共查询到20条相似文献,搜索用时 31 毫秒
1.
2.
Jeanine Brederoo Piet de Wildt Corrie Popp-Snijders Robin F. Irvine Alan Musgrave Herman van den Ende 《Planta》1991,184(2):175-181
InChlamydomonas eugametos gametes, phosphatidylinositol 4-phosphate (PtdInsP) and phosphatidylinositol 4,5-bisphosphate (PtdInsP2) comprised 0.4 and 0.3% of the whole-cell phospholipids. They were concentrated in the plasma membrane around the cell body and were present in low concentrations in the flagellar membrane. When gametes were fed32PO
4
-
, the label was rapidly incorporated into PtdInsP and PtdInsP2 and only slowly incorporated into structural lipids such as phosphatidylethanolamine and phosphatidylglycerol. Similarly, when a pulse of32PO
4
-
was chased with PO
4
-
, the label was rapidly lost from the polyphosphoinositol lipids but not from the structural lipids. The major fatty acids in the polyphosphoinositides were C-22 carbon polyenoic acids (70%). The significance of these results in relationship to intracellular signalling via inositol phosphates and Ca2+ is discussed.Abbreviations InsP3
inositol 1,4,5-trisphosphate
- mt–/mt+
mating-type plus or minus
- PtdA
phosphatidic acid
- PtdEtn
phosphatidylethanolamine
- PtdGro
phosphatidylglycerol
- PtdIns
phosphatidylinositol
- PtdInsP
phosphatidylinositol 4-phosphate;
- PtdInsP2
phosphatidylinositol 4,5-bisphosphate
- TCA
trichloroacetic acid
We thank Frank Schuring for Fig. 5A and Susan Kenter, Hans Kruisselbrink, Saskia Bijvank and Nelleke Corbett for their enthousiastic assistance. 相似文献
3.
Mastoparan induces Ca2+-dependent deflagellation of the unicellular green alga Chlamydomonas moewusii Gerloff, as well as the activation of phospholipase C and the production of inositol 1,4,5-trisphosphate (InsP3; T. Munnik et al., 1998, Planta 207: 133–145). Even in the absence of extracellular Ca2+, mastoparan still induces deflagellation (L.M. Quarmby and H.C. Hartzell, 1994, J Cell Biol 124: 807–815; J.A.J. van Himbergen
et al., 1999, J Exp Bot, in press) suggesting that InsP3 mediates Ca2+ release from intracellular stores. To test this hypothesis, cells were pre-loaded with 45Ca2+ and their plasma membranes permeabilized by digitonin. Subsequent treatment of the cells with mastoparan (3.5 μM) induced
release of intracellular 45Ca2+. Mastoparan also activated phospholipase C in permeabilized cells, as demonstrated by the breakdown of 32P-phosphatidylinositol 4,5-bisphosphate and the production of diacylglycerol. The mastoparan analogues mas7 and mas17 were
also effective and their efficacy was correlated with their biological activity. X-ray microanalysis showed that electron-dense
bodies (EDBs) are a major Ca2+ store in C. moewusii. Analysis of digitonin-permeabilized cells showed that EDBs lost calcium at digitonin concentrations that released radioactivity
from 45Ca2+-labelled cells, suggesting that 45Ca2+ monitored the content of EDBs. X-ray microanaysis of living cells treated with mastoparan also revealed that calcium was
released from EDBs.
Received: 30 December 1998 / Accepted: 25 June 1999 相似文献
4.
Rates of production and consumption of phosphatidic acid upon thrombin stimulation of human platelets 总被引:1,自引:0,他引:1
Human platelets were labelled with [32P]Pi and [3H]glycerol before gel filtration. In unstimulated cells, the specific 32P radioactivity in phosphatidic acid (PtdOH) was similar to that of phosphatidylinositol (PtdIns) but only 4% of that of the gamma-phosphate of ATP. Upon 3 min of stimulation with 0.5 U/ml of thrombin, there was a 20-fold increase in specific 32P radioactivity of PtdOH which approached that of the ATP gamma-phosphate. Based on constant rates of synthesis and removal, this thrombin-induced increase in specific 32P radioactivity in PtdOH allowed us to calculate the flux of phosphate through PtdOH upon stimulation. Synthesis and removal occurred at rates of 107 and 52 nmol min-1/10(11) cells, respectively. The specific [3H]glycerol radioactivity was similar in PtdIns, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate in unstimulated platelets. In PtdOH, it was 50% of that of the inositol phospholipids. Thrombin stimulation induced no changes in the specific 3H radioactivity of the inositol phospholipids whereas specific [3H]PtdOH increased to the level of these lipids. It is concluded that PtdIns, PtdInsP and PtdInsP2 exist in a metabolic homogenous pool in human platelets. 相似文献
5.
Within the first 2 h of sexual reproduction, gametes of the green alga Chlamydomonas eugametos agglutinate, fuse via their mating structures, de-agglutinate and swim off as vis-à-vis pairs. During this period, increases in intracellular inositol 1,4,5-trisphosphate levels and changes in polyphosphoinositide synthesis were associated with cell fusion. The protein-kinase-C inhibitor staurosporine (0.1–0.2 M) inhibited the de-agglutination of pairs and therefore prevented them swimming away, while earlier stages of mating such as agglutination or cell fusion were unaffected. The results suggest that inositol 1,4,5-trisphosphate and diacylglycerol are fertilization signals in C. eugametos. The idea that they could also be fertilization signals in higher plants is discussed in relation to in vitro embryogenesis.Abbreviations DAG
diacylglycerol
- InsP3
inositol 1,4,5-trisphosphate
- mt+/mt–
mating-type plus or minus
- PKC
protein kinase C
- PtdOH
phosphatidic acid
- PtdInsP
phosphatidylinositol
- 4-phosphate PtdInsP2
phosphatidylinositol 4,5-bisphosphate 相似文献
6.
Karen E. Anderson Anna Kielkowska Tom N. Durrant Veronique Juvin Jonathan Clark Len R. Stephens Phillip T. Hawkins 《PloS one》2013,8(3)
We disrupted the gene encoding lysophosphatidylinositol-acyltransferase-1 (LPIAT1) in the mouse with the aim of understanding its role in determining cellular phosphoinositide content. LPIAT1−/− mice were born at lower than Mendelian ratios and exhibited a severe developmental brain defect. We compared the phospholipid content of livers and brains from LPIAT1−/− and LPIAT1+/+ littermates by LC-ESI/MS. In accord with previous studies, the most abundant molecular species of each phosphoinositide class (PtdIns, PtdInsP, PtdInsP2 and PtdInsP3) possessed a C38∶4 complement of fatty-acyl esters (C18∶0 and C20∶4 are usually assigned to the sn-1 and sn-2 positions, respectively). LPIAT1−/− liver and brain contained relatively less of the C38∶4 species of PtdIns, PtdInsP and PtdInsP2 (dropping from 95–97% to 75–85% of the total species measured for each lipid class) and relatively more of the less abundant species (PtdInsP3 less abundant species were below our quantification levels). The increases in the less abundant PtdIns and PtdInsP2 species did not compensate for the loss in C38∶4 species, resulting in a 26–44% reduction in total PtdIns and PtdInsP2 levels in both brain and liver. LPIAT1−/− brain and liver also contained increased levels of C18∶0 lyso-PtdIns (300% and 525% respectively) indicating a defect in the reacylation of this molecule. LPIAT1−/− brain additionally contained significantly reduced C38∶4 PC and PE levels (by 47% and 55% respectively), possibly contributing to the phenotype in this organ. The levels of all other molecular species of PC, PE, PS and PA measured in the brain and liver were very similar between LPIAT1−/− and LPIAT1+/+ samples. These results suggest LPIAT1 activity plays a non-redundant role in maintaining physiological levels of PtdIns within an active deacylation/reacylation cycle in mouse tissues. They also suggest that this pathway must act in concert with other, as yet unidentified, mechanisms to achieve the enrichment observed in C38∶4 molecular species of phosphoinositides. 相似文献
7.
The first responses in spruce [Picea abies (L.) Karst.] cells induced by elicitors (N-acetylglucosamine oligomers) from ectomycorrhizal fungi have been described as
follows: efflux of Cl− and K+, influx of Ca2+, extracellular alkalinization, phosphorylation of a 63-kDa protein (pp63), dephosphorylation of a 65-kDa protein (pp65) and
synthesis of H2O2 (Salzer et al. 1996, Planta 198: 118–126). In order to obtain new insights into the triggering mechanism and the sequence
of these rapid responses we used compounds which are known to activate or block specific steps within an elicitor-induced
signal transduction cascade in plant cells. Comparable to elicitors the two protein phosphatase inhibitors, cantharidin and
calyculin A, as well as mastoparan, an activator of trimeric G-proteins, were able to induce the release of Cl− and K+ from spruce cells and the alkalinization of the medium. Half-maximal activation of the alkalinization occurred at 133 nM
calyculin A, 2.3 μM cantharidin and 1.6 μ mastoparan. The structural analogue of mastoparan, Mas 17, which has no G-protein-stimulating
properties, was unable to trigger the above-mentioned reactions. In addition, cantharidin and calyculin A induced an increased
synthesis of H2O2 in spruce cells which was prolonged in comparison to the elicitor-induced transient formation of H2O2. Also, the cantharidin-induced release of K+ was more pronounced and longer lasting than that caused by elicitors from the ectomycorrhizal fungus Hebeloma crustuliniforme (Bull. ex Fries.) and N-acetylglucosamine oligomers. Furthermore, cantharidin, calyculin A and mastoparan induced the phosphorylation
of pp63. Remarkably, the protein kinase inhibitor, staurosporine, inhibited all the rapid responses described above, no matter
whether they were triggered by fungal elicitors or by the protein phosphatase inhibitors. These results indicate that in the
initial signalling events in spruce cells, essential protein phosphorylations occur either as an (auto) phosphorylation of
a membrane-bound receptor kinase prior to the activation of a G-protein or (and) immediately downstream of the activated G-protein
in a phosphorylation cascade and are the basic requirements for the ion fluxes following downstream.
Received: 24 April 1998 / Accepted: 23 July 1998 相似文献
8.
F. Morishita A. Shimada Y. Takeda M. Fujimoto H. Katayama K. Yamada 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1996,166(8):467-472
To investigate the functions of GTP-binding protein(s) in the melanosome-aggregating response in fish melanophores, the effects
of activators of G-proteins, namely, mastoparan and compound 48/80, were examined in cultured melanophores of the balck-moor
goldfish, Carassius auratus. Both mastoparan and compound 48/80 induced an approximately 40% increase in the GTP-hydrolyzing activity in the melanophore
membranes compared to the basal level. In intact melanophores, these compounds inhibited the effect of 3-isobutyl-1-methylxanthine,
which induced the accumulation of intracellular cAMP. Pretreatment of melanophores with pertussis toxin at 1 μg ⋅ ml-1 for 15 h attenuated the inhibitory effect of mastoparan on the accumulation of cAMP. However, pretreatment with the toxin
only slightly attenuated the inhibitory effect of compound 48/80 on the accumulation of cAMP. In addition, compound 48/80
at 1 mg ⋅ ml-1 induced full aggregation of the melanosomes in melanophores, though mastoparan at 5 μmol ⋅ l-1 induced only 10–20% aggregation of melanophores. These results suggest that mastoparan and compound 48/80 can each activate
the inhibitory G-protein in goldfish melanophores, which results in inhibition of adenylate cyclase activity. This signal-transduction
pathway is involved in the aggregation of melanosomes in these cells.
Accepted: 3 June 1996 相似文献
9.
Comparison of PDGF-AA- and PDGF-BB-Induced Phosphoinositide Formation in Human and Mouse Fibroblasts
In certain cells, such as human fibroblasts (AG 1523), there is a clear difference in the cell motility response induced by the different isoforms of platelet-derived growth factor (PDGF). PDGF-BB induces extensive actin reorganization and is a potent chemotactic agent, whereas PDGF-AA has a limited effect on actin reorganization and is not chemotactic. In the present study, we wanted to compare these effects on cell motility with the effects of the PDGF isoforms on phosphoinositide (PtdIns) turnover. We find that stimulation of serum-starved AG 1523 cells with PDGF-AA or PDGF-BB caused an initial increase of the phosphatidylinositol phosphate and bisphosphate (PtdInsP and PtdInsP2) pools, suggesting that activation of the phosphoinositide kinases is an initial response to PDGF stimulation. Despite a lower number of PDGF α-receptors than β-receptors on these cells, the initial formation of PtdInsP and PtdInsP2 appears to be stimulated to a similar degree by the two PDGF isoforms. In contrast, PtdInsP2 hydrolysis, indirectly measured as formation of phosphatidic acid, was correlated to the number of receptors. During prolonged exposure to PDGF-BB the stimulated PtdIns turnover remained at a high level, whereas the effect of PDGF-AA appeared more transient. A marked increase in the synthesis of a component migrating as phosphatidylinositol trisphosphate (PtdInsPa) was also detected after stimulation with PDGF-BB for 5 min. With PDGF-AA minor amounts were found, indicating that activation of the PtdIns 3′-kinase occurs also via the PDGF α-receptor. Stimulation with PDGF-BB, but not -AA, also induced a 50% decrease in lyso-PtdIns. In murine fibroblasts (Swiss 3T3), where the two PDGF isoforms have a similar effect on cell motility, the two PDGF isoforms also similarly induced PtdIns turnover, PtdInsP3 formation, and a decrease in lyso-PtdIns. Thus, there seems to be a correlation between PDGF-induced PtdIns turnover and PDGF-induced actin reorganization. This is compatible with previous evidence suggesting the microfilament formation is directly linked to an increased turnover of polyphosphoinositides in stimulated cells. 相似文献
10.
E. SANTAVENERE R. DI PIETRO L. DI DOMENICANTONIO R. RANA 《Cell biology international》1996,20(9):643-646
The effects of recombinant Tumor Necrosis Factor α (r-TNF α) on polyphosphoinositide metabolism were examined in a Burkitt Lymphoma cell line (Daudi cells). After 1h of in vitro treatment with r-TNF α, the incorporation of32Pi into phosphatidylinositol 4,5-phosphate (PtdInsP2), phosphatidylinositol 4-phosphate (PtdInsP) and phosphatidylinositol (PtdIns) was reduced compared with controls, confirming previous findings observed in other cell lines of a specific PtdIns breakdown following r-TNF α treatment. The novelty of this study is therefore the demonstration of early changes in polyphosphoinositide metabolism during the antiproliferative response elicited by this cytokine in Daudi cells. 相似文献
11.
Yuying Yuan Lizhen Mao Qixian Shi E. R. S. Roldan Wenying Chen Shuqing Yu Yuanzhong Zhuang Shifang Xu 《中国科学:生命科学英文版》2001,44(4):345-355
To investigate whether GABA/progesterone (P4) stimulates PPI breakdown and its role in the acrosome reaction (AR), spermatozoa of guinea pig were preincubated in MCM-LCa2+ for 5.5 h and then labeled with [32P]pi for 1 h. Samples were washed through a three-step gradient Percoll, adjusted to 5×107 cells/mL and exposed to 2 mmol/L Ca2+, 5 ?mol/L GABA, 10 ?mol/L P4 and other agents. Lipids were separated by t.l.c. and radioactivity in spots determined by scintillation counting. The AR was assessed by phase-contrast microscopy. The results showed that (i) when spermatozoa were treated with GABA, 32P-label diminished rapidly in phosphatidylinositol 4, 5-bisphosphate (PIP2), phosphatidylinositol 4-phosphate (PIP), and increased in phosphatidic acid (PA). The loss of label from PPI was almost completed by 10 min. The time-course of the AR was much slower than PPI when spermatozoa reached a maximal response by 15 min; (ii) the pattern of PPI hydrolysis and stimulation of AR was similar for the three agonists tested;their potency followed the order A23187>progesterone≥GABA; (iii) GABA-induced PIP2 hydrolysis and rise in PA and the AR were prevented by inclusion of 10 mmol/L neomycin; (iv) the loss of PIP2 labeling and the increase in PA labeling abolished when spermatozoa were exposed to EGTA or Ca2+ channel blocker. These results indicate that GABA or P4-induced PPI breakdown is an important and essential event in the series of changes to membrane fusion during the AR of guinea pig spermatozoa and this effect is mediated via calcium by activation of phosphatidylinositol-specific phospholipase C. 相似文献
12.
We show that differentiation of zoospores of the late blight pathogen Phytophthora infestans into cysts, a process called encystment, was triggered by both phosphatidic acid (PA) and the G-protein activator mastoparan. Mastoparan induced the accumulation of PA, indicating that encystment by mastoparan most likely acts through PA. Likewise, mechanical agitation of zoospores, which often is used to induce synchronized encystment, resulted in increased levels of PA. The levels of diacylglycerolpyrophosphate (DGPP), the phosphorylation product of PA, increased simultaneously. Also in cysts, sporangiospores, and mycelium, mastoparan induced increases in the levels of PA and DGPP. Using an in vivo assay for phospholipase D (PLD) activity, it was shown that the mastoparan-induced increase in PA was due to a stimulation of the activity of this enzyme. Phospholipase C in combination with diacylglycerol (DAG) kinase activity also can generate PA, but activation of these enzymes by mastoparan was not detected under conditions selected to highlight 32P-PA production via DAG kinase. Primary and secondary butanol, which, like mastoparan, have been reported to activate G-proteins, also stimulated PLD activity, whereas the inactive tertiary isomer did not. Similarly, encystment was induced by n- and sec-butanol but not by tert-butanol. Together, these results show that Phytophthora infestans contains a mastoparan- and butanol-inducible PLD pathway and strongly indicate that PLD is involved in zoospore encystment. The role of G-proteins in this process is discussed. 相似文献
13.
Jamie M Furneisen George M Carman 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2000,1484(1):71-82
The product of the LPP1 gene in Saccharomyces cerevisiae is a membrane-associated enzyme that catalyzes the Mg2+-independent dephosphorylation of phosphatidate (PA), diacylglycerol pyrophosphate (DGPP), and lysophosphatidate (LPA). The LPP1-encoded lipid phosphatase was overexpressed 681-fold in Sf-9 insect cells and used to examine the enzymological properties of the enzyme using PA, DGPP, and LPA as substrates. The optimum pH values for PA phosphatase, DGPP phosphatase, and LPA phosphatase activities were 7.5, 7.0, and 7.0, respectively. Divalent cations (Mn2+, Co2+, and Ca2+), NaF, heavy metals, propranolol, phenylglyoxal, and N-ethylmaleimide inhibited the PA phosphatase, DGPP phosphatase, and LPA phosphatase activities of the enzyme. The inhibitory effects of N-ethylmaleimide and phenylglyoxal on the LPP1-encoded enzyme were novel properties when compared with other Mg2+-independent lipid phosphate phosphatases from S. cerevisiae and mammalian cells. The LPP1-encoded enzyme exhibited saturation kinetics with respect to the surface concentrations of PA (Km=0.05 mol%), DGPP (Km=0.07 mol%), and LPA (Km=0.08 mol%). Based on specificity constants (Vmax/Km), the order of substrate preference was PA (4.2 units/mg/mol%)>DGPP (3.5 units/mg/mol%)>LPA (1.3 units/mg/mol%). DGPP (Ki=0.12 mol%) was a competitive inhibitor with respect to PA, and PA (Ki=0.12 mol%) was a competitive inhibitor with respect to DGPP. This suggested that the binding sites for these substrates were the same. The enzymological properties of the LPP1-encoded enzyme differed significantly from those of the S. cerevisiae DPP1-encoded lipid phosphatase, a related enzyme that also utilizes PA, DGPP, and LPA as substrates. 相似文献
14.
Nucleoside di- and triphosphates and adenosine regulate several components of the mucocilairy clearance process (MCC) that
protects the lung against infections, via activation of epithelial purinergic receptors. However, assessing the contribution
of individual nucleotides to MCC functions remains difficult due to the complexity of the mechanisms of nucleotide release
and metabolism. Enzymatic activities involved in the metabolism of extracellular nucleotides include ecto-ATPases and secreted
nucleoside diphosphokinase (NDPK) and adenyl kinase, but potent and selective inhibitors of these activities are sparse. In
the present study, we discovered that ebselen markedly reduced NDPK activity while having negligible effect on ecto-ATPase
and adenyl kinase activities. Addition of radiotracer [γ
32P]ATP to human bronchial epithelial (HBE) cells resulted in rapid and robust accumulation of [32P]-inorganic phosphate (32Pi). Inclusion of UDP in the incubation medium resulted in conversion of [γ
32P]ATP to [32P]UTP, while inclusion of AMP resulted in conversion of [γ
32P]ATP to [32P]ADP. Ebselen markedly reduced [32P]UTP formation but displayed negligible effect on 32Pi or [32P]ADP accumulations. Incubation of HBE cells with unlabeled UTP and ADP resulted in robust ebselen-sensitive formation of
ATP (IC50 = 6.9 ± 2 μM). This NDPK activity was largely recovered in HBE cell secretions and supernatants from lung epithelial A549
cells. Kinetic analysis of NDPK activity indicated that ebselen reduced the V
max of the reaction (K
i = 7.6 ± 3 μM), having negligible effect on K
M values. Our study demonstrates that ebselen is a potent non-competitive inhibitor of extracellular NDPK. 相似文献
15.
The inositol 1,4,5-trisphosphate (InsP3) receptor was purified from bovine cerebellum and reconstituted in liposomes composed of phosphatidylcholine (PC) and phosphatidylethanolamine
(PE) (1:1) successfully. No effect of Ca2+ concentration on [3H]-InsP3 binding to unreconstituted InsP3 receptor could be observed either at 4°C or at 25°C, whereas the effect of [Ca2+] on reconstituted InsP3 receptor depended on the temperature. The Ca2+ concentration outside the proteolipsome ([Ca2+]o) had no detectable effect on InsP3 binding to InsP3 receptor at 4°C. In contrast, with increase of [Ca2+]o from 0 to 100 nmol/L at 25°C, the InsP3 binding activity increased gradually. Then the InsP3 binding activity was decreased drastically at higher [Ca2+]o and inhibited entirely at 50 μmol/L [Ca2+]o. Conformational studies on intrinsic fluorescence of the reconstituted InsP3 receptor and its quenching by KI and HB indicated that the global conformation of reconstituted InsP3 receptor could not be affected by [Ca2+]o at 4°C. While at 25°C, the effects of 10 μmol/L [Ca2+]o on global, membrane and cytoplasmic conformation of the reconstituted InsP3 receptor were different significantly from that of 100 nmol/L [Ca2+]o. 相似文献
16.
C. E. Crocker J. J. Cech Jr. 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1998,168(1):50-60
The effect of environmental hypercapnia on respiratory and acid-base variables was studied in white sturgeon, Acipenser transmontanus. Blood PCO2, PO2, pH, hemoglobin concentration, and plasma lactate, glucose, catecholamines and cortisol were measured first under normocapnia
(water PCO2 < 0.5 Torr, 1 Torr = 133.32 Pa), then under hypercapnia (25–35 Torr) and a final return to normocapnia at 19 ± 0.5 °C. Acute
(≤ 2h) hypercapnia significantly increased arterial PCO2 (8-fold increase), ventilation frequency (2-fold increase), plasma HCO3
− (2.3-fold) and decreased arterial pH (to 7.15 ± 0.02). After 24 h, norepinephrine, epinephrine and cortisol, were significantly
increased, and arterial pH reached its nadir (7.10 ± 0.03). During the 72- and 96-h-periods, arterial PCO2 (24 ± 4.4 Torr) and ventilatory frequency (105 ± 5 breaths min−1) stabilized, HCO3
− reached its apparent maximum (23.6 ± 0.0 mmol−1), glucose decreased by 32%, and pH increased significantly to 7.31 + 0.03. The return to normocapnia completely restored
arterial PCO2 (2.5 ± 0.14 Torr), HCO3
− (7.4 ± 0.59 mmol · l−1), ventilation frequency (71 ± 7 breaths · min−1), and pH (7.75 ± 0.04). Overall, hypercapnia produced a respiratory acidosis, hyperventilation, a transient norepinephrine
“spike”, and increased plasma catecholamines, cortisol, and arterial PO2. The respiratory acidosis was only partially compensated (35% pH restoration) 96 h after the onset of hypercapnia and resulted
in a significantly decreased blood-O2 affinity (Bohr effect), as determined by construction of in vitro blood O2 equilibrium curves at 15 °C and 20 °C. Prolonged exposure to hypercapnia may lead to acid-base disturbances and negatively
affect growth of white sturgeon.
Accepted: 17 August 1997 相似文献
17.
Nod factor-induced phosphatidic acid and diacylglycerol pyrophosphate formation: a role for phospholipase C and D in root hair deformation 总被引:9,自引:0,他引:9
den Hartog M Musgrave A Munnik T 《The Plant journal : for cell and molecular biology》2001,25(1):55-65
Rhizobium-secreted nodulation factors are lipochitooligosaccharides that trigger the initiation of nodule formation on host legume roots. The first visible effect is root hair deformation, but the perception and signalling mechanisms that lead to this response are still unclear. When we treated Vicia sativa seedlings with mastoparan root hairs deformed, suggesting that G proteins are involved. To investigate whether mastoparan and Nod factor activate lipid signalling pathways initiated by phospholipase C (PLC) and D (PLD), seedlings were radiolabelled with [(32)P]orthophosphate prior to treatment. Mastoparan stimulated increases in phosphatidic acid (PA) and diacylglycerol pyrophosphate, indicative of PLD or PLC activity in combination with diacylglycerol kinase (DGK) and PA kinase. Treatment with Nod factor had similar effects, although less pronounced. The inactive mastoparan analogue Mas17 had no effect. The increase in PA was partially caused by the activation of PLD that was monitored by its in vivo transphosphatidylation activity. The application of primary butyl alcohols, inhibitors of PLD activity, blocked root hair deformation. Using different labelling strategies, evidence was provided for the activation of DGK. Since the PLC antagonist neomycin inhibited root hair deformation and the formation of PA, we propose that PLC activation produced diacylglycerol (DAG), which was subsequently converted to PA by DGK. The roles of PLC and PLD in Nod factor signalling are discussed. 相似文献
18.
Maize (Zea mays L.) cell cultures incorporated radioactivity from [14C]cinnamate into hydroxycinnamoyl-CoA derivatives and then into polysaccharide-bound feruloyl residues. Within 5–20 min, the
CoA pool had lost its 14C by turnover and little or no further incorporation into polysaccharides then occurred. The system was thus effectively a
pulse–chase experiment. Kinetics of radiolabelling of diferulates (also known as dehydrodiferulates) varied with culture age.
In young (1–3 d) cultures, polysaccharide-bound [14C]feruloyl- and [14C]diferuloyl residues were both detectable within 1 min of [14C]cinnamate feeding. Thus, feruloyl residues were dimerised <1 min after their attachment to polysaccharides. For at least
the first 2.3 h after [14C]cinnamate feeding, polysaccharide-bound [14C]diferuloyl residues remained almost constant at ≈7% of the total polysaccharide-bound [14C]ferulate derivatives. Since feruloyl residues are attached to polysaccharides <1 min after the biosynthesis of the latter,
and >10 min before secretion, the data show that extensive feruloyl coupling occurred intra-protoplasmically. Exogenous H2O2 (1 mM) caused little additional feruloyl coupling; therefore, wall-localised coupling may have been peroxidase-limited. In
older (e.g. 4 d) cultures, less intraprotoplasmic coupling occurred: during the first 2.5 h, polysaccharide-bound [14C]diferuloyl residues were a steady 1.4% of the total polysaccharide-bound [14C]ferulate derivatives. In contrast to the situation in younger cultures, exogenous H2O2 induced a rapid 4- to 6-fold increase in all coupling products, indicating that coupling in the walls was H2O2-limited. In both 2- and 4-d-old cultures, polysaccharide-bound 14C-trimers and larger coupling products exceeded [14C]diferulates 3- to 4-fold, but followed similar kinetics. Thus, although all known dimers of ferulate can now be individually
quantified, it appears to be trimers and larger products that make the major contribution to cross-linking of wall polysaccharides
in cultured maize cells. We argue that feruloyl arabinoxylans that are cross-linked before and after secretion are likely
to loosen and tighten the cell wall, respectively. The consequences for the control of cell expansion and for the response
of cell walls to an oxidative burst are discussed.
Received: 19 January 2000 / Accepted: 13 April 2000 相似文献
19.
Harold J. G. Meijer Nullin Divecha Herman van den Ende Alan Musgrave Teun Munnik 《Planta》1999,208(2):294-298
Cells from several different plant species synthesised a polyphosphoinositide (PPI)-like lipid when osmo-stressed. Synthesis
was maximal after about 10 min and was stimulated by a variety of osmolytes. Using NaCl, the strongest response centred around
200 mM. The lipid was shown to be the novel PPI isomer phosphatidyl-inositol 3,5-bisphosphate [PtdIns-(3,5)P2] by analytical thin-layer chromatography and conversion to PtdIns(3,4,5)P3 using recombinant phosphoinositide 4-OH kinase. The results indicate that PtdIns-(3,5)P2 plays a role in a general osmo-signalling pathway in plants. Its potential role is discussed.
Received: 6 November 1998 / Accepted: 14 December 1998 相似文献
20.
《Plant Physiology and Biochemistry》2002,40(11):899-906
Membrane preparations of Coffea arabica suspension cells were incubated in the presence of 〚32P〛γ-ATP. After lipid extraction and separation by thin layer chromatography, the following phosphorylated lipids were detected: phosphatidylinositol 4,5 bis-phosphate (PtdIns4,5P2), lyso-phosphatidylinositol 4-phosphate (LPtdIns4P), phosphatidylinositol 4-phosphate (PtdIns4P), diacylglycerol pyrophosphate (DGPP), lyso-phosphatidic acid (LPA) and phosphatidic acid (PA). This suggests the presence of phosphatidylinositol (EC 2.7.1.67), phosphatidylinositol 4 phosphate (EC 2.7.1.68), diacylglycerol (EC 2.7.1.107) and monoacylglycerol (EC 2.1.1.94) kinases. The activities of these lipid kinases changed during the culture period of the C. arabica cells reaching peak at day 7 of culture; however, enzymatic activities were very low before and after day 7. The behavior of these lipid kinases in the presence of their respective substrates and exogenous substrates such as ATP was characterized. The apparent Km values for ATP of all the lipid kinase activities were lower than 30 μM. All kinase activities assayed were totally dependent on the presence of Mg2+ and were unable to use Mn2+ or Ca2+ which produced a strong inhibition of all the lipid kinase activities. By using polyclonal antibodies against PtdIns 4-kinase and PtdInsP 5-kinase, we were able to identify at least two putative isoforms for the PtdIns 4-kinase and one for the PtdInsP 5-kinase. In both cases, the correlation of the amount of these proteins with their respective kinase activities depended on the culture cycle. The present work describes for the first time the characterization of the lipid kinases of C. arabica suspension cells, and the correlation of these activities with the culture cycle. 相似文献