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Upon abiotic stress or senescence, the size and/or abundance of plastid-localized plastoglobules and cytosolic lipid droplets, both compartments devoted to neutral lipid storage, increase in leaves. Meanwhile, plant lipid metabolism is also perturbed, notably with the degradation of thylakoidal monogalactosyldiacylglycerol (MGDG) and the accumulation of neutral lipids. Although these mechanisms are probably linked, they have never been jointly studied, and the respective roles of plastoglobules and lipid droplets in the plant response to stress are totally unknown. To address this question, we determined and compared the glycerolipid composition of both lipid droplets and plastoglobules, followed their formation in response to nitrogen starvation, and studied the kinetics of lipid metabolism in Arabidopsis leaves. Our results demonstrated that plastoglobules preferentially store phytyl-esters, while triacylglycerols (TAGs) and steryl-esters accumulated within lipid droplets. Thanks to a pulse-chase labeling approach and lipid analyses of the fatty acid desaturase 2 (fad2) mutant, we showed that MGDG-derived C18:3 fatty acids were exported to lipid droplets, while MGDG-derived C16:3 fatty acids were stored within plastoglobules. The export of lipids from plastids to lipid droplets was probably facilitated by the physical contact occurring between both organelles, as demonstrated by our electron tomography study. The accumulation of lipid droplets and neutral lipids was transient, suggesting that stress-induced TAGs were remobilized during the plant recovery phase by a mechanism that remains to be explored., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Key Message: Assembly of PUFA-attached TAGs is intimately correlated to turnover of newly formed membrane lipids in starch-deficient Chlamydomonas exposed to high light and nitrogen stress under air-aerated mixotrophic conditions. Triacylglycerols (TAGs) rich in polyunsaturated fatty acids (PUFAs) in microalgae have attracted extensive attention due to its promising application in nutraceuticals and other high-value compounds. Previous studies revealed that PUFAs accumulated in TAG primarily derived from the dominant membrane lipids, monogalactosyldiacylglycerolipid, digalactosyldiacylglycerol and diacylglycerol-N,N,N-trimethylhomoserine (DGTS), in the model alga Chlamydomonas reinhardtii. However, their respective contribution to PUFA-attached TAG integration has not been clearly deciphered, particularly in starchless Chlamydomonas that hyper-accumulates TAG. In this study, the starchless C. reinhardtii BAFJ5 was mixotrophically cultivated in photobioreactors aerated with air (0.04% CO 2 ), and we monitored the dynamic changes in growth, cellular carbon and nitrogen content, photosynthetic activity, biochemical compositions, and glycerolipid remodeling under high light and nitrogen starvation conditions. The results indicated that multiple PUFAs continually accumulated in total lipids and TAG, and the primary distributors of these PUFAs gradually shifted from membrane lipids to TAG in stress-induced BAFJ5. The stoichiometry analyses showed that the PUFA-attached TAG assembly attributed to turnover of not only the major glycerolipids, but also the phospholipids, phosphatidylethanolamine (PE) and phosphatidylglycerol. Specifically, the augmented C16:3n3 and C18:3n3 in TAG mainly originated from de novo-synthesized galactolipids, while the cumulative C18:3n6 and C18:4n3 in TAG were intimately correlated with conversion of the newly formed DGTS and PE. These findings emphasized significance of PUFA-attached TAG formation dependent on turnover of de novo assembled membrane lipids in starch-deficient Chlamydomonas, beneficial for enhanced production of value-added lipids in microalgae., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Alka(e)nes are produced by many living organisms and exhibit diverse physiological roles, reflecting a high functional versatility. Alka(e)nes serve as waterproof wax in plants, communicating pheromones for insects, and microbial signaling molecules in some bacteria. Although alka(e)nes have been found in cyanobacteria and algal chloroplasts, their importance for photosynthetic membranes has remained elusive. In this study, we investigated the consequences of the absence of alka(e)nes on membrane lipid composition and photosynthesis using the cyanobacterium Synechocystis PCC6803 as a model organism. By following the dynamics of membrane lipids and the photosynthetic performance in strains defected and altered in alka(e)ne biosynthesis, we show that drastic changes in the glycerolipid contents occur in the absence of alka(e)nes, including a decrease in the membrane carotenoid content, a decrease in some digalactosyldiacylglycerol (DGDG) species and a parallel increase in monogalactosyldiacylglycerol (MGDG) species. These changes are associated with a higher susceptibility of photosynthesis and growth to high light in alka(e)ne-deficient strains. All these phenotypes are reversed by expressing an algal photoenzyme producing alka(e)nes from fatty acids. Therefore, alkenes, despite their low abundance, are an essential component of the lipid composition of membranes. The profound remodeling of lipid composition that results from their absence suggests that they play an important role in one or more membrane properties in cyanobacteria. Moreover, the lipid compensatory mechanism observed is not sufficient to restore normal functioning of the photosynthetic membranes, particularly under high-light intensity. We conclude that alka(e)nes play a crucial role in maintaining the lipid homeostasis of thylakoid membranes, thereby contributing to the proper functioning of photosynthesis, particularly under elevated light intensities., Competing Interests: Conflict of interest statement. The authors declare that there is no potential conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Chlorella sorokiniana , isolated from a pond adjacent to a cement plant, was cultured using flue gas collected directly from kiln emissions using 20 L and 25000 L photobioreactors. Lipids, proteins, and polysaccharides were analyzed to understand their overall composition for potential applications. The lipid content ranged from 17.97% to 21.54% of the dry biomass, with carotenoid concentrations between 8.4 and 9.2 mg/g. Lutein accounted for 55% of the total carotenoids. LC/MS analysis led to the identification of 71 intact triacylglycerols, 8 lysophosphatidylcholines, 10 phosphatidylcholines, 9 monogalactosyldiacylglycerols, 12 digalactosyldiacylglycerols, and 1 sulfoquinovosyl diacylglycerol. Palmitic acid, oleic acid, linoleic acid, and α-linolenic acid were the main fatty acids. Polyunsaturated fatty acid covers ≥ 56% of total fatty acids. Protein isolates and polysaccharides were also extracted. Protein purity was determined to be ≥75% by amino acid analysis, with all essential amino acids present. Monomer analysis of polysaccharides suggested that they are composed of mainly D-(+)-mannose, D-(+)-galactose, and D-(+)-glucose. The results demonstrate that there is no adverse effect on the metabolite profile of C. sorokiniana biomass cultured using flue gas as the primary carbon source, revealing the possibility of utilizing such algal biomass in industrial applications such as animal feed, sources of cosmeceuticals, and as biofuel.

The plant endoplasmic reticulum (ER) contacts heterotypic membranes at membrane contact sites (MCSs) through largely undefined mechanisms. For instance, despite the well-established and essential role of the plant ER-chloroplast interactions for lipid biosynthesis, and the reported existence of physical contacts between these organelles, almost nothing is known about the ER-chloroplast MCS identity. Here we show that the Arabidopsis ER membrane-associated VAP27 proteins and the lipid-binding protein ORP2A define a functional complex at the ER-chloroplast MCSs. Specifically, through in vivo and in vitro association assays, we found that VAP27 proteins interact with the outer envelope membrane (OEM) of chloroplasts, where they bind to ORP2A. Through lipidomic analyses, we established that VAP27 proteins and ORP2A directly interact with the chloroplast OEM monogalactosyldiacylglycerol (MGDG), and we demonstrated that the loss of the VAP27-ORP2A complex is accompanied by subtle changes in the acyl composition of MGDG and PG. We also found that ORP2A interacts with phytosterols and established that the loss of the VAP27-ORP2A complex alters sterol levels in chloroplasts. We propose that, by interacting directly with OEM lipids, the VAP27-ORP2A complex defines plant-unique MCSs that bridge ER and chloroplasts and are involved in chloroplast lipid homeostasis., (© 2024. The Author(s).)

Isotope labeling coupled with mass spectrometry imaging (MSI) presents a potent strategy for elucidating the dynamics of metabolism at cellular resolution, yet its application to plant systems is scarce. It has the potential to reveal the spatio-temporal dynamics of lipid biosynthesis during plant development. In this study, we explore its application to galactolipid biosynthesis of an aquatic plant, Lemna minor, with D2O labeling. Specifically, matrix-assisted laser desorption/ionization-MSI data of two major galactolipids in L. minor, monogalactosyldiacylglycerol and digalactosyldiacylglycerol, were studied after growing in 50% D2O media over a 15-day time period. When they were partially labeled after 5 d, three distinct binomial isotopologue distributions were observed corresponding to the labeling of partial structural moieties: galactose only, galactose and a fatty acyl chain and the entire molecule. The temporal change in the relative abundance of these distributions follows the expected linear pathway of galactolipid biosynthesis. Notably, their mass spectrometry images revealed the localization of each isotopologue group to the old parent frond, the intermediate tissues and the newly grown daughter fronds. Besides, two additional labeling experiments, (i) 13CO2 labeling and (ii) backward labeling of completely 50% D2O-labeled L. minor in H2O media, confirm the observations in forward labeling. Furthermore, these experiments unveiled hidden isotopologue distributions indicative of membrane lipid restructuring. This study suggests the potential of isotope labeling using MSI to provide spatio-temporal details in lipid biosynthesis in plant development., (© The Author(s) 2024. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Monogalactosyldiacylglycerol (MGDG) is the main lipid constituent of thylakoids and a structural component of photosystems and photosynthesis-related proteo-lipid complexes in green tissues. Previously reported changes in MGDG abundance upon stress treatments are hypothesized to reflect mobilization of MGDG-based polyunsaturated lipid intermediates to maintain extraplastidial membrane integrity. While exchange of lipid intermediates between compartmental membranes is well documented, physiological consequences of mobilizing an essential thylakoid lipid, such as MGDG, for an alternative purpose are not well understood. Arabidopsis seedlings exposed to mild (50 mM) salt treatment displayed significantly increased abundance of both MGDG and the extraplastidial lipid, phosphatidylcholine (PC). Interestingly, similar increases in MGDG and PC were observed in Arabidopsis fad3 mutant seedlings defective in endoplasmic reticulum (ER)-localized linolenic acid formation, in which compensatory plastid-to-ER-directed mobilization of linolenic acid-containing intermediates takes place. The postulated (salt) or evident (fad3) plastid-ER exchange of intermediates concurred with altered thylakoid function according to parameters of photosynthetic performance. While salt treatment of wild-type seedlings inhibited photosynthetic parameters in a dose-dependent manner, interestingly, untreated fad3 mutants did not show overall reduced photosynthetic quantum yield. By contrast, we observed a reduction specifically of non-photochemical quenching (NPQ) under high light, representing only part of observed salt effects. The decreased NPQ in the fad3 mutant was accompanied by reduced activity of the xanthophyll cycle, leading to a reduced concentration of the NPQ-effective pigment zeaxanthin. The findings suggest that altered ER-located fatty acid unsaturation and ensuing inter-organellar compensation impacts on the function of specific thylakoid enzymes, rather than globally affecting thylakoid function., (© The Author(s) 2023. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The chloroplast thylakoid membrane is composed of membrane lipids and photosynthetic protein complexes, and the orchestration of thylakoid lipid biosynthesis and photosynthesis-associated protein accumulation is considered important for thylakoid development. Galactolipids consist of ∼80% of the thylakoid lipids, and their biosynthesis is fundamental for chloroplast development. We previously reported that the suppression of galactolipid biosynthesis decreased the expression of photosynthesis-associated nuclear-encoded genes (PhAPGs) and photosynthesis-associated plastid-encoded genes (PhAPGs). However, the mechanism for coordinative regulation between galactolipid biosynthesis in plastids and the expression of PhANGs and PhAPGs remains largely unknown. To elucidate this mechanism, we investigated the gene expression patterns in galactolipid-deficient Arabidopsis seedlings during the de-etiolation process. We found that galactolipids are crucial for inducing both the transcript accumulation of PhANGs and PhAPGs and the accumulation of plastid-encoded photosynthesis-associated proteins in developing chloroplasts. Genetic analysis indicates the contribution of the GENOMES UNCOUPLED1 (GUN1)-mediated plastid-to-nucleus signaling pathway to PhANG regulation in response to galactolipid levels. Previous studies suggested that the accumulation of GUN1 reflects the state of protein homeostasis in plastids and alters the PhANG expression level. Thus, we propose a model that galactolipid biosynthesis determines the protein homeostasis in plastids in the initial phase of de-etiolation and optimizes GUN1-dependent signaling to regulate the PhANG expression. This mechanism might contribute to orchestrating the biosynthesis of lipids and proteins for the biogenesis of functional chloroplasts in plants., (© The Author(s) 2024. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Galactolipids comprise the majority of chloroplast membranes in plants, and their biosynthesis requires dephosphorylation of phosphatidic acid at the chloroplast envelope membranes. In Arabidopsis (Arabidopsis thaliana), the lipid phosphate phosphatases LPPγ, LPPε1, and LPPε2 have been previously implicated in chloroplast lipid assembly, with LPPγ being essential, as null mutants were reported to exhibit embryo lethality. Here, we show that lppγ mutants are in fact viable and that LPPγ, LPPε1, and LPPε2 do not appear to have central roles in the plastid pathway of membrane lipid biosynthesis. Redundant LPPγ and LPPε1 activity at the outer envelope membrane is important for plant development, and the respective lppγ lppε1 double mutant exhibits reduced flux through the ER pathway of galactolipid synthesis. While LPPε2 is imported and associated with interior chloroplast membranes, its role remains elusive and does not include basal nor phosphate limitation-induced biosynthesis of glycolipids. The specific physiological roles of LPPγ, LPPε1, and LPPε2 are yet to be uncovered, as does the identity of the phosphatidic acid phosphatase required for plastid galactolipid biosynthesis., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

The ability to harvest light effectively in a changing environment is necessary to ensure efficient photosynthesis and crop growth. One mechanism, known as qE, protects photosystem II (PSII) and regulates electron transfer through the harmless dissipation of excess absorbed photons as heat. This process involves reversible clustering of the major light-harvesting complexes of PSII (LHCII) in the thylakoid membrane and relies upon the ΔpH gradient and the allosteric modulator protein PsbS. To date, the exact role of PsbS in the qE mechanism has remained elusive. Here, we show that PsbS induces hydrophobic mismatch in the thylakoid membrane through dynamic rearrangement of lipids around LHCII leading to observed membrane thinning. We found that upon illumination, the thylakoid membrane reversibly shrinks from around 4.3 to 3.2 nm, without PsbS, this response is eliminated. Furthermore, we show that the lipid digalactosyldiacylglycerol (DGDG) is repelled from the LHCII-PsbS complex due to an increase in both the p K a of lumenal residues and in the dipole moment of LHCII, which allows for further conformational change and clustering in the membrane. Our results suggest a mechanistic role for PsbS as a facilitator of a hydrophobic mismatch-mediated phase transition between LHCII-PsbS and its environment. This could act as the driving force to sort LHCII into photoprotective nanodomains in the thylakoid membrane. This work shows an example of the key role of the hydrophobic mismatch process in regulating membrane protein function in plants.

The use of Nuclear Magnetic Resonance spectroscopy for studying lipid digestion in vitro most often consists of quantifying lipolysis products after they have been extracted from the reaction medium using organic solvents. However, the current sensitivity level of NMR spectrometers makes possible to avoid the extraction step and continuously quantify the lipids directly in the reaction medium. We used real-time 1 H NMR spectroscopy and guinea pig pancreatic lipase-related protein 2 (GPLRP2) as biocatalyst to monitor in situ the lipolysis of monogalactosyl diacylglycerol (MGDG) in the form of mixed micelles with the bile salt sodium taurodeoxycholate (NaTDC). Residual substrate and lipolysis products (monogalactosyl monoacylglycerol (MGMG); monogalactosylglycerol (MGG) and octanoic acid (OA) were simultaneously quantified throughout the reaction thanks to specific proton resonances. Lipolysis was complete with the release of all MGDG fatty acids. These results were confirmed by thin layer chromatography (TLC) and densitometry after lipid extraction at different reaction times. Using diffusion-ordered NMR spectroscopy (DOSY), we could also estimate the diffusion coefficients of all the reaction compounds and deduce the hydrodynamic radius of the lipid aggregates in which they were present. It was shown that MGDG-NaTDC mixed micelles with an initial hydrodynamic radius r H of 7.3 ± 0.5 nm were changed into smaller micelles of NaTDC-MGDG-MGMG of 2.3 ± 0.5 nm in the course of the lipolysis reaction, and finally into NaTDC-OA mixed micelles (r H of 2.9 ± 0.5 nm) and water soluble MGG. These results provide a better understanding of the digestion of galactolipids by PLRP2, a process that leads to the complete micellar solubilisation of their fatty acids and renders their intestinal absorption possible., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Glycerol-3-phosphate acyltransferase GPAT9 catalyzes the first acylation of glycerol-3-phosphate (G3P), a committed step of glycerolipid synthesis in Arabidopsis . The role of GPAT9 in Brassica napus remains to be elucidated. Here, we identified four orthologs of GPAT9 and found that BnaGPAT9 encoded by BnaC01T0014600WE is a predominant isoform and promotes seed oil accumulation and eukaryotic galactolipid synthesis in Brassica napus . BnaGPAT9 is highly expressed in developing seeds and is localized in the endoplasmic reticulum (ER). Ectopic expression of BnaGPAT9 in E. coli and siliques of Brassica napus enhanced phosphatidic acid (PA) production. Overexpression of BnaGPAT9 enhanced seed oil accumulation resulting from increased 18:2-fatty acid. Lipid profiling in developing seeds showed that overexpression of BnaGPAT9 led to decreased phosphatidylcholine (PC) and a corresponding increase in phosphatidylethanolamine (PE), implying that BnaGPAT9 promotes PC flux to storage triacylglycerol (TAG). Furthermore, overexpression of BnaGPAT9 also enhanced eukaryotic galactolipids including monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), with increased 36:6-MGDG and 36:6-DGDG, and decreased 34:6-MGDG in developing seeds. Collectively, these results suggest that ER-localized BnaGPAT9 promotes PA production, thereby enhancing seed oil accumulation and eukaryotic galactolipid biosynthesis in Brassica napus .

Plastoglobules (PGs) contiguous with the outer leaflets of thylakoid membranes regulate lipid metabolism, plastid developmental transitions, and responses to environmental stimuli. However, the function of OsFBN7, a PG-core fibrillin gene in rice, has not been elucidated. Using molecular genetics and physiobiochemical approaches, we observed that OsFBN7 overexpression promoted PG clustering in rice chloroplasts. OsFBN7 interacted with two KAS I enzymes, namely OsKAS Ia and OsKAS Ib, in rice chloroplasts. Lipidomic analysis of chloroplast subcompartments, including PGs in the OsFBN7 overexpression lines, confirmed that levels of diacylglycerol (DAG), a chloroplast lipid precursor and monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), the main chloroplast membrane lipids, were increased in PGs and chloroplasts. Furthermore, OsFBN7 enhanced the abundances of OsKAS Ia/Ib in planta and their stability under oxidative and heat stresses. In addition, RNA sequencing and real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) analyses showed that the expression of the DAG synthetase gene PAP1 and MGDG synthase gene MDG2 was upregulated by OsFBN7. In conclusion, this study proposes a new model in which OsFBN7 binds to OsKAS Ia/Ib in chloroplast and enhances their abundance and stability, thereby regulating the chloroplast and PG membrane lipids involved in the formation of PG clusters., (© 2023 The Authors. New Phytologist © 2023 New Phytologist Foundation.)

Glycerolipids are the most abundant lipids in microalgae, and glycerol-3-phosphate:acyl-CoA acyltransferase (GPAT) plays an important role in their biosynthesis. However, the biochemical and biological functions of algal GPAT remain poorly characterized. Here, we characterized the endoplasmic reticulum (ER)-associated GPAT of the model unicellular green alga Chlamydomonas reinhardtii (CrGPATer). Enzymatic assays indicated that CrGPATer is an sn-1 acyltransferase using a variety of acyl-CoAs as the acyl donor. Subcellular localization revealed that CrGPATer was associated with ER membranes and lipid droplets. We constructed overexpression (OE) and knockdown (KD) transgenic C. reinhardtii lines to investigate the in vivo function of CrGPATer. Lipidomic analysis indicated that CrGPATer OE enhanced the cellular content of galactolipids, especially monogalactosyldiacylglycerol, under nitrogen deficiency stress. Correspondingly, CrGPATer KD lines contained lower contents of galactolipids than the control. Feeding experiments with labeled phosphatidic acid revealed that the intermediate of the eukaryotic Kennedy pathway could be used for galactolipid biosynthesis in the chloroplasts. These results provided multiple lines of evidence that the eukaryotic Kennedy pathway mediated by CrGPATer may be involved in galactolipid biosynthesis in C. reinhardtii. OE of CrGPATer significantly increased the content of triacylglycerol and the yield of biomass. Moreover, the content and yield of 1, 3-olein-2-palmitin, a high-value lipid that can be used as an alternative for human milk fat in infant formula, were significantly enhanced in the OE transgenic lines. Taken together, this study provided insights into the biochemical and biological functions of CrGPATer and its potential as a genetic engineering target in functional lipid manufacturing., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Galactolipids are the main lipids from plant photosynthetic membranes and they can be digested by pancreatic lipase related protein 2 (PLRP2), an enzyme found in the pancreatic secretion in many animal species. Here, we used transmission Fourier-transform infrared spectroscopy (FTIR) to monitor continuously the hydrolysis of galactolipids by PLRP2, in situ and in real time. The method was first developed with a model substrate, a synthetic monogalactosyl diacylglycerol with 8-carbon acyl chains (C8-MGDG), in the form of mixed micelles with a bile salt, sodium taurodeoxycholate (NaTDC). The concentrations of the residual substrate and reaction products (monogalactosylmonoglyceride, MGMG; monogalactosylglycerol, MGG; octanoic acid) were estimated from the carbonyl and carboxylate vibration bands after calibration with reference standards. The results were confirmed by thin layer chromatography analysis (TLC) and specific staining of galactosylated compounds with thymol and sulfuric acid. The method was then applied to the lipolysis of more complex substrates, a natural extract of MGDG with long acyl chains, micellized with NaTDC, and intact chloroplasts isolated from spinach leaves. After a calibration performed with α-linolenic acid, the main fatty acid (FA) found in plant galactolipids, FTIR allowed quantitative measurement of chloroplast lipolysis by PLRP2. A full release of FA from membrane galactolipids was observed, that was not dependent on the presence of bile salts. Nevertheless, the evolution of amide vibration band in FTIR spectra suggested the interaction of membrane proteins with NaTDC and lipolysis products., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The lipid composition of thylakoid membranes is conserved from cyanobacteria to green plants. However, the biosynthetic pathways of galactolipids, the major components of thylakoid membranes, are known to differ substantially between cyanobacteria and green plants. We previously reported on a transformant of the unicellular rod-shaped cyanobacterium Synechococcus elongatus PCC 7942, namely SeGPT, in which the synthesis pathways of the galactolipids monogalactosyldiacylglycerol and digalactosyldiacylglycerol are completely replaced by those of green plants. SeGPT exhibited increased galactolipid content and could grow photoautotrophically, but its growth rate was slower than that of wild-type S. elongatus PCC 7942. In the present study, we investigated pleiotropic effects that occur in SeGPT and determined how its increased lipid content affects cell proliferation. Microscopic observations revealed that cell division and thylakoid membrane development are impaired in SeGPT. Furthermore, physiological analyses indicated that the bioenergetic state of SeGPT is altered toward energy storage, as indicated by increased levels of intracellular ATP and glycogen. We hereby report that we have identified a new promising candidate as a platform for material production by modifying the lipid synthesis system in this way., (© 2023. The Author(s).)

Photosystem II (PSII) contains many lipid molecules that are essential for the function and maintenance of PSII. Under strong light conditions, PSII complexes are dynamically modified during the repair process; however, the molecular mechanism of the dynamic changes in the PSII structure is still unclear. In the present study, we investigated the role of a lipase in the repair of PSII in Synechocystis sp. PCC 6803. We identified a protein encoded by the sll1969 gene, previously named lipase A (lipA), in the Synechocystis sp. PCC 6803 genome as a candidate for the lipase involved in PSII repair. Recombinant protein expressed in Escherichia coli cells hydrolyzed fatty acids at the sn-1 position of monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol as well as triacylglycerol esterified with stearic acids. PSII repair in a disrupted mutant of the lipA gene was suppressed by the slow degradation of damaged D1 protein under strong light. The level of the PSII dimer remained higher in lipA mutant cells than wild-type (WT) cells under strong light. LipA protein was associated with the PSII dimer in vivo, and recombinant LipA protein decomposed PSII dimers purified from WT cells to monomers by reducing MGDG content in the PSII complex. These results indicate that LipA reacts with PSII dimers, dissociates them into monomers by digesting MGDG, and enhances D1 degradation during PSII repair., (© American Society of Plant Biologists 2022. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Most lipids in our diet come under the form of triacylglycerols that are often redispersed and stabilized by surfactants in processed foods. In plant however, lipid assemblies constitute interesting sources of natural bioactive and functional ingredients. In most photosynthetic sources, polar lipids rich in ω3 fatty acids are concentrated. The objective of this review is to summarize all the knowledge about the physico-chemical composition, digestive behavior and oxidative stability of plant polar lipid assemblies to emphasize their potential as functional ingredients in human diet and their potentialities to substitute artificial surfactants/antioxidants. The specific composition of plant membrane assemblies is detailed, including plasma membranes, oil bodies, and chloroplast; emphasizing its concentration in phospholipids, galactolipids, peculiar proteins, and phenolic compounds. These molecular species are hydrolyzed by specific digestive enzymes in the human gastrointestinal tract and reduced the hydrolysis of triacylglycerols and their subsequent absorption. Galactolipids specifically can activate ileal break and intrinsically present an antioxidant (AO) activity and metal chelating activity. In addition, their natural association with phenolic compounds and their physical state (Lα state of digalactosyldiacylglycerols) in membrane assemblies can enhance their stability to oxidation. All these elements make plant membrane molecules and assemblies very promising components with a wide range of potential applications to vectorize ω3 polyunsaturated fatty acids, and equilibrate human diet.

In previous research, xylem sap of angiosperms has been found to include low concentrations of nanoparticles and polar lipids. A major goal of this study was to test predictions arising from the hypothesis that the nanoparticles consist largely of polar lipids from the original cell content of vessel elements. These predictions included that polar lipid and nanoparticle concentrations would be correlated, that they both do not pass through pit membranes and that they do not vary seasonally because they originate from living vessel element cells. We collected xylem sap of six temperate angiosperm species over the whole year to consider seasonal variation. Concentrations of nanoparticles and lipids in xylem sap and contamination control samples were measured with a NanoSight device and mass spectrometry. We found that the concentration of nanoparticles and polar lipids was (i) diluted when an increasing amount of sap was extracted, (ii) significantly correlated to each other for three species, (iii) affected by vessel anatomy, (iv) very low and largely different in chemical composition from contamination controls and (v) hardly variable among seasons. Moreover, there was a minor freezing-thawing effect with respect to nanoparticle amount and size. Xylem sap lipids included polar galactolipids and phospholipids in all species and neutral triacylglycerols in two species. These findings support the predictions and, by implication, the underlying hypothesis that nanoparticles in xylem sap consist of polar lipids from the original cell content of living vessel element cells. Further research is needed to examine the formation and stability of nanoparticles concerning lipid composition and multiphase interactions among gas, liquid and solid phases in xylem conduits of living plants., (© The Author(s) 2022. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Digalactosyldiacylglycerol- (DGDG-) monoestolide is a characteristic glycolipid in oats. DGDG-monoestolides possess a unique structure whereby a fatty acid of DGDG is replaced by a fatty acid ester of hydroxy fatty acid (FAHFA). While the physiological effects of DGDG and FAHFA have been reported previously, the effects of DGDG-monoestolides are unknown. Hence, we isolated a major DGDG-monoestolide molecular species from oats, analyzed its structure, and evaluated its anti-inflammatory effect. Based on GC-MS, MS/MS, and NMR analyses, the isolated compound was identified as a DGDG-monoestolide that contains the linoleic acid ester of 15-hydroxy linoleic acid (LAHLA) and linoleic acid (i.e., DGDG-LAHLA). The isolated DGDG-LAHLA was evaluated for its anti-inflammatory effect on LPS-stimulated RAW264 cells. The production of nitric oxide and cytokines (IL-6, TNF-α, and IL-10) were significantly decreased by DGDG-LAHLA, suggesting the anti-inflammatory effect of DGDG-LAHLA for the first time. In addition, our data showed a pronounced uptake of DGDG-LAHLA by cells. Some compounds corresponding to the predicted DGDG-LAHLA metabolites were also detected, suggesting that both intact DGDG-LAHLA and its metabolites may contribute to the above anti-inflammatory activities. These results are expected to expand the availability of oats as a functional food.

Arabidopsis thaliana monogalactosyldiacylglycerol synthase 1 (AtMGD1) and digalactosyldiacylglycerol synthase 2 (AtDGD2) genes introduced into a Bacillus subtilis chromosome with disrupted galE, which encodes UDP-glucose 4-epi merase, enabled the mutant to produce monogalactosyldiacylglycerol. When galE mutant cells are cultivated in galactose containing medium they show ab normal morphology. This phenotype is correlated with a decrease in the amount of glucolipids. Nucleoids of the ugtP and galE mutants were stained by propidium iodide, which does not permeate intact cell membranes, whereas nucleoids of wild type and of a pssA mutant we examined were not stained. Expression of the AtMGD1 gene in a ugtP galE double mutant restored cell membrane integrity. Expression of galactolipid synthase genes from a multi-copy plasmid, pDGHisN4, allowed higher production of galactolipids. Activation of the extracytoplasmic function sigma factors SigM, SigV, and SigX, in the ugtP mutant was decreased by expression of AtMGD1, and SigX activity was strongly repressed when both AtMGD1 and AtDGD2 genes were expressed in the mutant. We conclude that the number of sugars that bind to diacylglycerol - rather than the exact sugar species - is important for glycolipid function in B. subtilis.

Senescence in plants enables resource recycling from senescent leaves to sink organs. Under stress, increased production of reactive oxygen species (ROS) and associated signalling activates senescence. However, senescence is not always associated with stress since it has a prominent role in plant development, in which the role of ROS signalling is less clear. To address this, we investigated lipid metabolism and patterns of lipid peroxidation related to signalling during sequential senescence in first-emerging barley leaves grown under natural light conditions. Leaf fatty acid compositions were dominated by linolenic acid (75% of total), the major polyunsaturated fatty acid (PUFA) in galactolipids of thylakoid membranes, known to be highly sensitive to peroxidation. Lipid catabolism during senescence, including increased lipoxygenase activity, led to decreased levels of PUFA and increased levels of short-chain saturated fatty acids. When normalised to leaf area, only concentrations of hexanal, a product from the 13-lipoxygenase pathway, increased early upon senescence, whereas reactive electrophile species (RES) from ROS-associated lipid peroxidation, such as 4-hydroxynonenal, 4-hydroxyhexenal and acrolein, as well as β-cyclocitral derived from oxidation of β-carotene, decreased. However, relative to total chlorophyll, amounts of most RES increased at late-senescence stages, alongside increased levels of α-tocopherol, zeaxanthin and non-photochemical quenching, an energy dissipative pathway that prevents ROS production. Overall, our results indicate that lipid peroxidation derived from enzymatic oxidation occurs early during senescence in first barley leaves, while ROS-derived lipid peroxidation associates weaker with senescence., (© 2022 The Authors. Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

The appearance of oxygenic photosynthesis in cyanobacteria is a major event in evolution. It had an irreversible impact on the Earth, promoting the Great Oxygenation Event (GOE) ~2.4 billion years ago. Ancient cyanobacteria predating the GOE were Gloeobacter-type cells lacking thylakoids, which hosted photosystems in their cytoplasmic membrane. The driver of the GOE was proposed to be the transition from unicellular to filamentous cyanobacteria. However, the appearance of thylakoids expanded the photosynthetic surface to such an extent that it introduced a multiplier effect, which would be more coherent with an impact on the atmosphere. Primitive thylakoids self-organize as concentric parietal uninterrupted multilayers. There is no robust evidence for an origin of thylakoids via a vesicular-based scenario. This review reports studies supporting that hexagonal II-forming glucolipids and galactolipids at the periphery of the cytosolic membrane could be turned, within nanoseconds and without any external source of energy, into membrane multilayers. Comparison of lipid biosynthetic pathways shows that ancient cyanobacteria contained only one anionic lamellar-forming lipid, phosphatidylglycerol. The acquisition of sulfoquinovosyldiacylglycerol biosynthesis correlates with thylakoid emergence, possibly enabling sufficient provision of anionic lipids to trigger a hexagonal II-to-lamellar phase transition. With this non-vesicular lipid-phase transition, a framework is also available to re-examine the role of companion proteins in thylakoid biogenesis., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Plants that are starved of phosphate trigger membrane lipid remodeling, which hydrolyses phospholipids and presumably allows their phosphate to be utilized, whilst replacing them with galactolipids to maintain the integrity of the membrane system. In addition to the two concurrent pathways of phospholipid hydrolysis by phospholipases C and D that have already been established, an emerging third pathway has been proposed that includes a reaction step catalysed by glycerophosphodiester phosphodiesterases (GDPDs). However, its functional involvement in phosphate-starved plants remains elusive. Here, we show that Arabidopsis GDPD6 is a functional isoform responsible for glycerophosphocholine hydrolysis in vivo. Overexpression of GDPD6 promoted root growth whilst gdpd6 mutants showed impaired root growth under phosphate starvation, and this defect was rescued by supplementing with the reaction product glycerol 3-phosphate but not with choline. Since GDPD6 is induced by phosphate starvation, we suggest that GDPD6 might be involved in root growth via the production of glycerol 3-phosphate in phosphate-starved plants., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Phosphorus (P) is an essential nutrient for plants. Membrane lipid remodeling is an adaptive mechanism for P-starved plants that replaces membrane phospholipids with non-P galactolipids, presumably to retrieve scarce P sources and maintain membrane integrity. Whereas metabolic pathways to convert phospholipids to galactolipids are well-established, the mechanism by which phospholipid biosynthesis is involved in this process remains elusive. Here, we report that phospho-base N-methyltransferases 1 and 2 (PMT1 and PMT2), which convert phosphoethanolamine to phosphocholine (PCho), are transcriptionally induced by P starvation. Shoots of seedlings of pmt1 pmt2 double mutant showed defective growth upon P starvation; however, membrane lipid profiles were unaffected. We found that P-starved pmt1 pmt2 with defective leaf growth had reduced PCho content, and the growth defect was rescued by exogenous supplementation of PCho. We propose that PMT1 and PMT2 are induced by P starvation to produce PCho mainly for leaf growth maintenance, rather than for phosphatidylcholine biosynthesis, in membrane lipid remodeling., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. 
For permissions, please email: journals.permissions@oup.com.)

Rhomboid-like proteins are intramembrane proteases with a variety of regulatory roles in cells. Though many rhomboid-like proteins are predicted in plants, their detailed molecular mechanisms or cellular functions are not yet known. Of the 13 predicted rhomboids in Arabidopsis thaliana, one, RBL10, affects lipid metabolism in the chloroplast, because in the respective rbl10 mutant the transfer of phosphatidic acid through the inner envelope membrane is disrupted. Here we show that RBL10 is part of a high-molecular-weight complex of 250 kDa or greater in size. Nine likely components of this complex are identified by two independent methods and include Acyl Carrier Protein 4 (ACP4) and Carboxyltransferase Interactor1 (CTI1), which have known roles in chloroplast lipid metabolism. The acp4 mutant has decreased C16:3 fatty acid content of monogalactosyldiacylglycerol, similar to the rbl10 mutant, prompting us to offer a mechanistic model of how an interaction between ACP4 and RBL10 might affect chloroplast lipid assembly. We also demonstrate the presence of a seventh transmembrane domain in RBL10, refining the currently accepted topology of this protein. Taken together, the identity of possible RBL10 complex components as well as insights into RBL10 topology and distribution in the membrane provide a stepping-stone towards a deeper understanding of RBL10 function in Arabidopsis lipid metabolism., (© 2021 Society for Experimental Biology and John Wiley & Sons Ltd.)

Monogalactosyldiacylglycerol (MGDG), the most abundant lipid in thylakoid membranes, is involved in photosynthesis and chloroplast development. MGDG lipase has an important role in lipid remodeling in Chlamydomonas reinhardtii. However, the process related to turnover of the lysogalactolipid that results from MGDG degradation, monogalactosylmonoacylglycerol (MGMG), remains to be clarified. Here we identified a homolog of Arabidopsis thaliana lysophosphatidylcholine acyltransferase (LPCAT) and characterized two independent knockdown (KD) alleles in C. reinhardtii. The enzyme designated as C. reinhardtiiLysolipid Acyltransferase 1 (CrLAT1) has a conserved membrane-bound O-acyl transferase domain. LPCAT from Arabidopsis has a key role in deacylation of phosphatidylcholine (PC). Chlamydomonas reinhardtii, however, lacks PC, and thus we hypothesized that CrLAT1 has some other important function in major lipid flow in this organism. In the CrLAT1 KD mutants, the amount of MGMG was increased, but triacylglycerols (TAGs) were decreased. The proportion of more saturated 18:1 (9) MGDG was lower in the KD mutants than in their parental strain, CC-4533. In contrast, the proportion of MGMG has decreased in the CrLAT1 overexpression (OE) mutants, and the proportion of 18:1 (9) MGDG was higher in the OE mutants than in the empty vector control cells. Thus, CrLAT1 is involved in the recycling of MGDG in the chloroplast and maintains lipid homeostasis in C. reinhardtii., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Acyl-lipids are vital components for all life functions of plants. They are widely studied using often in vitro conditions to determine inter alia the impact of genetic modifications and the description of biochemical and physiological functions of enzymes responsible for acyl-lipid metabolism. What is currently lacking is knowledge of if these results also hold in real environments-in in vivo conditions. Our study focused on the comparative analysis of both in vitro and in vivo growth conditions and their impact on the acyl-lipid metabolism of Camelina sativa leaves. The results indicate that in vitro conditions significantly decreased the lipid contents and influenced their composition. In in vitro conditions, galactolipid and trienoic acid (16:3 and 18:3) contents significantly declined, indicating the impairment of the prokaryotic pathway. Discrepancies also exist in the case of acyl-CoA:lysophospholipid acyltransferases (LPLATs). Their activity increased about 2-7 times in in vitro conditions compared to in vivo . In vitro conditions also substantially changed LPLATs' preferences towards acyl-CoA. Additionally, the acyl editing process was three times more efficient in in vitro leaves. The provided evidence suggests that the results of acyl-lipid research from in vitro conditions may not completely reflect and be directly applicable in real growth environments.

The lipid bilayer matrix of the thylakoid membrane of cyanobacteria and chloroplasts of plants and algae is mainly composed of uncharged galactolipids, but also contains anionic lipids sulfoquinovosyldiacylglycerol (SQDG) and phosphatidylglycerol (PG) as major constituents. The necessity of PG for photosynthesis is evident in all photosynthetic organisms examined to date, whereas the requirement of SQDG varies with species. In plants, although PG and SQDG are also found in non-photosynthetic plastids, their importance for the growth and functions of non-photosynthetic organs remains unclear. In addition, plants synthesize another anionic lipid glucuronosyldiacylglycerol (GlcADG) during phosphorus starvation, but its role in plant cells is not elucidated yet. To understand the functional relationships among PG, SQDG, and GlcADG, we characterized several Arabidopsis thaliana mutants defective in biosynthesis of these lipids. The mutants completely lacking both PG and SQDG biosynthesis in plastids showed developmental defects of roots, hypocotyls, and embryos in addition to leaves, which suggests that these lipids are pleiotropically required for the development of both photosynthetic and non-photosynthetic organs. Furthermore, our analysis revealed that SQDG, but not GlcADG, is essential for complementing the role of PG, particularly in photosynthesis under PG-deficient conditions such as phosphorus starvation.

Lipids have been observed attached to lumen-facing surfaces of mature xylem conduits of several plant species, but there has been little research on their functions or effects on water transport, and only one lipidomic study of the xylem apoplast. Therefore, we conducted lipidomic analyses of xylem sap from woody stems of seven plants representing six major angiosperm clades, including basal magnoliids, monocots and eudicots, to characterize and quantify phospholipids, galactolipids and sulfolipids in sap using mass spectrometry. Locations of lipids in vessels of Laurus nobilis were imaged using transmission electron microscopy and confocal microscopy. Xylem sap contained the galactolipids di- and monogalactosyldiacylglycerol, as well as all common plant phospholipids, but only traces of sulfolipids, with total lipid concentrations in extracted sap ranging from 0.18 to 0.63 nmol ml -1 across all seven species. Contamination of extracted sap from lipids in cut living cells was found to be negligible. Lipid composition of sap was compared with wood in two species and was largely similar, suggesting that sap lipids, including galactolipids, originate from cell content of living vessels. Seasonal changes in lipid composition of sap were observed for one species. Lipid layers coated all lumen-facing vessel surfaces of L. nobilis, and lipids were highly concentrated in inter-vessel pits. The findings suggest that apoplastic, amphiphilic xylem lipids are a universal feature of angiosperms. The findings require a reinterpretation of the cohesion-tension theory of water transport to account for the effects of apoplastic lipids on dynamic surface tension and hydraulic conductance in xylem., (© 2020 Society for Experimental Biology and John Wiley & Sons Ltd.)

Head group-acylated chloroplast lipids were discovered in the 1960s, but interest was renewed about 15 years ago with the discovery of Arabidopsides E and G, acylated monogalactosyldiacylglycerols with oxidized fatty acyl chains originally identified in Arabidopsis thaliana . Since then, plant biologists have applied the power of mass spectrometry to identify additional oxidized and non-oxidized chloroplast lipids and quantify their levels in response to biotic and abiotic stresses. The enzyme responsible for the head-group acylation of chloroplast lipids was identified as a cytosolic protein closely associated with the chloroplast outer membrane and christened acylated galactolipid-associated phospholipase 1 (AGAP1). Despite many advances, critical questions remain about the biological functions of AGAP1 and its head group-acylated products.

Under certain abiotic conditions (elevated irradiance, temperature and sediment salinity) observed mostly during the Mediterranean summer, the halophyte Sarcocornia fruticosa suffers a metabolic shift evidenced by a red coloration, evidencing the presence of two physiotypes (green and red). Previous works indicated that this metabolic shift has severe implications in the primary photochemistry of this species, impairing the light and carbon harvesting. Under stress plants have lower light use efficiencies and are more prone to photoinhibition, and thus this metabolic shift is essential for this species to deal with the high light intensities characteristic from this time of the year. Nevertheless, the fatty acid and lipid remodelling in green and red S. fruticosa physiotypes was not previously evaluated nor its relations with this metabolic shift. The evaluation of the lipid landscape suggests several lipid and fatty acid remodelling when comparing both red and green physiotype, as strategies to overcome stress. The galactolipids of the red physiotype suffer several changes aiming to keep chloroplast membrane structural and functional stability during water stress and can also be related to an improvement of the plants response to osmotic stress. At the phospholipid level, a readjustment of its fatty acid profiles was also observable. This remodelling allows the plants to adjust membrane fluidity the imposed osmotic stress, being this action transversal to choroplastidial, extraplastidial, and involves the action of the different phospholipids. Additionally, neutral lipids (NLs) also appear to play a role in osmotic stress adaptation, with an increase content in C18 fatty acids in the red physiotype. The resulting lipid landscape in both physiotypes presents very specific signatures that can be used as biomarkers to track this kind of metabolic shifts, in future studies with similar species., (Copyright © 2020 Elsevier Masson SAS. All rights reserved.)

Plant chloroplasts have complex membrane systems. Among these, thylakoids serve as the sites for photosynthesis and photosynthesis-related adaptation. In addition to the photosynthetic membrane complexes and associated molecules, lipids in the thylakoid membranes, are predominantly composed of MGDG (monogalactosyldiacylglycerol), DGDG (digalactosyldiacylglycerol), SQDG (sulfoquinovosyldiacylglycerol) and PG (phosphatidylglycerol), play essential roles in shaping the thylakoid architecture, electron transfer, and photoregulation. In this review, we discuss the effect of abiotic stress on chloroplast structure, the changes in membrane lipid composition, and the degree of unsaturation of fatty acids. Advanced understanding of the mechanisms regulating chloroplast membrane lipids and unsaturated fatty acids in response to abiotic stresses is indispensable for improving plant resistance and may inform the strategies of crop breeding.

Aluminium (Al) is a key element that plays a major role in inhibiting plant growth and productivity under acidic soils. While lipids may be involved in plant tolerance/sensitivity to Al, the role of monogalactosyldiacylglycerol (MGDG) in Al response remains unknown. In this study, Arabidopsis MGDG synthase (AtMGD) mutants (mgd1, mgd2 and mgd3) and wild-type (Col-0) plants were treated with AlCl 3 ; the effect of aluminium on root growth, aluminium distribution, plasma membrane integrity, lipid peroxidation, hydrogen peroxide content and membrane lipid compositions were analysed. Under Al stress, mgd mutants exhibited a more severe root growth inhibition, plasma membrane integrity damage and lipid peroxidation compared to Col-0. Al accumulation in root tips showed no difference between Col-0 and mutants under Al stress. Lipid analysis demonstrated that under Al treatment the MGDG content in all plants and MGDG/DGDG (digalactosyldiacylglycerol) remarkably reduced, especially in mutants impairing the stability and permeability of the plasma membrane. These results indicate that the Arabidopsis mgd mutants are hypersensitive to Al stress due to the reduction in MGDG content, and this is of great significance in the discovery of effective measures for plants to inhibit aluminium toxicity., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In cereals, C-repeat binding factor genes have been defined as key components of the light quality-dependent regulation of frost tolerance by integrating phytochrome-mediated light and temperature signals. This study elucidates the differences in the lipid composition of barley leaves illuminated with white light or white light supplemented with far-red light at 5 or 15 °C. According to LC-MS analysis, far-red light supplementation increased the amount of monogalactosyldiacylglycerol species 36:6, 36:5, and 36:4 after 1 day at 5 °C, and 10 days at 15 °C resulted in a perturbed content of 38:6 species. Changes were observed in the levels of phosphatidylethanolamine, and phosphatidylserine under white light supplemented with far-red light illumination at 15 °C, whereas robust changes were observed in the amount of several phosphatidylserine species at 5 °C. At 15 °C, the amount of some phosphatidylglycerol species increased as a result of white light supplemented with far-red light illumination after 1 day. The ceramide (42:2)-3 content increased regardless of the temperature. The double-bond index of phosphatidylglycerol, phosphatidylserine, phosphatidylcholine ceramide together with total double-bond index changed when the plant was grown at 15 °C as a function of white light supplemented with far-red light. white light supplemented with far-red light increased the monogalactosyldiacylglycerol/diacylglycerol ratio as well. The gene expression changes are well correlated with the alterations in the lipidome.

Galactolipids, mainly monogalactosyl diglycerides and digalactosyl diglycerides are the main lipids found in the membranes of plants, algae and photosynthetic microorganisms like microalgae and cyanobacteria. As such, they are the main lipids present at the surface of earth. They may represent up to 80% of the fatty acid stocks, including a large proportion of polyunsaturated fatty acids mainly α-linolenic acid (ALA). Nevertheless, the interest in these lipids for nutrition and other applications remains overlooked, probably because they are dispersed in the biomass and are not as easy to extract as vegetable oils from oleaginous fruit and oil seeds. Another reason is that galactolipids only represent a small fraction of the acylglycerolipids present in modern human diet. In herbivores such as horses, fish and folivorous insects, galactolipids may however represent the main source of dietary fatty acids due to their dietary habits and digestion physiology. The development of galactolipase assays has led to the identification and characterization of the enzymes involved in the digestion of galactolipids in the gastrointestinal tract, as well as by microorganisms. Pancreatic lipase-related protein 2 (PLRP2) has been identified as an important factor of galactolipid digestion in humans, together with pancreatic carboxyl ester hydrolase (CEH). The levels of PLRP2 are particularly high in monogastric herbivores thus highlighting the peculiar role of PLRP2 in the digestion of plant lipids. Similarly, pancreatic lipase homologs are found to be expressed in the midgut of folivorous insects, in which a high galactolipase activity can be measured. In fish, however, CEH is the main galactolipase involved. This review discusses the origins and fatty acid composition of galactolipids and the physiological contribution of galactolipid digestion in various species. This overlooked aspect of lipid digestion ensures not only the intake of ALA from its main natural source, but also the main lipid source of energy for growth of some herbivorous species.

Chloroplast membranes have a high content of the uncharged galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG). These galactolipids are essential for the biogenesis of plastids and functioning of the photosynthetic machinery. A monotopic glycosyltransferase, monogalactosyldiacylglycerol synthase synthesizes the bulk of MGDG. It is embedded in the outer leaflet of the inner envelope membrane of chloroplasts. The protein transfers a galactose residue from UDP-galactose to diacylglycerol (DAG); it needs anionic lipids such as phosphatidylglycerol (PG) to be active. The intricacy of the organization and the process of active complex assembly and synthesis have been investigated at the Coarse-Grained and All-Atom of computer simulation levels to cover large spatial and temporal scales. The following self-assembly process and catalytic events can be drawn; (1) in the membrane, in the absence of protein, there is a spontaneous formation of PG clusters to which DAG molecules associate, (2) a reorganization of the clusters occurs in the vicinity of the protein once inserted in the membrane, (3) an accompanying motion of the catalytic domain of the protein brings DAG in the proper position for the formation of the active complex MGD1/UDP-Gal/DAG/PG for which an atomistic model of interaction is proposed.

Microalgae, due to their huge taxonomic and metabolic diversity, have been shown to be a valuable and eco-friendly source of bioactive natural products. The increasing number of genomic and transcriptomic data will give a great boost for the study of metabolic pathways involved in the synthesis of bioactive compounds. In this study, we analyzed the presence of the enzymes involved in the synthesis of monogalactosyldiacylglycerols (MGDGs) and sulfoquinovosyldiacylglycerols (SQDG). Both compounds have important biological properties. MGDGs present both anti-inflammatory and anti-cancer activities while SQDGs present immunostimulatory activities and inhibit the enzyme glutaminyl cyclase, which is involved in Alzheimer's disease. The Ocean Global Atlas (OGA) database and the Marine Microbial Eukaryotic Transcriptome Sequencing Project (MMETSP) were used to search MGDG synthase (MGD), UDP-sulfoquinovose synthase (SQD1), and sulfoquinovosyltransferase (SQD2) sequences along microalgal taxa. In silico 3D prediction analyses for the three enzymes were performed by Phyre2 server, while binding site predictions were performed by the COACH server. The analyzed enzymes are distributed across different taxa, which confirms the importance for microalgae of these two pathways for thylakoid physiology. MGD genes have been found across almost all analyzed taxa and can be separated in two different groups, similarly to terrestrial plant MGD. SQD1 and SQD2 genes are widely distributed along the analyzed taxa in a similar way to MGD genes with some exceptions. For Pinguiophyceae, Raphidophyceae, and Synurophyceae, only sequences coding for MGDG were found. On the contrary, sequences assigned to Ciliophora and Eustigmatophyceae were exclusively corresponding to SQD1 and SQD2. This study reports, for the first time, the presence/absence of these enzymes in available microalgal transcriptomes, which gives new insights on microalgal physiology and possible biotechnological applications for the production of bioactive lipids.

The increasing demand for triacylglycerol (TAG) enriching polyunsaturated fatty acids (PUFAs) has led to a surge of interest in microalgal TAG metabolism. Polar membrane lipids serve as the desaturation carrier for PUFA, and the functional group of PUFA can be incorporated into TAG. Monogalactoglycerolipid has been found to provide the de novo synthesized oleate acyl group or the nascent polyunsaturated diacylglycerol backbone for TAG biosynthesis in the model green alga, Chlamydomonas reinhardtii. However, whether other membrane lipids take part in the formation of PUFA-attached TAG has not been clearly discovered. A time course study of glycerolipidomics in the starchless mutant of C. reinhardtii, BAFJ5, which hyper-accumulates TAG, revealed that digalactosyldiacylglycerol (DGDG) and diacylglycerol-N,N,N-trimethylhomoserine (DGTS) turned into the main components of membrane lipids, accounting for 62% of the total polar lipids, under nitrogen deprivation combined with high light conditions. In addition, the membrane lipid molecules DGDG 18:3n3/16:0 and DGTS 16:0/18:3n6 were presumed to be involved in the consecutive integration of the de novo synthesized linolenates into TAG. Based on the stoichiometry calculation, DGDG and DGTS were demonstrated to provide a major contribution to the accumulation of linolenate-attached TAG. Our study gives insights into the potential PUFA-attached TAG formation pathway mediated by the turnover of de novo synthesized DGDG and DGTS in the starchless mutant of Chlamydomonas., (© The Author(s) 2020. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Integral membrane proteins are exposed to a complex and dynamic lipid environment modulated by nonbilayer lipids that can influence protein functions by lipid-protein interactions. The nonbilayer lipid monogalactosyldiacylglycerol (MGDG) is the most abundant lipid in plant photosynthetic thylakoid membranes, but its impact on the functionality of energy-converting membrane protein complexes is unknown. Here, we optimized a detergent-based reconstitution protocol to develop a proteoliposome technique that incorporates the major light-harvesting complex II (LHCII) into compositionally well-defined large unilamellar lipid bilayer vesicles to study the impact of MGDG on light harvesting by LHCII. Using steady-state fluorescence spectroscopy, CD spectroscopy, and time-correlated single-photon counting, we found that both chlorophyll fluorescence quantum yields and fluorescence lifetimes clearly indicate that the presence of MGDG in lipid bilayers switches LHCII from a light-harvesting to a more energy-quenching mode that dissipates harvested light into heat. It is hypothesized that in the in vitro system developed here, MGDG controls light harvesting of LHCII by modulating the hydrostatic lateral membrane pressure profile in the lipid bilayer sensed by LHCII-bound peripheral pigments.

Cyanobacterial monoglucosyldiacylglycerol (MGlcDG) not only serves as a precursor for monogalactosyldiacylglycerol (MGDG) synthesis, but also participates in stress acclimation. Two genes (mgdA and mgdE) related to MGDG synthesis of Synechococcus sp. PCC 7942 were identified. The mgdE-suppressed mutant (AE) accumulated MGlcDG (4.2%) and showed better growth and photosynthetic activities compared with WT and other mutants (mgdA/mgdE-overexpressed and mgdA-suppressed strains), which suggested that MGlcDG was involved in phosphate stress adaptation for Synechococcus sp. PCC 7942. A notable increase in contents of 18:1 fatty acid (FA) of MGDG (127%), DGDG (68%), and SQDG (105%) in AE were found under phosphate starvation. However, the expression of △9 desaturase (desC) was not higher in AE than that in WT during phosphate-starved period. These results suggested that MGlcDG might be involved in the process of FA desaturation, which contributed to membrane fluidity and cell basic metabolism for stress acclimation in cyanobacteria. In complementary experiments of E. coli, although the expression of mgdA and desC in the mgdA and desC coexpressed strain (OEAC) reduced by 22% and 35% compared with that of the strains only overexpressing mgdA (OEA) or desC (OEC), the content of unsaturated FA in OEAC was the highest. This further implied that the accumulation of MGlcDG could prompt FA desaturation in E. coli. Therefore, we propose that an overproduction of MGlcDG is responsible for FA desaturation and participates in phosphate stress adaptation in cyanobacteria., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The removal of intact chloroplasts from their cell wall confinement offers a novel way to obtain lipophilic nutrients from green biomass, especially carotenoids and galactolipids. These latter are the main membrane lipids in plants and they represent a major source of the essential α-linolenic acid (18:3; ALA). Nevertheless, knowledge on their digestion is still limited. We have developed a physical method of recovering a chloroplast-rich fraction (CRF) from green biomass and tested its digestibility in vitro under simulated gastrointestinal conditions. Using a two-step static model, CRF from both spinach leaves and postharvest, pea vine field residue (haulm) were first exposed to enzymes from rabbit gastric extracts and then either to pancreatic enzymes from human pancreatic juice (HPJ) or to porcine pancreatic extracts (PPE). The lipolysis of monogalactosyldiacylglycerol (MGDG) and digalactosyl diacylglycerol (DGDG) was monitored by thin layer chromatography and gas chromatography of fatty acid methyl esters. For both CRF preparations, MGDG and DGDG were converted to monogalactosylmonoacylglycerol (MGMG) and digalactosylmonoacylglycerol (DGMG), respectively, during the intestinal phase and ALA was the main fatty acid released. Galactolipids were more effectively hydrolysed by HPJ than by PPE, and PPE showed a higher activity on MGDG than on DGDG. These findings may be explained by the higher levels of galactolipase activity in HPJ compared to PPE, which mainly results from pancreatic lipase-related protein 2. Thus, we showed that CRF galactolipids are well digested by pancreatic enzymes and represent an interesting vehicle for ALA supplementation in human diet.

Organisms use various adaptive strategies against phosphate stress, including lipid remodeling. Here, the response of major membrane lipids to phosphate stress was analyzed in Synechococcus sp. PCC 7942. Unlike plants and eukaryotic microalgae, no significant increases in neutral lipids were found, whereas glycolipids content increased to as high as 6.13% (of dry cell weight, DCW) and phospholipids decreased to 0.34% (of DCW) after 16 days of cultivation without phosphate. Glycolipids accumulation were mainly attributed to the significant increase of digalactosyldiacylglycerol (DGDG) by 50% and sulfoquinovosyldiaclglycerol (SQDG) by 90%, both of which acted as complementary lipids for phosphatidylglycerol (PG) in the cyanobacterial membrane. Also, a notable increase in content (by 48%) of C18 fatty acids (especially C18:1) was observed in all glycolipids at the expense of C12 and C14 (72%). These changes may contribute to membrane fluidity and photosynthetic activity for basic cell metabolism and phosphate stress adaptation. Lipidomic analyses showed the reduction of PG 18:1/16: 0 (by 52%) with the increase of DGDG 18:1/16:0 (133%) and SQDG 18:1/16:0 (245%), strongly suggesting a direct conversion of PG to DGDG and SQDG. Moreover, the decreasing amount of monogalactosyldiacylglycerol (MGDG) 16:1/16:0 (22%) was consistent with the increase of free fatty acids (125%) on day 2 of phosphate absence, which suggested that MGDG is more likely to provide a pool of fatty acids for de novo synthesis of glycolipids. This study provides valuable insight into cyanobacteria adaptation strategies to phosphate stress by membrane lipid remodeling and unveils the underlying acyl chain fluxes into glycolipids., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Lipids are only minor wheat flour constituents but play major roles in bread making (BM). Here, the importance of a well-balanced lipid population in BM was studied by applying a lipase from Fusarium oxysporum in the process. Monogalactosyldiacylglycerols and N-acyl phosphatidylethanolamines were the most accessible lipase substrates. Hydrolysis thereof into their corresponding lysolipids was largely if not entirely responsible for loaf volume increases upon lipase application. Degradation of endogenously present lipids and enzymatically released lysolipids caused loaf volume to decrease, confirming that an appropriate balance between different types of lipids is crucial in BM. For optimal dough gas cell stability, the level of lipids promoting lamellar mesophases and, thus, liquid condensed monolayers needs to be maximal while maintaining an appropriate balance between lipids promoting hexagonal I phases, non-polar lipids and lipids promoting hexagonal II or cubic phases., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Proteinaceous inclusions containing alpha-synuclein (α-Syn) have been implicated in neuronal toxicity in Parkinson's disease, but the pathways that modulate toxicity remain enigmatic. Here, we used a targeted proteomic assay to simultaneously measure 269 pathway activation markers and proteins deregulated by α-Syn expression across a panel of 33 Saccharomyces cerevisiae strains that genetically modulate α-Syn toxicity. Applying multidimensional linear regression analysis to these data predicted Pah1, a phosphatase that catalyzes conversion of phosphatidic acid to diacylglycerol at the endoplasmic reticulum membrane, as an effector of rescue. Follow-up studies demonstrated that inhibition of Pah1 activity ameliorates the toxic effects of α-Syn, indicate that the diacylglycerol branch of lipid metabolism could enhance α-Syn neuronal cytotoxicity, and suggest a link between α-Syn toxicity and the biology of lipid droplets., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Key Message: Here we show that accumulation of galactose-containing lipids in plastid membranes in shoots and the other membranes in roots maintains Arabidopsis growth under acidic stress and acidic phosphate deficiency. Soil acidification and phosphate deficiency are closely related to each other in natural environments. In addition to the toxicity of high proton concentrations, acid soil can lead to imbalances of ion availability and nutritional deficiencies, including inorganic phosphate (Pi). Among plants, activation of non-phosphorus-containing galactolipid, digalactosyldiacylglycerol (DGDG), synthesis concomitant with phospholipid degradation, namely membrane lipid remodeling, is crucial for coping with Pi starvation. However, regulation mechanisms of membrane lipid composition during acidic stress have not been clarified. Here, we investigated lipid metabolism in Arabidopsis thaliana grown under acidic stress with or without Pi. Under Pi-sufficient acidic conditions, DGDG was increased in shoot membranes, and some Pi starvation-responsive genes that are involved in lipid remodeling were upregulated without reducing Pi content in leaves. In contrast, under acidic Pi deficiency, membrane lipid remodeling in roots was partially repressed at a lower external pH. Nevertheless, phenotypic comparison between wild type and the double mutant of MGD2/3, which are responsible for DGDG accumulation during Pi starvation, indicated that the complete absence of lipid remodeling in roots resulted in a loss of tolerance to Pi deficiency rather specifically under acidic conditions. This result suggested important physiological roles of galactolipid-enriched membranes under acidic Pi deficiency.

The thylakoid membranes of the chloroplast harbor the photosynthetic machinery that converts light into chemical energy. Chloroplast membranes are unique in their lipid makeup, which is dominated by the galactolipids mono- and digalactosyldiacylglycerol (MGDG and DGDG). The most abundant galactolipid, MGDG, is assembled through both plastid and endoplasmic reticulum (ER) pathways in Arabidopsis, resulting in distinguishable molecular lipid species. Phosphatidic acid (PA) is the first glycerolipid formed by the plastid galactolipid biosynthetic pathway. It is converted to substrate diacylglycerol (DAG) for MGDG Synthase (MGD1) which adds to it a galactose from UDP-Gal. The enzymatic reactions yielding these galactolipids have been well established. However, auxiliary or regulatory factors are largely unknown. We identified a predicted rhomboid-like protease 10 (RBL10), located in plastids of Arabidopsis thaliana, that affects galactolipid biosynthesis likely through intramembrane proteolysis. Plants with T-DNA disruptions in RBL10 have greatly decreased 16:3 (acyl carbons:double bonds) and increased 18:3 acyl chain abundance in MGDG of leaves. Additionally, rbl10-1 mutants show reduced [ 14 C]-acetate incorporation into MGDG during pulse-chase labeling, indicating a reduced flux through the plastid galactolipid biosynthesis pathway. While plastid MGDG biosynthesis is blocked in rbl10-1 mutants, they are capable of synthesizing PA, as well as producing normal amounts of MGDG by compensating with ER-derived lipid precursors. These findings link this predicted protease to the utilization of PA for plastid galactolipid biosynthesis potentially revealing a regulatory mechanism in chloroplasts., (© 2019 The Authors The Plant Journal © 2019 John Wiley & Sons Ltd.)

Problem: Sperm are the major cells in semen. Human sperm possess a number of HIV-1 gp120 binding ligands including sulfogalactosylglycerolipid (SGG). However, the mechanisms of how sperm capture HIV-1 onto their surface are unclear. Furthermore, the ability of sperm to deliver HIV-1 to vaginal/cervical epithelial cells lining the lower female reproductive tract, as a first step in HIV-1 transmission, needs to be determined., Method of Study: Sperm from healthy donors were incubated with dual-tropic HIV-1 CS204 (clinical isolate), and virus capture was determined by p24 antigen ELISA. The involvement of SGG in HIV-1 capture was assessed by determining K d values of HIV-1 gp120-SGG binding as well as computational docking of SGG to the gp120 V3 loop. The ability of sperm-associated HIV-1 to infect peripheral blood mononuclear cells (PBMCs) and TZM-bl indicator cells was determined. Lastly, infection of vaginal (Vk2/E6E7), ectocervical (Ect1/E6E7), and endocervical (End1/E6E7) epithelial cells mediated by HIV-1-associated sperm was evaluated., Results: Sperm were able to capture HIV-1 in a dose-dependent manner, and the capture reached a maximum within 5 minutes. Captured HIV-1, however, could be removed from sperm by Percoll-gradient centrifugation. Affinity of gp120 for SGG was substantial, implicating sperm SGG in HIV-1 capture. Sperm-associated HIV-1 could productively infect PBMCs and TZM-bl cells, and was capable of being transmitted into vaginal/cervical epithelial cells., Conclusion: Sperm are able to capture HIV-1, which remains infectious and is able to be transmitted into vaginal/cervical epithelial cells, a result indicating the importance of sperm in HIV transmission., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Etioplasts developed in angiosperm cotyledon cells in darkness rapidly differentiate into chloroplasts with illumination. This process involves dynamic transformation of internal membrane structures from the prolamellar bodies (PLBs) and prothylakoids (PTs) in etioplasts to thylakoid membranes in chloroplasts. Although two galactolipids, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), are predominant lipid constituents of membranes in both etioplasts and chloroplasts, their roles in the structural and functional transformation of internal membranes during etioplast-to-chloroplast differentiation are unknown. We previously reported that a 36% loss of MGDG by an artificial microRNA targeting major MGDG synthase (amiR-MGD1) only slightly affected PLB structures but strongly impaired PT formation and protochlorophyllide biosynthesis. Meanwhile, strong DGDG deficiency in a DGDG synthase mutant (dgd1) disordered the PLB lattice structure in addition to impaired PT development and protochlorophyllide biosynthesis. In this study, thylakoid biogenesis after PLB disassembly with illumination was strongly perturbed by amiR-MGD1. The amiR-MGD1 expression impaired the accumulation of Chl and the major light-harvesting complex II protein (LHCB1), which may inhibit rapid transformation from disassembled PLBs to the thylakoid membrane. As did amiR-MGD1 expression, dgd1 mutation impaired the accumulation of Chl and LHCB1 during etioplast-to-chloroplast differentiation. Furthermore, unlike in amiR-MGD1 seedlings, in dgd1 seedlings, disassembly of PLBs after illumination was retarded. Because DGDG but not MGDG prefers to form the bilayer lipid phase in membranes, the MGDG-to-DGDG ratio may strongly affect the transformation of PLBs to the thylakoid membrane during etioplast-to-chloroplast differentiation., (� The Author(s) 2019. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Diatoms are eukaryotic microalgae that play a pivotal role in biological and geochemical marine cycles. These microorganisms are at the basis of the trophic chain and their lipids are essential components (e.g. eicosapentaenoic acid, EPA) of aquatic food webs. Galactolipids are the primary lipid components of plastid membranes and form the largest lipid family of diatoms. As source of polyunsaturated fatty acids (PUFAs), these compounds are also involved in the synthesis of lipoxygenase (LOX) products such as non-volatile oxylipins and polyunsaturated aldehydes. Here, we report the first identification of two genes, namely PmLAH1 and PaLAH1, coding for lipolytic enzymes in two diatoms of the genus Pseudo-nitzschia. Functional and modeling studies evidence a patatin-like domain endowed with galactolipase and phospholipase activity at the C-terminus of both proteins. Homologues of Pseudo-nitzschia LAH1 genes were retrieved in other diatom species so far sequenced in agreement with conservation of the functional role of these proteins within the lineage., (Copyright © 2018 Elsevier B.V. All rights reserved.)