Phosphatidylinositol, phosphatidylglycerol and cardiolipin synthesis in adult Dirofilaria immitis females

Srivastava A. K. and Jaffe J. J. 1987. Phosphatidylinositol, phosphatidylglycerol, and cardiolipin synthesis in adult Dirofilaria immitis females. International Journal for Parasitology17:917–920. The pathways leading to the formation of phosphatidylinositol (PI), phosphatidylglycerol (PG) and cardiolipin (CL) in adult Dirofilaria immitis females were investigated. PI was synthesized by both de novo as well as via base exchange pathway in the worms. Under specified assay conditions, the respective rates of PI formation by way of these pathways in crude homogenates of the worms in the order given were around 3.0 and 0.75 nmol min⁻¹ mg⁻¹ protein. PG synthesizing activity in the worms was mainly associated with the particulate fractions and the rate of formation by these fractions was around 1.5 nmol min⁻¹mg⁻¹ protein. The worms were unable to synthesize CL by the pathway found in mammals.     

Phospholipid Composition and Metabolism of Micrococcus denitrificans

The phospholipid composition of Micrococcus denitrificans was unusual in that phosphatidyl choline (PC) was a major phospholipid (30.9%). Other phospholipids were phosphatidyl glycerol (PG, 52.4%), phosphatidyl ethanolamine (PE, 5.8%), an unknown phospholipid (5.3%), cardiolipin (CL, 3.2%), phosphatidyl dimethylethanolamine (PDME, 0.9%), phosphatidyl monomethylethanolamine (PMME, 0.6%), phosphatidyl serine (PS, 0.5%), and phosphatidic acid (0.4%). Kinetics of ³²P incorporation suggested that PC was formed by the successive methylations of PE. Pulse-chase experiments with pulses of ³²P or acetate-1-¹⁴C to exponentially growing cells showed loss of isotopes from PMME, PDME, PS, and CL with biphasic kinetics suggesting the same type of multiple pools of these lipids as proposed in other bacteria. The major phospholipids, PC, PG, and PE, were metabolically stable under these conditions. The fatty acids isolated from the complex lipids were also unusual in being a simple mixture of seven fatty acids with oleic acid representing 86% of the total. Few free fatty acids and no non-extractable fatty acids associated with the cell wall or membrane were found.     

Oral transmission of Brugia pahangi and Dipetalonema viteae to adult and neonatal jirds

Sullivan J. J. and Chernin E. 1976. Oral transmission of Brugia pahangi and Dipetalonema viteae to adult and neonatal jirds. International Journal for Parasitology6: 75–78. Confirming previous studies, anaesthetized adult jirds became infected after oral doses of 100 infective larvae of B. pahangi; 4 of 5 jirds became microfilaria-positive and all 5 harbored adult worms. Among 7 unanaesthetized adult jirds similarly exposed, none developed microfilaraemia although 5 each harbored a few adult worms. In these unanaesthetized jirds, presumably, rapid passage of the inoculum through the mouth permitted fewer larvae to penetrate the mucosa, and the rest were probably killed in the stomach. Unanaesthetized 4-day old jirds proved highly and equally susceptible to oral or subcutaneous infection with B. pahangi as indicated by microfilaraemias and large worm-burdens. Direct and indirect evidence suggest that the baby's stomach and small intestine are not inimical to swallowed larvae, thus accounting for the relatively numerous mature worms in the peritoneal cavity. Third-stage larvae of D. viteae, readily infective subcutaneously, succeeded relatively infrequently in maturing when given orally to anaesthetized adult or to unanaesthetized baby jirds. Consistent oral infectivity may thus be a feature of filariae more closely related to B. pahangi.     

Changes in the pattern of phospholipid synthesis during the induction by cytokinin of cell division in soybean suspension cultures

A method is described for preparing fully viable, cytokinin-starved soybean (Glycine max (L.) Merr. cv. Acme) cells from a suspension-culture of callus tissue. The cells respond to kinetin treatment by re-initiating cell division. We present evidence, from the pattern of incorporation of ³²P-labelled inorganic phosphate into individual phospholipids during the first hour of this response, that the synthesis of phosphatidylinositol (PI) and of phosphatidic-acid head-groups is affected within 15 min. The polyphosphoinositide phosphatidylinositol 4-phosphate, but not phosphatidylinositol 4,5-bisphosphate, was detected in the tissue. The characteristics of cytokinin-induced PI synthesis in cytokinin-starved soybean cells appear to resemble the PI response of animal cells.Abbreviations DPG diphosphatidylglycerol - PA phosphatidic acid - PC phosphatidylcholine - PE phosphatidylethanolamine - PG phosphatidylglycerol - PI phosphatidylinositol - PIP phosphatidylinositol 4-phosphate - PIP₂ phosphatidylinositol 4,5-bisphosphate - PS phosphatidylserine - Pi inorganic phosphate - TLC thin-layer chromatography     

Phospholipid and phospholipase changes by jasmonic acid during stolon to tuber transition of potato

Potato tuber formation starts with the stolon swelling and is regulated by jasmonates. The cascade of events leading to tuber formation is not completely understood. The aim of this study was to evaluate phospholipid composition and phospholipase activities during four stages of stolon-to-tuber transition of Solanum tuberosum L., cv. Spunta, and involvement of phosphatidic acid (PA) in stolon cell expansion during early stages. Effects of jasmonic acid (JA) treatment on phospholipid content and activation of phospholipase D (PLD) (EC 3.1.4.4) and phosphatidylinositol-4,5-bisphosphate-specific phospholipase C (PIP₂-PLC) (EC 3.1.4.3) were studied in the early stages (first stage, hooked apex stolon; second stage, initial swelling stolon) of tuberization. All the phospholipid species identified, phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), PA, and cardiolipin (CL), decreased as tuber formation progressed. PLD and PLC were activated in control tissues at an early stage. JA treatment caused a decrease of PC and PS in first stage stolons, accumulation of PA in second stage stolons, and modification of PLD and PLC activities. PA increased stolon cell area in the first and second stages. These findings indicate that phospholipid catabolism is activated from the early stages of tuber formation, and that JA treatment modifies the pattern of phospholipid (PC, PS, and PA) composition and phospholipase (PLD and PLC) activity. These phospholipids therefore may play a role in activation of an intracellular mechanism that switches the developmental fate of stolon meristem cells, causing differentiation into a tuber.     

Brugia pahangi: uptake and incorporation of adenosine and thymidine.

The uptake and incorporation of adenosine and thymidine by infective larvae, 10-day-old juvenile, and adult stages of Brugia pahangi were investigated using scintillation counting and autoradiographic techniques. No evidence of thymidine incorporation by the worm was obtained in this study. Scintillation counting methods demonstrated that ¹⁴C-labeled adenosine was incorporated by all three stages of this filarial worm. Autoradiography, performed on worms incubated in [³H] adenosine from 5 min through 2 hr, revealed that following 5–15 min incubation the greatest degree of adenosine incorporation occurred in the hypodermis and somatic cords. Adenosine incorporation into the deeper body tissues, including the gut, increased significantly with longer periods of incubation. The results obtained further support the concept that nutrient uptake in B. pahangi occurs by a transcuticular route.     

Studies with Brugia pahangi. II. The effect of repeated infection on parasite levels in cats

Denham D. A., Ponnudurai T., Nelson G. S., Rogers Rosemary and Guy Frances 1972. Studies with Brugia pahangi—II. The effect of repeated infection on parasite levels in cats. International Journal for Parasitology2: 401–407. 21 cats were given a primary infection of 100–200 infective larvae of Brugia pahangi followed, some time later, by repeated challenge with 50 larvae per time at 10-day intervals. In most cats the microfilarial levels increased considerably but in a minority the levels remained the same as those seen in cats given only one infection. Adult worm recoveries were very much higher than after a single infection but after about 20 challenges there was no further increase in the number of worms establishing an infection. After a long series of challenge infections, the microfilarial counts of some cats suddenly fell and the blood became free of microfilariae.     

Brugia pahangi: infections and their effect on the lymphatic system of Mongolian jirds (Meriones unguiculatus)

Brugia pahangi has been found to be primarily a lymphatic-dwelling parasite in jirds when infections are induced by the subcutaneous injection of infective larvae or by allowing infected Aedes aegypti to feed.Migration to the regional lymphatics occurred as early as 1–4 days. Although some injected larvae remained in the skin for as long as 30 days and some became localized in the heart, lungs, pleural cavity, or peritoneal cavity, about three-fourths of the recovered filariae were found in the regional lymphatics. In contrast, when larvae were injected peritoneally they remained largely in the peritoneal cavity for at least 30 days.The relevant lymphatics and their drainage patterns in jirds have been described.The major pathological changes noted in jirds involved the regional lymphatic vessels and nodes, which were severely affected when they contained dead worms. Pulmonary granulomas due to dead microfilariae and occasionally to dead larvae or adult worms were noted.Observations are included on the susceptibility and course of B. pahangi infections in jirds.     

Yeast-Based High-Throughput Screens to Identify Novel Compounds Active against Brugia malayi

Background

Lymphatic filariasis is caused by the parasitic worms Wuchereria bancrofti, Brugia malayi or B. timori, which are transmitted via the bites from infected mosquitoes. Once in the human body, the parasites develop into adult worms in the lymphatic vessels, causing severe damage and swelling of the affected tissues. According to the World Health Organization, over 1.2 billion people in 58 countries are at risk of contracting lymphatic filariasis. Very few drugs are available to treat patients infected with these parasites, and these have low efficacy against the adult stages of the worms, which can live for 7–15 years in the human body. The requirement for annual treatment increases the risk of drug-resistant worms emerging, making it imperative to develop new drugs against these devastating diseases.

Methodology/Principal Findings

We have developed a yeast-based, high-throughput screening system whereby essential yeast genes are replaced with their filarial or human counterparts. These strains are labeled with different fluorescent proteins to allow the simultaneous monitoring of strains with parasite or human genes in competition, and hence the identification of compounds that inhibit the parasite target without affecting its human ortholog. We constructed yeast strains expressing eight different Brugia malayi drug targets (as well as seven of their human counterparts), and performed medium-throughput drug screens for compounds that specifically inhibit the parasite enzymes. Using the Malaria Box collection (400 compounds), we identified nine filarial specific inhibitors and confirmed the antifilarial activity of five of these using in vitro assays against Brugia pahangi.

Conclusions/Significance

We were able to functionally complement yeast deletions with eight different Brugia malayi enzymes that represent potential drug targets. We demonstrated that our yeast-based screening platform is efficient in identifying compounds that can discriminate between human and filarial enzymes. Hence, we are confident that we can extend our efforts to the construction of strains with further filarial targets (in particular for those species that cannot be cultivated in the laboratory), and perform high-throughput drug screens to identify specific inhibitors of the parasite enzymes. By establishing synergistic collaborations with researchers working directly on different parasitic worms, we aim to aid antihelmintic drug development for both human and veterinary infections.     

Fatty acid composition and incorporation of arachidonic acid into phospholipids of hemocytes from the tobacco hornworm Manduca sexta

The fatty acid compositions were determined for total lipids, triacylglycerols, phospholipids and four phospholipid fractions, including phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine/phosphatidylinositol (PS/PI) and cardiolipin (CA) obtained from hemocytes and cell-free serum from second day, fifth instar larvae of the tobacco hornworm Manduca sexta and the standard Manduca rearing medium. The hemocyte fatty acid profiles were considerably different from the profiles of the medium the insects were reared on and from the profiles of the cell-free serum. Hemocyte neutral lipids had lower proportions of polyunsaturated fatty acids than phospholipids. The fatty acid profiles of PC, PE, PS/PI and CA differ from each other and from the total lipid profiles, indicating selective fatty acid incorporation into hemocyte phospholipid species. Studies with radioactive arachidonic acid similarly indicated selective incorporation of polyunsaturated fatty acids into hemocyte lipids. Under our in vitro conditions, >40% of the total radioactivity was incorporated into hemocyte lipids. About 93% of the incorporated radioactivity was found in phospholipids. Within phospholipids. most of the radioactivity was associated with PC (46%), and less with PE (28%) and PS/PI (21%). Very little radioactivity was recovered in CA (0.9%).     

Studies with Brugia pahangi. I. Parasitological observations on primary infections of cats (Felis catus)

The large majority of cats given a single inoculation of third stage larvae of Brugia pahangi became microfilaraemic. Some cats had microfilariae in their blood 53 or 54 days after infection and most had become positive before 72 days after infection. In the majority of cats microfilarial counts remained very steady between 2 and 10 microfilariae per mm³ for long periods. At autopsy 10·7% of the infective larvae injected were recovered as adult worms. The recovery of adult worms was directly related to the number of larvae injected. The microfilarial level did not increase significantly with an increase in the number of adult worms.     

Discovery of a bifunctional cardiolipin/phosphatidylethanolamine synthase in bacteria

Phosphatidylethanolamine (PE) and cardiolipin (CL) are major components of bacterial and eukaryotic membranes. In bacteria, synthesis of PE usually occurs via decarboxylation of phosphatidylserine (PS) by PS decarboxylases (Psd). CL is produced by various CL synthases (Cls). Membranes of the plant pathogen Xanthomonas campestris predominantly contain PE, phosphatidylglycerol (PG) and CL. The X. campestris genome encodes one Psd and six putative CLs. Deletion of psd resulted in loss of PE and accumulation of PS. The mutant was severely affected in growth and cell size. PE synthesis, growth and cell division were partially restored when cells were supplied with ethanolamine (EA) suggesting a previously unknown PE synthase activity. Via mutagenesis, we identified a Cls enzyme (Xc_0186) responsible for EA‐dependent PE biosynthesis. Xanthomonas lacking xc_0186 not only lost its ability to utilize EA for PE synthesis but also produced less CL suggesting a bifunctional enzyme. Recombinant Xc_0186 in E. coli and in cell‐free extracts uses cytidine diphosphate diacylglycerol (CDP‐DAG) and PG for CL synthesis. It is also able to use CDP‐DAG and EA for PE synthesis. Owing to its dual function in CL and PE production, we consider Xc_0186 the founding member of a new class of enzymes called CL/PE synthase (CL/PEs).     

The molecular species of phospholipids of the cold-water soft coral <Emphasis Type="Italic">Gersemia rubiformis</Emphasis> (Ehrenberg, 1834) (Alcyonacea,Nephtheidae)

The chemical structures and contents of 68 molecular species of ethanolamine glycerophospholipids (PE), choline glycerophospholipids (PC), serine glycerophospholipids (PS), and inositol glycerophospholipids (PI) were determined in the asymbiotic soft coral Gercemia rubiformis (Ehrenberg, 1834) from the Sea of Okhotsk by the method of tandem high-resolution mass spectrometry. The major molecular species were 16:1e/20:4 PE, 16:0e/20:4 PC, 20:1/24:5 PS, and 16:0/24:5 PI. This study showed a significant similarity of the polar lipidomes of G. rubiformis and tropical species of alcyonarians with symbiotic microalgae and revealed the basic characteristic of the polar lipidome of these alcyonarians. It was found that the 24:5n-6 and 24:6n-3 acids (chemotaxonomic markers of soft corals) concentrate in the acyl groups of the molecular species of PS and PI, which can be used as molecular lipid markers in the study of the symbiotic and trophic relationships of soft corals.     

Lipid and fatty acids composition of photoautotrophically and heterotrophically grownChlamydomonas reinhardtii

Monogalactosyl diacylglycerol (MGDG), digalactosyl diacylglycerol (DGDG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), phosphatidylinositol (PI) and sulfoquinovosyl diacylglycerol (SQDG) are the most abundant lipid classes present in both the autotrophically and heterotrophically grownChlamydomonas reinhardtii. However, phosphatidylcholine (PC) and diacylglycerol (N,N,N-trimethyl)-homoserine (DGTS) were absent in both alga types. The polyne index B was higher in heterotrophic than photoautotrophic algae, but the unsaturation index was higher in photoautotrophic algae PI, PE and DGDG. The proportion of linolenic acid decreased under heterotrophy with compensatory increases in hexadecadienoic (16 : 3), oleic (18 : 1) and linoleic (18 : 3) acids.     

Specific induction of TaAAPT1, an ER- and Golgi-localized ECPT-type aminoalcoholphosphotransferase, results in preferential accumulation of the phosphatidylethanolamine membrane phospholipid during cold acclimation in wheat

Cold acclimation requires substantial alteration in membrane property. In contrast to well-documented fatty acid unsaturation during cold acclimation, changes in phospholipid biosynthesis during cold acclimation are less understood. Here, we isolated and characterized two aminoalcoholphosphotransferase (AAPT) cDNAs, TaAAPT1 and TaAAPT2, from wheat. AAPTs utilize diacylglycerols and CDP-choline/ethanolamine as substrates and catalyze the final step of the CDP-choline/ethanolamine pathway for phosphatidylcholine (PC)/phosphatidylethanolamine (PE) synthesis, respectively. Functionality of TaAAPT1 and TaAAPT2 was demonstrated by heterologous expression in a yeast cpt1Δ ept1Δ double mutant that lacks both AAPT activities. Detailed characterization of AAPT activities from the transformed mutant cells indicated that TaAAPT1 is an ECPT-type enzyme with higher ethanolamine phosphotransferase (EPT) activity than choline phosphotransferase (CPT) activity, while TaAAPT2 is a CEPT-type with the opposite substrate preference. Transient expression of GFP-fused TaAAPT1 and TaAAPT2 proteins in wheat and onion cells indicated they are localized to both the endoplasmic reticulum and Golgi apparatus, suggesting that the final synthesis of PE and PC via the CDP-choline/ethanolamine pathway occurs in these organella. Quantitative PCR analyses revealed that TaAAPT1 expression is strongly induced by cold, while TaAAPT2 was constitutively expressed at lower levels. Measurement of phospholipid content in wheat leaves indicated that PE is more prominently increased in response to cold than PC and accordingly PE/PC ratio increased from 0.385 to 0.530 during 14 days of cold acclimation. Together, these data suggested that an increase in the PE/PC ratio during cold acclimation is regulated at the final step of the biosynthetic pathway.     

NaCl-induced changes in plasma membrane lipids and proteins of <Emphasis Type="Italic">Zea mays</Emphasis> L. cultivars differing in their response to salinity

Two contrasting maize (Zea mays L.) cultivars, i.e., Giza 2 (salt tolerant) and Trihybrid 321 (salt sensitive), were grown hydroponically to study NaCl effect (100 mM) on root plasma membrane (PM) lipid and protein alterations. The PM total sterols of Trihybrid 321 were decreased while that of Giza 2 was increased in response to salt. Salt imposition had no significant effect on PM total glycolipids and proteins of both cultivars. The PM total phospholipids were increased in Trihybrid 321 but it did not change significantly in Giza 2 after salinity stress. Molecular percentage of PM phospholipids and fatty acids of both cultivars was different in absence (0 mM) and presence (100 mM) of salt. The most abundant phospholipids in untreated Trihybrid 321 PM were phosphatidylglycerol (PG), phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylserine (PS), which changed into PG, PS, phosphatidylinositol (PI) and PC after salt treatment. However, the dominant phospholipids of the control PM of Giza 2 were PC, PE, PS and PG, which changed into PG, PE, PS and diphosphatidylglycerol (DPG) after salt imposition. Over 60% of the total fatty acids were saturated in control and salinized PM of both cultivars, which was increased after salt stress. The predominant fatty acid in the control and salinized PM of Trihybrid 321 was C18:1 and C17:0, respectively. However, in control and treated PM of Giza 2, the predominant fatty acid was C17:0 and C20:0, respectively. Qualitative and quantitative differences in PM protein patterns were found in both cultivars with and without salt. PM lipid changes enhanced membrane integrity, reflected in different ion accumulation (Mansour et al. 2005), and hence salt tolerance of Giza 2.     

Carbohydrate metabolism in Brugia pahangi (Nematoda: Filarioidea)

Carbohydrate metabolism in Brugia pahangi (Nematoda:Filarioidea). International Journal for Parasitology16: 465–469. Adult B. pahangi have a complete glycolytic sequence and tricarboxylic acid cycle. The activities of the glycolytic enzymes are extremely high, but their relative activities are similar to those found in other glycolytic tissues. A comparison of the mass action ratios of the glycolytic enzymes with their apparent equilibrium constants indicates that phosphorylase, hexokinase, phosphofructokinase and pyruvate kinase are non-equilibrium regulatory enzymes. There is also some evidence that aldolase may be pseudoregulatory. Adult B. pahangi have measurable amounts of fumarate reductase as well as phosphoenolpyruvate carboxykinase and NADP-linked malic enzyme. Apart from lactate. the only other acidic end-product detected was traces of alanine; lactate production was little affected by the presence or absence of oxygen, unless exogenous glucose was present.     

Membrane phospholipid composition affects function of potassium channels from rabbit colon epithelium

We tested the effects of membrane phospholipids on the functionof high-conductance,Ca²⁺-activatedK⁺ channels from the basolateralcell membrane of rabbit distal colon epithelium by reconstituting thesechannels into planar bilayers consisting of different 1:1 mixtures ofphosphatidylethanolamine (PE), phosphatidylcholine (PC),phosphatidylserine (PS), and phosphatidylinositol (PI). At low ambientK⁺ concentrations single-channelconductance is higher in PE/PS and PE/PI bilayers than in PE/PCbilayers. At high K⁺concentrations this difference in channel conductance is abolished. Introducing the negatively charged SDS into PE/PC bilayersincreases channel conductance, whereas the positively chargeddodecyltrimethylammonium has the opposite effect. All these findingsare consistent with modulation of channel current by the charge of thelipid membrane surrounding the channel. But theK⁺ that permeates the channelsenses only a small fraction of the full membrane surface potential ofthe charged phospholipid bilayers, equivalent to separation of theconduction pathway from the charged phospholipid head groups by 20 Å. This distance appears to insulate the channel entrancefrom the bilayer surface potential, suggesting large dimensions of thechannel-forming protein. In addition, in PE/PC and PE/PI bilayers, butnot in PE/PS bilayers, the open-state probability of the channeldecreases with time ("channel rundown"), indicating thatphospholipid properties other than surface charge are required tomaintain channel fluctuations.

     

Comparative Plasma Lipidome between Human and Cynomolgus Monkey: Are Plasma Polar Lipids Good Biomarkers for Diabetic Monkeys?

Background

Non-human primates (NHP) are now being considered as models for investigating human metabolic diseases including diabetes. Analyses of cholesterol and triglycerides in plasma derived from NHPs can easily be achieved using methods employed in humans. Information pertaining to other lipid species in monkey plasma, however, is lacking and requires comprehensive experimental analysis.

Methodologies/Principal Findings

We examined the plasma lipidome from 16 cynomolgus monkey, Macaca fascicularis, using liquid chromatography coupled with mass spectrometry (LC/MS). We established novel analytical approaches, which are based on a simple gradient elution, to quantify polar lipids in plasma including (i) glycerophospholipids (phosphatidylcholine, PC; phosphatidylethanolamine, PE; phosphatidylinositol, PI; phosphatidylglycerol, PG; phosphatidylserine, PS; phosphatidic acid, PA); (ii) sphingolipids (sphingomyelin, SM; ceramide, Cer; Glucocyl-ceramide, GluCer; ganglioside mannoside 3, GM3). Lipidomic analysis had revealed that the plasma of human and cynomolgus monkey were of similar compositions, with PC, SM, PE, LPC and PI constituting the major polar lipid species present. Human plasma contained significantly higher levels of plasmalogen PE species (p<0.005) and plasmalogen PC species (p<0.0005), while cynomolgus monkey had higher levels of polyunsaturated fatty acyls (PUFA) in PC, PE, PS and PI. Notably, cynomolgus monkey had significantly lower levels of glycosphingolipids, including GluCer (p<0.0005) and GM₃ (p<0.0005), but higher level of Cer (p<0.0005) in plasma than human. We next investigated the biochemical alterations in blood lipids of 8 naturally occurring diabetic cynomolgus monkeys when compared with 8 healthy controls.

Conclusions

For the first time, we demonstrated that the plasma of human and cynomolgus monkey were of similar compositions, but contained different mol distribution of individual molecular species. Diabetic monkeys exhibited decreased levels of sphingolipids, which are microdomain-associated lipids and are thought to be associated with insulin sensitivity. Significant increases in PG species, which are precursors for cardiolipin biosynthesis in mitochondria, were found in fasted diabetic monkeys (n = 8).     

The effect of temperature on hairy root cultures of Catharanthus roseus: Growth,indole alkaloid accumulation and membrane lipid composition

Cultivation of Catharanthus roseus hairy root cultures at different temperatures was found to have an effect on growth rate and indole alkaloid content as well as lipid composition. When lowering the temperature, the roots responded by increasing the degree of unsaturation of cellular lipids, which was mainly due to an increased proportion of linolenic acid in the main lipid classes. The modifications in lipid composition were obviously necessary for the roots to retain the proper cell membrane fluidity at each temperature. Despite of changes in membrane lipids, no effect on the distribution of indole alkaloids between the roots and the medium could be detected. Instead, the level of alkaloid accumulation showed a clear increase with lowering temperature.Abbreviations PC phosphatidylcholine - PE phosphatidylethanolamine - PI phosphatidylinositol - PS phosphatidylserine - PG phosphatidylglycerol - CL cardiolipin - DGD digalactosyldiglyceride - MGD monogalactosyldiglyceride - NL neutral lipids - DU degree of fatty acid unsaturation