Intracellular levels of Mg2+ are tightly regulated, as Mg2+ deficiency or excess affects normal plant growth and development. In Arabidopsis, we determined that phospholipase Dα1 (PLDα1) is involved in the stress response to high-magnesium conditions. The T-DNA insertion mutant pldα1 is hypersensitive to increased concentrations of magnesium, exhibiting reduced primary root length and fresh weight. PLDα1 activity increases rapidly after high-Mg2+ treatment, and this increase was found to be dose dependent. Two lines harbouring mutations in the HKD motif, which is essential for PLDα1 activity, displayed the same high-Mg2+ hypersensitivity of pldα1 plants. Moreover, we show that high concentrations of Mg2+ disrupt K+ homeostasis, and that transcription of K+ homeostasis-related genes CIPK9 and HAK5 is impaired in pldα1. Additionally, we found that the akt1, hak5 double mutant is hypersensitive to high-Mg2+ . We conclude that in Arabidopsis, the enzyme activity of PLDα1 is vital in the response to high-Mg2+ conditions, and that PLDα1 mediates this response partially through regulation of K+ homeostasis.
- Klíčová slova
- Arabidopsis thaliana, CIPK9, HAK5, high magnesium, ion homeostasis, phospholipase D, potassium,
- MeSH
- Arabidopsis enzymologie metabolismus fyziologie MeSH
- draslík metabolismus MeSH
- fosfolipasa D metabolismus fyziologie MeSH
- fyziologický stres MeSH
- homeostáza MeSH
- hořčík metabolismus MeSH
- proteiny huseníčku metabolismus fyziologie MeSH
- transkriptom MeSH
- western blotting MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- draslík MeSH
- fosfolipasa D MeSH
- hořčík MeSH
- PLDA1 protein, Arabidopsis MeSH Prohlížeč
- proteiny huseníčku MeSH
The plant selective autophagy cargo receptor neighbour of breast cancer 1 gene (NBR1) has been scarcely studied in the context of abiotic stress. We wanted to expand this knowledge by using Arabidopsis thaliana lines with constitutive ectopic overexpression of the AtNBR1 gene (OX lines) and the AtNBR1 Knock-Out (KO lines). Transcriptomic analysis of the shoots and roots of one representative OX line indicated differences in gene expression relative to the parental (WT) line. In shoots, many differentially expressed genes, either up- or down-regulated, were involved in responses to stimuli and stress. In roots the most significant difference was observed in a set of downregulated genes that is mainly related to translation and formation of ribonucleoprotein complexes. The link between AtNBR1 overexpression and abscisic acid (ABA) signalling was suggested by an interaction network analysis of these differentially expressed genes. Most hubs of this network were associated with ABA signalling. Although transcriptomic analysis suggested enhancement of ABA responses, ABA levels were unchanged in the OX shoots. Moreover, some of the phenotypes of the OX (delayed germination, increased number of closed stomata) and the KO lines (increased number of lateral root initiation sites) indicate that AtNBR1 is essential for fine-tuning of the ABA signalling pathway. The interaction of AtNBR1 with three regulatory proteins of ABA pathway (ABI3, ABI4 and ABI5) was observed in planta. It suggests that AtNBR1 might play role in maintaining the balance of ABA signalling by controlling their level and/or activity.
- MeSH
- Arabidopsis genetika metabolismus MeSH
- autofagie * MeSH
- geneticky modifikované rostliny MeSH
- klíčení MeSH
- kyselina abscisová metabolismus MeSH
- proteiny huseníčku genetika metabolismus MeSH
- regulace genové exprese u rostlin MeSH
- semena rostlinná genetika MeSH
- semenáček MeSH
- signální transdukce * MeSH
- transportní proteiny genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kyselina abscisová MeSH
- NBR1 protein, Arabidopsis MeSH Prohlížeč
- proteiny huseníčku MeSH
- transportní proteiny MeSH
The phytohormone salicylic acid (SA) has a crucial role in plant physiology. Its role is best described in the context of plant response to pathogen attack. During infection, SA is rapidly accumulated throughout the green tissues and is important for both local and systemic defences. However, some genetic/metabolic variations can also result in SA overaccumulation in plants, even in basal conditions. To date, more than forty Arabidopsis thaliana mutants have been described as having enhanced endogenous SA levels or constitutively activated SA signalling pathways. In this study, we established a collection of mutants containing different SA levels due to diverse genetic modifications and distinct gene functions. We chose prototypic SA-overaccumulators (SA-OAs), such as bon1-1, but also "non-typical" ones such as exo70b1-1; the selection of OA is accompanied by their crosses with SA-deficient lines. Here, we extensively studied the plant development and SA level/signalling under various growth conditions in soil and in vitro, and showed a strong negative correlation between rosette size, SA content and PR1/ICS1 transcript signature. SA-OAs (namely cpr5, acd6, bon1-1, fah1/fah2 and pi4kβ1β2) had bigger rosettes under high light conditions, whereas WT plants did not. Our data provide new insights clarifying a link between SA and plant behaviour under environmental stresses. The presented SA mutant collection is thus a suitable tool to shed light on the mechanisms underlying trade-offs between growth and defence in plants.
- Klíčová slova
- Arabidopsis mutants, Salicylic acid, gene transcription, growth, light,
- MeSH
- Arabidopsis genetika metabolismus mikrobiologie MeSH
- interakce hostitele a patogenu MeSH
- kyselina salicylová metabolismus MeSH
- mutace * MeSH
- nemoci rostlin genetika mikrobiologie MeSH
- proteiny huseníčku genetika metabolismus MeSH
- regulace genové exprese u rostlin * MeSH
- regulátory růstu rostlin metabolismus MeSH
- signální transdukce genetika MeSH
- vývoj rostlin genetika MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kyselina salicylová MeSH
- proteiny huseníčku MeSH
- regulátory růstu rostlin MeSH
Recognition of pathogen-associated molecular patterns (PAMPs) is crucial for plant defence against pathogen attack. The best characterized PAMP is flg22, a 22 amino acid conserved peptide from flagellin protein. In Arabidopsis thaliana, flg22 is recognized by the flagellin sensing 2 (FLS2) receptor. In this study, we focused on biotic stress responses triggered by flg22 after exposure to temporary heat stress (HS). It is important to study the reactions of plants to multiple stress conditions because plants are often exposed simultaneously to a combination of both abiotic and biotic stresses. Transient early production of reactive oxygen species (ROS) is a well-characterized response to PAMP recognition. We demonstrate the strong reduction of flg22-induced ROS production in A. thaliana after HS treatment. In addition, a decrease in FLS2 transcription and a decrease of the FLS2 presence at the plasma membrane are shown after HS. In summary, our data show the strong inhibitory effect of HS on flg22-triggered events in A. thaliana. Subsequently, temporary HS strongly decreases the resistance of A. thaliana to Pseudomonas syringae. We propose that short exposure to high temperature is a crucial abiotic stress factor that suppresses PAMP-triggered immunity, which subsequently leads to the higher susceptibility of plants to pathogens.
- Klíčová slova
- Arabidopsis thaliana, Pseudomonas syringae, PAMP-triggered immunity, flagellin sensing 2 receptor, flg22, heat stress, reactive oxygen species,
- MeSH
- alarminy metabolismus MeSH
- Arabidopsis účinky léků genetika imunologie mikrobiologie MeSH
- flagelin farmakologie MeSH
- genetická transkripce účinky léků MeSH
- imunita rostlin * účinky léků MeSH
- nemoci rostlin imunologie mikrobiologie MeSH
- odolnost vůči nemocem imunologie MeSH
- proteiny huseníčku genetika metabolismus MeSH
- Pseudomonas syringae účinky léků fyziologie MeSH
- reakce na tepelný šok * účinky léků MeSH
- regulace genové exprese u rostlin účinky léků MeSH
- respirační vzplanutí účinky léků MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- alarminy MeSH
- flagelin MeSH
- proteiny huseníčku 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
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
