Metabolism of Unsaturated Monogalactosyldiacylglycerol Molecular Species in Arabidopsis thaliana Reveals Different Sites and Substrates for Linolenic Acid Synthesis

Synthesis of unsaturated monogalactosyldiacylglycerol (MGDG) was examined in a mutant of Arabidopsis thaliana (L.) Heynh. containing reduced levels of hexadecatrienoic (16:3) and linolenic (18:3) acids in leaf lipids. Molecular species composition and labeling kinetics following the incorporation of exogenous [¹⁴C]fatty acids suggest that at least two pathways and multiple substrates are involved in desaturation of linoleic acid (18:2) to 18:3 for production of unsaturated galactolipids. A reduction in 18:3/16:3 MGDG and an increase in 18:2/16:2 MGDG, together with labeling kinetics of these molecular species following the incorporation of exogenous [¹⁴C]12:0 fatty acids, suggests that a chloroplastic pathway for production of 18:3 at the sn-1 position of MGDG utilizes 18:2/16:2 MGDG as a substrate. This chloroplastic (prokaryotic) pathway is deficient in the mutant. When exogenous [¹⁴C]18:1 was supplied, a eukaryotic (cytoplasmic) pathway involving the desaturation of 18:2 to 18:3 on phosphatidylcholine serves as the source of 18:3 for the sn-2 position of MGDG. This eucaryotic pathway predominates in the mutant.     

Polyunsaturated Fatty Acid Synthesis by a Mixture of Chloroplasts and Microsomes from Spinach Leaves: Evidence for Two Distinct Pathways of the Biosynthesis of Trienoic Acids

In a mixture of chloroplasts and microsomes from spinach leaves,all the leaf lipids were synthesized from (1-¹⁴C)-acetate. Inthis system, all the lipids contained labelled oleate, linoleateand linolenate but labelled linolenate was mainly concentratedinto diacylgalactosylglycerol (MGDG). A small but significantlabelling was found in the linolenate of the diacyldigalactosylglycerol(DGDG). On the other hand, labelled hexadecamonoenoic acid (C^16:1),hexadecadienoic acid (C^16:2) and hexadecatrienoic acid (C^16:3)were only found into MGDG. In such a reconstituted system, atthe end of the incubation period, labelled MGDG was almost exclusivelyrecovered into the chloroplast while the labelled phosphatidylcholine(PC) was found highly concentrated in the microsomes In the MGDG of the chloroplast, C^16:1, C^16:2 and C^16:3 werefound at the C2 position of the glycerol while oleic acid (C^18:1),linoleic acid (C^18:2) and a-linolenic acid (^18:3) esterifiedspecifically the position 1 of the glycerol. No C¹⁸ acids werefound in position 2. In the PC of the microsomes, C^18:1, C^18:2and C^18:3 were found at the Cl and C2 positions of the glycerolwhile palmitic acid esterified exclusively the Cl of the glycerol. The biosynthetic pathway of trienoic fatty acids in leaves ofhigher plants is discussed. (Received July 19, 1982; Accepted October 18, 1982)     

Differential Effects of a Substituted Pyridazinone, BASF 13-338, on Pathways of Monogalactosyldiacylglycerol Synthesis in Arabidopsis

We have examined the effects of the substituted pyridazinone herbicide, 4-chloro-5-(dimethylamino)-2-phenyl-3(2H)pyridazinone (BASF 13-338, Sandoz 9785), on the desaturation of linoleic acid (18:2) on different molecular species of monogalactosyldiacylglycerol (MGDG) and phosphatidylcholine (PC) in leaf tissue of Arabidopsis thaliana (L.) Heynh. Specific changes in lipid composition allowed identification of different substrates for desaturation of 18:2 to linolenic acid (18:3). 18:2/16:2 MGDG was desaturated in the chloroplast to form 18:3/16:3 MGDG. Levels of 18:3/16:3 MGDG were reduced by treatment with BASF 13-338, suggesting that both the formation of 18:3 at the sn-1 position, and the formation of 16:3 at the sn-2 position of 18:2/16:2 MGDG were inhibited by this compound. Kinetic studies using exogenously incorporated [¹⁴C] 18:1 indicated that 18:2/18:3 MGDG originated from an 18:2/18:3 diglyceride precursor derived from PC. The formation of 18:3 at the sn-1 position of 18:2/18:3 MGDG was also inhibited by BASF 13-338. In contrast the desaturation of 18:2 proposed to occur at the sn-2 position of PC outside the chloroplast, was not affected.     

Lipids of Ectocarpus fasciculatus (Phaeophyceae). Incorporation of [l-14C]Oleate and the Role of TAG and MGDG in Lipid Metabolism

Lipids and fatty acids of Ectocarpus fasciculatus (Ectocarpales,Phaeophyceae) were analyzed. Major polar lipids are monogalactosyldiacylglycerol(MGDG), digalactosyldiacylglycerol (DGDG), sulfoquinovosyldiacylglycerol(SQDG), diacylglycerylhydroxymethyl-N,N,N-trimethyl-rß-alanine(DGTA), phosphatidylcholine (PC), phospha-tidylethanolamine(PE), phosphatidylglycerol (PG) and phosphatidylinositol (PI).Diphosphatidylglycerol (DPG), phosphatidic acid (PA) and phosphatidyl-O-[N-(2-hydroxy-ethyl)glycine](PHEG) were also present in small amounts. Nonpolar lipids mainlyconsist of triacylglycerol (TAG) and diacylglycerol (DAG). Majorfatty acids are 16:0,18:1, 18:3, 18:4, 20:4 and 20:5. The positionaldistribution of fatty acids showed that molecular species ofeukaryotic structure account for 99% in MGDG, 98% in DGDG, 62%in PG and 23% in SQDG. On incubation with [1-¹⁴C]18:1 for 30min, 33% of the total label was detected in TAG, 16% in PG,14% in PE, 10% in PC and 8% in MGDG. During 7 days of chase,the label in TAG, PG, PE and PC decreased and simultaneouslyincreased in MGDG up to 41% of the total. In SQDG, labelledfatty acids were found in prokaryotic as well as in eukaryoticmolecular species. During the experiment, the label shiftedfrom 18:1 to 18:2, 18:3, 18:4 and, to a minor extent, to 20:4and 20:5 acids indicating 18:1 to be processed by elongationand/or desaturation. These results suggest TAG to act as a majorprimary acceptor of exogenous oleate and to be involved in thetransfer of fatty acids to MGDG and other polar lipids. (Received March 24, 1997; Accepted June 11, 1997)     

Fatty Acid Composition of Polar Lipids in Ceratodon purpureus and Pleurozium schreberi

The total amount of fatty acids in the mono- (MGDG) and diglycosyl diglyceride (DGDG) and more polar lipid fractions of frozen Ceratodon purpureus shoots was 4.6, 3.4 and 4.0 mg/g dry weight, respectively. The respective values for the tops of frozen Pleurozium schreberi were 2.6, 3.3 and 3.8 mg/g dry weight. The molar ratios MGDG/DGDG and MGDG + DGDG/chlorophyll were 1.3 and 3.7, respectively, for C. purpureus and 0.8 and 3.5 for P. schreberi. In C. purpureus the main fatty acids in the MGDG fraction were C 18:3ω3 (44% of the total fatty acids) and C 16:3ω3 (26%); in the DGDG fraction C 18:3ω3 (70%); and in the more polar lipid fraction C 18: 3ω3 (26%) and C 16:0 (25%). The proportion of C 20 polyunsaturated fatty acids was 15, 12 and 19% of the total fatty acids found in the MGDG, DGDG and more polar lipid fractions, respectively. In P. schreberi the proportion of C 20 polyunsaturated fatty acids was high in all polar lipid fractions (47, 42 and 25% in MGDG, DGDG and more polar lipid fractions, respectively). In addition, MGDG and DGDG fractions contained abundantly C 18:3ω3 (32 and 45%, respectively), and the more polar lipid fraction both C 18: 3ω3 (24%) and C 16:0 (27%).     

Drought stress affects chloroplast lipid metabolism in rape (Brassica napus) leaves

Rape ( Brassica napus L. var. Bienvenue) is a 16:3 plant which contains predominantly prokaryotic species of monogalactosyldiacylglycerol i.e. sn-1 C18, sn-2 C16 (C18/C16 MGDG). Rape plants were exposed to a restricted water supply for 12 days. Under drought conditions, considerable changes in lipid metabolism were observed. Drought stress provoked a decline in leaf polar lipids, which is mainly due to a decrease in MGDG content. Determination of molecular species in phosphatidylcholine (PC) and MGDG indicated that the prokaryotic molecular species of MGDG (C18/C16) decreased after drought stress while the eukaryotic molecular species (C18/C18) remained stable. Drought stress had different effects on two key enzymes of PC and MGDG synthesis. The in vitro activity of MGDG synthase (EC. 2.4.1.46) was reduced in drought stressed plants whereas cholinephosphotransferase (EC. 2.7.8.2) activity was not affected. Altogether these results suggest that the prokaryotic pathway leading to MGDG synthesis was strongly affected by drought stress while the eukaryotic pathway was not. It was also observed that the molecular species of leaf PC became more saturated in drought stressed plants. This could be due to a specific decrease in oleate desaturase activity.     

Conversion of monogalactosyldiacylglycerols to triacylglycerols in ozone-fumigated spinach leaves

Molecular species and fatty acid distribution of triacylglycerol (TG) accumulated in spinach (Spinacia oleracea L.) leaves fumigated with ozone (0.5 microliter per liter) were compared with those of monogalactosyldiacylglycerol (MGDG). Analysis of positional distribution of the fatty acids in MGDG and the accumulated TG by the enzymatic digestion method showed that hexadecatrienoate (16:3) was restricted to sn-2 position of the glycerol backbone in both MGDG and TG, whereas α-linolenate (18:3) was preferentially located at sn-1 position in MGDG, and sn-1 and/or sn-3 positions in TG, suggesting that 1,2-diacylglycerol moieties of MGDG are the direct precursor of TG in ozonefumigated leaves. Further analysis of TG molecular species by argentation chromatography and mass spectrometry showed that TG increased with ozone fumigation consisted of approximately an equal molar ratio of sn-1,3-18:3-2-16:3 and sn-1,2,3-18:3. Because the molecular species of MGDG in spinach leaves is composed of a similar molar ratio of sn-1-18:3-2-16:3 and sn-1,2-18:3, we concluded that MGDG was converted to 1,2-diacylglycerol and acylated with 18:3 to TG in ozone-fumigated spinach leaves.     

Biosynthesis and desaturation of prokaryotic galactolipids in leaves and isolated chloroplasts from spinach

Mono- and digalactosyldiacylglycerol (MGDG and DGDG) were isolated from the leaves of sixteen 16:3 plants. In all of these plant species, the sn-2 position of MGDG was more enriched in C₁₆ fatty acids than sn-2 of DGDG. The molar ratios of prokaryotic MGDG to prokaryotic DGDG ranged from 4 to 10. This suggests that 16:3 plants synthesize more prokaryotic MGDG than prokaryotic DGDG. In the 16:3 plant Spinacia oleracea L. (spinach), the formation of prokaryotic galactolipids was studied both in vivo and in vitro. In intact spinach leaves as well as in chloroplasts isolated from these leaves, radioactivity from [1-¹⁴C]acetate accumulated 10 times faster in MGDG than in DGDG. After 2 hours of incorporation, most labeled galactolipids from leaves and all labeled galactolipids from isolated chloroplasts were in the prokaryotic configuration. Both in vivo and in vitro, the desaturation of labeled palmitate and oleate to trienoic fatty acids was higher in MGDG than in DGDG. In leaves, palmitate at the sn-2 position was desaturated in MGDG but not in DGDG. In isolated chloroplasts, palmitate at sn-2 similarly was desaturated only in MGDG, but palmitate and oleate at the sn-1 position were desaturated in MGDG as well as in DGDG. Apparently, palmitate desaturase reacts with sn-1 palmitate in either galactolipid, but does not react with the sn-2 fatty acid of DGDG. These results demonstrate that isolated spinach chloroplasts can synthesize and desaturate prokaryotic MGDG and DGDG. The finally accumulating molecular species, MGDG(18:3/16:3) and DGDG(18:3/16:0), are made by the chloroplasts in proportions similar to those found in leaves.     

On the metabolic relationships between monogalactosyldiacylglycerol and digalactosyldiacylglycerol molecular species in Dunaliella salina

Dunaliella salina cells were pulse-labeled for 2 min with [14C]palmitic acid, [14C]oleic acid, or [14C]lauric acid in order to trace the pathway of galactolipid biosynthesis and desaturation. Through the use of high performance liquid chromatography it was possible to follow the movement of radioactivity through many individual molecular species of monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) for periods of 24 h and, in some cases, as much as 120 h. Analysis of the fatty acid fluxes permitted us to refine current views regarding biosynthesis of the predominantly "prokaryotic" galactolipids. The initial D. salina MGDG molecular species, containing paired oleate and palmitate (18:1/16:0), can follow two metabolic routes. If the palmitoyl chain is desaturated to 16:1, the resulting 18:1/16:1 MGDG is subject to rapid further desaturation to varying degrees, and a part of these products is subsequently galactosylated to DGDG. Contrary to widely held opinions, these DGDG molecular species can themselves be further desaturated toward a 18:3/16:4 final product. In a separate series of reactions, a smaller portion of the nascent 18:1/16:0 MGDG is directly galactosylated to 18:1/16:0 DGDG. This molecular species can then be sequentially desaturated to 18:2/16:0 DGDG and 18:3/16:0 DGDG. However, there is only very limited desaturation of the palmitoyl group attached to these molecular species.     

Lipid profiling of Synechococcus sp. PCC7002 and two related strains by HPLC coupled to ESI-(ion trap)-MS/MS

The lipid profiles of Synechococcus sp. PCC7002 and two related 16S rDNA (99% identity) strains were established by a new method of high-performance liquid chromatography coupled to electrospray-mass spectrometry (HPLC-MS). Lipids were analysed in the positive and negative ionization mode, and fragmentation patterns are reported. No differences in the lipid profile between the three strains could be observed, but the relative content of some species differed. Major lipid species were found to be 1-octadecatrienoyl-2-hexadecanoyl-3-(6'-sulfo-alpha-D-quinovosyl)-sn-glycerol [SQDG (18:3/16:0)] and 1-octadecatrienoyl-2-hexadecenoyl-3-beta-D-monogalactosyl-sn-glycerol [MGDG (18:3/16:1)]. Ten species of SQDG, six species of PG (phosphatidyl-glycerol), seven species of MGDG, and two species of DGDG (digalactosyl-diacyl-glycerol) were detected. A PG species (m/z 761) containing hydroxylinolenic acid or oxophytodienoic acid acyl ester (C18H32O3), and SQDG species containing C17:1 and C17:3 fatty acyl esters are reported for the first time in cyanobacteria. The method also allowed the separation of two pairs of closely related isobaric MGDG species (m/z 770 and m/z 772 in positive ionization).     

Radiolabelling Studies on the Lipid Metabolism in the Marine Brown Alga Dictyopteris membranacea

The lipid metabolism of the marine brown alga D. membranaceawas investigated using [2–¹⁴C]acetate, [1–¹⁴C]myristate,[l–^I4C]oleate and [l–¹⁴C]arachidonate as precursors.On incubation with [2–¹⁴C]acetate, 18:1 and 16:0 werethe main products formed by de novo synthesis and incorporatedinto polar lipids. With all the exogenous substrates used, DGTAwas strongly labelled and the subsequent rapid turnover of radioactivitysuggested a key role for this lipid in the redistribution ofacyl chains and most likely also in the biosynthesis of theeukaryotic galacto-lipids produced in the absence of PC. Inthe glycolipids a continuous accumulation of radioactivity wasobserved with all the substrates used. The labelling kineticsof molecular species of MGDG suggested the desaturation of 18:1to 18:4 and of 20:4 (n-6) to 20:5 (n–3) acids on thislipid. Both PG and PE were primary acceptors of de novo synthesizedfatty acids and exogenous [l–¹⁴C]oleate, but no evidenceexists for a further processing of acyl chains on these lipids.TAG, although strongly labelled with all exogenous [l–¹⁴CJacids,was not labelled when [2–¹⁴C]acetate was used as a precursorindicating the flux of endogenous fatty acids to be differentof that of exogenously supplied fatty acids. (Received November 4, 1997; Accepted February 23, 1998)     

Comparison of glycerolipid biosynthesis in non-green plastids from sycamore (Acer pseudoplatanus) cells and cauliflower (Brassica oleracea) buds.

The availability of methods to fractionate non-green plastids and to prepare their limiting envelope membranes [Alban, Joyard & Douce (1988) Plant Physiol. 88, 709-717] allowed a detailed analysis of the biosynthesis of lysophosphatidic acid, phosphatidic acid, diacylglycerol and monogalactosyl-diacylglycerol (MGDG) in two different types of non-green starch-containing plastids: plastids isolated from cauliflower buds and amyloplasts isolated from sycamore cells. An enzyme [acyl-ACP (acyl carrier protein):sn-glycerol 3-phosphate acyltransferase) recovered in the soluble fraction of non-green plastids transfers oleic acid from oleoyl-ACP to the sn-1 position of sn-glycerol 3-phosphate to form lysophosphatidic acid. Then a membrane-bound enzyme (acyl-ACP:monoacyl-sn-glycerol 3-phosphate acyltransferase), localized in the envelope membrane, catalyses the acylation of the available sn-2 position of 1-oleoyl-sn-glycerol 3-phosphate by palmitic acid from palmitoyl-ACP. Therefore both the soluble phase and the envelope membranes are necessary for acylation of sn-glycerol 3-phosphate. The major difference between cauliflower (Brassica oleracea) and sycamore (Acer pseudoplatanus) membranes is the very low level of phosphatidate phosphatase activity in sycamore envelope membrane. Therefore, very little diacylglycerol is available for MGDG synthesis in sycamore, compared with cauliflower. These findings are consistent with the similarities and differences described in lipid metabolism of mature chloroplasts from 'C18:3' and 'C16:3' plants (those with MGDG containing C18:3 and C16:3 fatty acids). Sycamore contains only C18 fatty acids in MGDG, and the envelope membranes from sycamore amyloplasts have a low phosphatidate phosphatase activity and therefore the enzymes of the Kornberg-Pricer pathway have a low efficiency of incorporation of sn-glycerol 3-phosphate into MGDG. By contrast, cauliflower contains MGDG with C16:3 fatty acid, and the incorporation of sn-glycerol 3-phosphate into MGDG by the enzymes associated with envelope membranes is not limited by the phosphatidate phosphatase. These results demonstrate that: (1) non-green plastids employ the same biosynthetic pathway as that previously established for chloroplasts (the formation of glycerolipids is a general property of all plastids, chloroplasts as well as non-green plastids), (2) the envelope membranes are the major structure responsible for the biosynthesis of phosphatidic acid, diacylglycerol and MGDG, and (3) the enzymes of the envelope Kornberg-Pricer pathway have the same properties in non-green starch-containing plastids as in mature chloroplasts from C16:3 and C18:3 plants.     

Radiolabelling Studies of Lipids in the Marine Cryptomonad Chroomonas salina in Relation to Fatty Acid Desaturation

Cells of Chroomonas salina were exposed to [¹⁴C]acetate, [^l4C]16:0,[¹⁴C]18:0, [¹⁴C]18:1(n-9), [¹⁴C]18:2(n–6) or [¹⁴C]18:3(n–3)for 1 h and then incubated for 24 h in non-radioactive medium.At the end of the pulse period, non-glycolipid polar lipidscontained the highest proportions of radioactivity incorporatedfrom [¹⁴C]acetate and [¹⁴C]18:3(n–3) whereas with [¹⁴C]16:0,[¹⁴C]18:1 and [¹⁴C]18:2(n–6), triacylglycerols were mosthighly labelled. ¹⁴C-18:0 was recovered mainly as non-esterifiedfatty acid. Monogalactosyldiacylglycerol initially contained17% of label incorporated from [¹⁴C]acetate but less than 3%of that from [¹⁴C]fatty acids. With all substrates, excluding[¹⁴C]18:0, a gradual transfer of label from polar lipids totriacylglycerols was observed during the chase period. Saturatesand monoenes synthesised from [¹⁴C]acetate were mostly transferedfrom phospholipids and glycolipids to neutral lipid withoutfurther desaturation. Most of the incorporated ¹⁴C-fatty acidsremained unchanged and only with [¹⁴C]18:3(n–3) was substantialamounts of label recovered in penta- and hexaenoic fatty acids.The results indicate that, under the conditions of the study,lipid synthesis in the algae was heavily dominated by triacylglycerolformation and that the mechanisms of fatty acid desaturationin this species may differ from those in higher plants. (Received December 10, 1991; Accepted March 6, 1992)     

Effect of Growth Temperature on the Biosynthesis of Chloroplastic Galactosyldiacylglycerol Molecular Species in Brassica napus Leaves

Brassica napus leaves developed at low temperature display rapid in situ desaturation of monogalactosyldiacylglycerol (MGDG) fatty acids leading to the production of hexadecatrienoic/linolenic acid. This was shown by radioactivity-tracer experiments to occur via a sequence of desaturations proceeding from the initially synthesized palmitic/oleic acid molecular species to palmitic/linoleic acid, palmitoleic/linoleic acid, hexadecadienoic/linoleic acid, hexadecadienoic/linolenic acid, and finally to hexadecatrienoic/linolenic acid. The results suggest that there is increased activity in all five desaturation steps in leaves developed at low temperatures. Labeling data also indicate that there is another pool of MGDG which is more slowly desaturated before galactosylation to digalactosyldiacylglycerol (DGDG). Our data further suggest that relative rates of galactosylation of chloroplastic and cytosolic MGDG molecular species may regulate the final amounts of chloroplastic and cytosolic MGDG and DGDG in the leaf. We have proposed a model for chloroplastic biosynthesis and desaturation of galactosyldiacylglycerols in the leaves of Brassica napus, a 16:3 plant.     

The Effect of Light Intensity, Day Length, and Temperature on Fatty Acid Synthesis and Desaturation in Vicia faba L.

Some effects of light intensity, day length, and temperatureon the fatty acid composition of the major glycerolipids ofleaves of Vicia faba L. (cv. Giant Windsor) were observed. Increasinglight intensity caused an increase in the relative concentrationsof 16 : 1 in PG and 18 : 3 in MGDG and DGDG. Increasing daylength during growth (and continuous illumination of leaf tissue)had no effect on 16 : 1 in PG but caused a decrease in the 18: 3 content of PG, PC, MGDG, and DGDG. Since the quantitiesof these lipids increased under these conditions, the decreasewas not due to photodestruction but to the differences in therelative rates of biosynthesis and desaturation of fatty acids.Incubation of leaf tissue in the dark for 4 d had little effecton the fatty acid composition of MGDG, DGDG, and PG. Temperaturealso controls fatty acid synthesis and desaturation. Above theoptimum growth temperature (20 °C), the 18 : 3 content ofMGDG, DGDG, PG, and PC decreased. In mature leaf tissue, thedegree of unsaturation of MGDG may be modified upward in responseto temperature changes. When plants were grown at 30 °Cand transferred to 20 °C the level of 18 : 3 in MGDG ofthe leaf tissue increased to levels found in plants grown onlyat 20 °C. The level of 18 : 3 in MGDG does not decreaseas rapidly when plants grown at 20 °C were transferred to30 °C. This suggests that the lower temperature induceddesaturation of 18 : 2 to 18 : 3.     

Fatty acids of monogalactosyl diglycerides from Citrus

Fatty acids in vesicular and leaf monogalactosyl diglycerides (MGDG) of citrus were studied. Vesicular MGDG contained front 94.4 to 97.3% C₁₆, C_16:1, C_18:1, C_18:2, and C_18:3; whereas leaf MGDG contained ca 90% C_18:3, 3% C₁₆ and 1.8 to 9.5% C_18:2. Species varied considerably in their percentages of vesicular C_18:2, C1_8:3 and to a lesser degree, C_16:1 and C_18:1 fatty acids with lemons being the most distinctive. Branched fatty acids were present to the extent of 5.6% in vesicular and to only 0.1% in leaf MGDG.     

Glycerolipid synthesis in Chlorella kessleri 11h. I. Existence of a eukaryotic pathway

The fatty acid distributions at the sn-1 and sn-2 positions in major chloroplast lipids of Chlorella kessleri 11h, monogalactosyl diacylglycerol (MGDG) and digalactosyl diacylglycerol (DGDG), were determined to show the coexistence of both C16 and C18 acids at the sn-2 position, i.e. of prokaryotic and eukaryotic types in these galactolipids. For investigation of the biosynthetic pathway for glycerolipids in C. kessleri 11h, cells were fed with [14C]acetate for 30 min, and then the distribution of the radioactivity among glycerolipids and their constituent fatty acids during the subsequent chase period was determined. MGDG and DGDG were labeled predominantly as the sn-1-C18-sn-2-C16 (C18/C16) species as early as by the start of the chase, which suggested the synthesis of these lipids within chloroplasts via a prokaryotic pathway. On the other hand, the sn-1-C18-sn-2-C18 (C18/C18) species of these galactolipids gradually gained radioactivity at later times, concomitant with a decrease in the radioactivity of the C18/C18 species of phosphatidylcholine (PC). The change at later times can be explained by the conversion of the C18/C18 species of PC into galactolipids through a eukaryotic pathway. The results showed that C. kessleri 11h, distinct from most of other green algal species that were postulated mainly to use a prokaryotic pathway for the synthesis of chloroplast lipids, is similar to a group of higher plants designated as 16:3 plants in terms of the cooperation of prokaryotic and eukaryotic pathways to synthesize chloroplast lipids. We propose that the physiological function of the eukaryotic pathway in C. kessleri 11h is to supply chloroplast membranes with 18:3/18:3-MGDG for their functioning, and that the acquisition of a eukaryotic pathway by green algae was favorable for evolution into land plants.     

Fatty-acid composition of polar lipids in fruit and leaf chloroplasts of “16:3”- and “18:3”-plant species

The fatty-acid composition of polar lipids from fruit and leaf chloroplasts was compared in five Solanaceous and two cucurbit species. The acylated fatty acids in monogalactosyl diglycerides (MGDG) from leaf chloroplasts of all five Solanaceous species included substantial amounts of ^7,10,13-hexadecatrienoic acid (16:3). In contrast, the MGDG from fruit chloroplasts of the Solanaceae contained very little of this plastid-specific polyunsaturate, and instead included a proportionately greater percentage of linoleic acid (18:2). In MGDG from leaf chloroplasts of two cucurbits, -linolenic acid (18:3) constituted 94–95% of the acylated fatty acids. Fruit-chloroplast galactolipids of the cucurbits had a greater abundance of 18:2, and hence a higher 18:2/18:3 ratio, than found in the corresponding leaf lipids. Among the phosphoglycerides, the unusual fatty acid 3-trans-hexadecenoate (trans-16:1) constituted from 15 to 24% of the acylated fatty acids in phosphatidyl glycerol (PG) from leaf chloroplasts (all species). In sharp contrast, trans-16:1 was virtually absent in PG from fruit chloroplasts of both Solanaceous and cucurbit species, and was replaced by a proportionate increase in the content of palmitate (16:0). The observed differences in the polar lipid fatty-acid composition of fruit and leaf chloroplasts are discussed in terms of the relative activity of several intrachloroplastic enzymes involved in lipid synthesis and fatty-acyl desaturation.Abbreviations MGDG monogalactosyldiglyceride - DGDG digalactosyl diglyceride - PC phosphatidyl choline - PE phosphatidyl ethanolamine - PG phosphatidyl glycerol     

Arachidonic acid is important for efficient use of light by the microalga Lobosphaera incisa under chilling stress

The oleaginous microalga Lobosphaera incisa (Trebouxiophyceae, Chlorophyta) contains arachidonic acid (ARA, 20:4 n − 6) in all membrane glycerolipids and in the storage lipid triacylglycerol. The optimal growth temperature of the wild-type (WT) strain is 25 °C; chilling temperatures (≤ 15 °C) slow its growth. This effect is more pronounced in the delta-5-desaturase ARA-deficient mutant P127, in which ARA is replaced with dihomo-γ-linolenic acid (DGLA, 20:3 n − 6). In nutrient-replete cells grown at 25 °C, the major chloroplast lipid monogalactosylglycerol (MGDG) was dominated by C18/C16 species in both strains. Yet ARA constituted over 10% of the total fatty acids in the WT MGDG as a component of C20/C18 and C20/C20 species, whereas DGLA was only a minor component of MGDG in P127. Both strains increased the percentage of 18:3 n − 3 in membrane lipids under chilling temperatures. The temperature downshift led to a dramatic increase in triacylglycerol at the expense of chloroplast lipids. WT and P127 showed a similarly high photochemical quantum yield of photosystem II, whereas non-photochemical quenching (NPQ) and violaxanthin de-epoxidation were drastically higher in P127, especially at 15 °C. Fluorescence anisotropy measurements indicated that ARA-containing MGDG might contribute to sustaining chloroplast membrane fluidity upon dropping to the chilling temperature. We hypothesize that conformational changes in chloroplast membranes and increased rigidity of the ARA-deficient MGDG of P127 at chilling temperatures are not compensated by trienoic fatty acids. This might ‘lock’ violaxanthin de-epoxidase in the activated state causing high constitutive NPQ and alleviate the risk of photodamage under chilling conditions in the mutant.