSH Prohlížeč
• 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene MeSH Prohlížeč
• benzylalkohol MeSH
• diglyceridy MeSH
• fosfolipasy typu C MeSH
• hliník MeSH
• ionty MeSH
• sloučeniny boru MeSH

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 β-glucuronidase 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.

• Klíčová slova
• Arabidopsis, aluminum toxicity, diacylglycerol, non-specific phospholipase C, plasma membrane, pollen tube, signaling, tobacco,
• Publikační typ
• časopisecké články MeSH

Phosphatidylcholine-hydrolysing phospholipase C, also known as non-specific phospholipase C (NPC), is a new member of the plant phospholipase family that reacts to environmental stresses such as phosphate deficiency and aluminium toxicity, and has a role in root development and brassinolide signalling. Expression of NPC4, one of the six NPC genes in Arabidopsis, was highly induced by NaCl. Maximum expression was observed from 3 h to 6 h after the salt treatment and was dependent on salt concentration. Results of histochemical analysis of P(NPC4):GUS plants showed the localization of salt-induced expression in root tips. On the biochemical level, increased NPC enzyme activity, indicated by accumulation of diacylglycerol, was observed as early as after 30 min of salt treatment of Arabidopsis seedlings. Phenotype analysis of NPC4 knockout plants showed increased sensitivity to salinity as compared with wild-type plants. Under salt stress npc4 plants had shorter roots, lower fresh weight, and reduced seed germination. Expression levels of abscisic acid-related genes ABI1, ABI2, RAB18, PP2CA, and SOT12 were substantially reduced in salt-treated npc4 plants. These observations demonstrate a role for NPC4 in the response of Arabidopsis to salt stress.

• MeSH
• Arabidopsis účinky léků   enzymologie   genetika   metabolismus   MeSH
• chlorid sodný metabolismus   farmakologie   MeSH
• fosfolipasy typu C genetika   metabolismus   fyziologie   MeSH
• geneticky modifikované rostliny MeSH
• kořeny rostlin účinky léků   enzymologie   genetika   metabolismus   MeSH
• kyselina abscisová genetika   metabolismus   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• proteiny huseníčku genetika   metabolismus   fyziologie   MeSH
• regulace genové exprese u rostlin MeSH
• signální transdukce MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chlorid sodný MeSH
• fosfolipasy typu C MeSH
• kyselina abscisová MeSH
• NPC4 protein, Arabidopsis MeSH Prohlížeč
• proteiny huseníčku MeSH