Regulation Of Membrane Lipids Research Articles

An association analysis of lipidome and transcriptome highlights the involvement of MmGDPD1 in regulating low phosphorus tolerance in Malus mandshurica

Phosphorus (Pi) plays a crucial role in the growth and development of plants. Membrane lipid regulation is one of the main mechanisms underlying plant adaptation to Pi deficiency. Previously, the high tolerance to low-Pi stress was justified in an elite line, MSDZ 109, which was obtained from Malus mandshurica. To better understand the mechanism underlying high adaptation to low-Pi stress, currently, lipidomic and transcriptomic analysis, as well as CRISPR/Cas9 and MmGDPD1-overexpressing methodologies were comprehensively integrated into a strategy for elucidating the high tolerance to low-Pi stress. Totally, 770 differential metabolites were identified from the roots between the low-Pi and stress-free, belonging to 21 sub-classes of lipid compounds. Fatty acids (FA) constituted the predominant lipid component, accounting for approximately 50%–60% of the total lipids, and triglycerides (TAG) ranked the second, comprising around 12% of the total, consecutively followed by phosphatidylcholine (PC) and diacylglycerol (DAG) with approximately 10% and 8% of the total, respectively. The synchronous transcriptomic analysis revealed a significant up-regulation of genes related to glycerophospholipid and glycerolipid metabolism, specifically those (e.g., MmGDPD1, MmDGDG1, MmMGDG1, MmSQDG, etc.) involved in phospholipid and galactosyl synthesis in response to low-Pi stress. GUS fusing reporter assay showed that MmGDPD1 promoter induced GUS gene expression and demonstrated initiation activity. Based on expression analysis, a dual-luciferase reporter assay, as well as yeast one-hybrid (Y1H) identification, MmPHR1 was justified to bind with the MmGDPD1 promoter and positively regulate plant tolerance to low-Pi stress. To further elucidate the role of MmGDPD1, CRISPR/Cas9 and MmGDPD1-overexpressing vectors were successfully introduced into apple (‘Royal Gala’) calli. Interestingly, the MmGDPD1-KO line calli exhibited the remarkable decreases in the contents of phosphodiesterase (PDE), activity, as well as the contents of total Pi, and Pi in comparison with those of the wild type. Conversely, MmGDPD1-OE ones demonstrated the significant elevation in Pi accumulations, further justifying its potential role in Pi remobilization in apple. Therefore, MmGDPD1 substantially involves elevating low-Pi tolerance via promoting Pi release in M. mandshurica.

Identification and Transcriptome Analysis of Genes Related to Membrane Lipid Regulation in Sweet Sorghum under Salt Stress.

Sweet sorghum has strong stress resistance and is considered a promising energy crop. In the present study, the effects of salt on the membrane lipid metabolism of two sweet sorghum inbred lines (salt-tolerant M-81E and salt-sensitive Roma) were analyzed. After treatment with 150 mM NaCl, higher levels of fresh weight and chlorophyll fluorescence, as well as lower levels of malondialdehyde (MDA) were found in salt-tolerant M-81E. Concomitantly, 702 and 1339 differentially expression genes (DEGs) in M-81E and Roma were identified in response to salt stress. We determined that most DEGs were related to glycerophospholipid metabolism, glycerolipid metabolism, and other membrane lipid metabolisms. Under NaCl treatment, the expression of the membrane-associated phospholipase A1 was down-regulated at the transcriptional level, along with an increased content of phosphatidylcholine (PC) in both cultivars. The inhibition of triacylglycerol (TAG) mobilization in M-81E delayed salt-induced leaf senescence. Furthermore, enhanced levels of glycerol-3-phosphate acyltransferase (GPAT) expression contributed to improved salt resistance in M-81E. The results of this study demonstrate membrane the role of lipid regulation in mediating salt-defensive responses in sweet sorghum and expand our understanding of the relationship between changes in membrane lipid content and salt resistance.

Isolation and Functional Analysis of Arteriolar Endothelium of Mouse Brain Parenchyma.

Cerebral blood flow is conveyed by vascular resistance arteries and downstream parenchymal arterioles. Steady-state vascular resistance to blood flow increases with decreasing diameter from arteries to arterioles that ultimately feed into capillaries. Due to their smaller size and location in the parenchyma, arterioles have been relatively understudied and with less reproducibility in findings than surface pial arteries. Regardless, arteriolar endothelial cell structure and function-integral to the physiology and etiology of chronic degenerative diseases-requires extensive investigation. In particular, emerging evidence demonstrates that compromised endothelial function precedes and exacerbates cognitive impairment and dementia. In the parenchymal microcirculation, endothelial K+ channel function is the most robust stimulus to finely control the spread of vasodilation to promote increases in blood flow to areas of neuronal activity. This paper illustrates a refined method for freshly isolating intact and electrically coupled endothelial "tubes" (diameter, ~25 µm) from mouse brain parenchymal arterioles. Arteriolar endothelial tubes are secured during physiological conditions (37 °C, pH 7.4) to resolve experimental variables that encompass K+ channel function and their regulation, including intracellular Ca2+ dynamics, changes in membrane potential, and membrane lipid regulation. A distinct technical advantage versus arterial endothelium is the enhanced morphological resolution of cell and organelle (e.g., mitochondria) dimensions, which expands the usefulness of this technique. Healthy cerebral perfusion throughout life entails robust endothelial function in parenchymal arterioles, directly linking blood flow to the fueling of neuronal and glial activity throughout precise anatomical regions of the brain. Thus, it is expected that this method will significantly advance the general knowledge of vascular physiology and neuroscience concerning the healthy and diseased brain.

Fluidity is the way to life: lipid phase separation in bacterial membranes.

A hallmark of biological membranes is the dynamic localization of lipids and proteins. Lipids respond to temperature reduction below a critical point with phase separation, and poikilothermic animals and also bacteria adapt their lipid content to prevent gel phase formation in membranes. In a new study, Gohrbandt etal (2022) show that reduced membrane fluidity in bacterial cells causes reversible phase separation without membrane rupture invivo, highlighting the physical robustness of biological membranes.

Proper regulation of inositolphosphorylceramide levels is required for acquirement of low pH resistance in budding yeast

All organisms have stress response systems to protect themselves from various environmental stresses, and regulation of membrane lipids is thought to play an important role in acquirement of stress tolerance. Complex sphingolipids in the yeast Saccharomyces cerevisiae are classified into three types based on differences in the structure of the polar head group, and the compositions and quantities of complex sphingolipids in biomembranes are tightly regulated. In this study, we found that the accumulation of inositol phosphorylceramides (IPCs) due to a defect of mannosylinositol phosphorylceramide biosynthesis (sur1∆ csh1∆), i.e., disruption of the balance of the composition of complex sphingolipids, causes hypersensitivity to low pH conditions (pH 4.0–2.5). Furthermore, screening of suppressor mutations that confer low pH resistance to sur1∆ csh1∆ cells revealed that a change in ergosterol homeostasis at plasma membranes can rescue the hypersensitivity, suggesting the functional relationship between complex sphingolipids and ergosterol under low pH conditions. Under low pH conditions, wild-type yeast cells exhibited decreases in IPC levels, and forced enhancement of the biosynthesis of IPCs causes low pH hypersensitivity. Thus, it was suggested that the accumulation of IPCs is detrimental to yeast under low pH conditions, and downregulation of IPC levels is one of the adaptation mechanisms for low pH conditions.

Gene identification and functional characterization of a Δ12 fatty acid desaturase in Tetrahymena thermophila and its influence in homeoviscous adaptation to low temperature

Homeoviscous adaptation in poikilotherms is based in the regulation of the level of desaturation of fatty acids, variation in phospholipids head groups and sterol content in the membrane lipids, in order to maintain the membrane fluidity in response to changes in environmental temperature. Increased proportion of unsaturated fatty acids is thought to be the main response to low-temperature acclimation, which is mostly achieved by fatty acid desaturases. Genome analysis of the ciliate Tetrahymena thermophila and a gene knockout approach has allowed us to identify one Δ12 FAD and to study its activity in the original host and in a yeast heterologous expression system. The “PUFA index” -relative content of polyunsaturated fatty acids compared to the sum of saturated and monounsaturated fatty acid content- was ~57% lower at 15 °C and 35 °C in the Δ12 FAD gene knockout strain (KOΔ12) compared to WT strain. We characterized the role of T. thermophila Δ12 FAD on homeoviscous adaptation and analyzed its involvement in cellular growth, cold stress response, and membrane fluidity, as well as its expression pattern during temperature shifts. Although these alterations allowed normal growth in the KOΔ12 strain at 30 °C or higher temperatures, growth was impaired at temperatures of 20 °C or lower, where homeoviscous adaptation is impaired. These results stress the importance of Δ12 FAD in the regulation of cold adaptation processes, as well as the suitability of T. thermophila as a valuable model to investigate the regulation of membrane lipids and evolutionary conservation and divergence of the underlying mechanisms.

Regulation of KcsA by Bilayer-Modifying Molecules

The coupling between membrane proteins and their surrounding lipid bilayer make understanding the membrane lipid regulation of membrane proteins key to understanding their function. The membrane lipid regulation of membrane protein function ranges from specific lipid binding to more general lipid bilayer regulation. The latter mechanism arises from energetic coupling between the lipid bilayer and membrane protein where protein conformational change will alter the organization of the adjacent bilayer. To gain insight into, and tease apart, the specific and general mechanism(s) of membrane protein regulation by the membrane lipids, we examine how the prototypical bacterial potassium channel, KcsA, is regulated by bilayer modifying compounds and drugs using a fluorescence-based technique. The concentration ranges at which the compounds alter the lipid bilayer is determined using gramicidin channels as probes. Comparing the concentration ranges at which molecules alter KcsA function and lipid bilayer properties, enable us to determine whether the underlying mechanism primarily involves specific interactions with KcsA or is bilayer-mediated. Most compounds tested slowed activation and accelerated inactivation at concentrations below those where they alter bilayer properties, suggesting that KcsA likely has a non-specific site that allows hydrophobic molecules to bind and alter channel properties. This site may be the fenestrations proposed to be located on the cytosolic side of aqueous pore of potassium channels (and evident in the x-ray structure). The same compounds alter KcsA function at the concentrations where they alter lipid bilayer properties. These, bilayer-modifying, molecules slow recovery from inactivation at concentrations at which they alter bilayer properties. These results suggest that conformational changes responsible for recovery from inactivation are regulated by a bilayer-mediated mechanism. Our results point to a complicated relationship between direct and bilayer-mediated mechanisms, which both are rather non-specific and may affect a diverse range of ion channels.

Intra-Colonial Functional Differentiation-Related Modulation of the Cellular Membrane in a Pocilloporid Coral Seriatopora caliendrum.

Scleractinian corals have displayed phenotypic gradients of polyps within a single genotypic colony, and this has profound implications for their biology. The intrinsic polymorphism of membrane lipids and the molecular interactions involved allow cells to dynamically organize their membranes to have physicochemical properties appropriate for their physiological requirements. To gain insight into the accommodation of the cellular membrane during ontogenetic shifts, intra-colony differences in the glycerophosphocholine profiling of a pocilloporid coral, Seriatopora caliendrum, were characterized using a previously validated method. Specifically, several major polyunsaturated phosphatidylcholines showed higher levels in the distal tissue of coral branches. In contrast, the corresponding molecules with 1-2-degree less unsaturation and plasmanylcholines were expressed more highly in the proximal tissue. The lipid profiles of these two colonial positions also contrasted sharply with regard to the saturated, monounsaturated, and lyso-glycerophosphocholine ratios. Based on the biochemical and biophysical properties of these lipids, the associated modulation of cellular membrane properties could be related to the physiological requirements, including coral growth and aging, of the functionally differentiated polyps. In this study, the metabolic regulation of membrane lipids involved in the functional differentiation of polyps within a S. caliendrum colony was identified.

Lipid Modulation of Calcium Flux through CaV2.3 Regulates Acrosome Exocytosis and Fertilization

Membrane lipid regulation of cell function is poorly understood. In early development, sterol efflux and the ganglioside GM1 regulate sperm acrosome exocytosis (AE) and fertilization competence through unknown mechanisms. Here, we show that sterol efflux and focal enrichment of GM1 trigger Ca(2+) influx necessary for AE through CaV2.3, whose activity has been highly controversial in sperm. Sperm lacking CaV2.3's pore-forming α1E subunit showed altered Ca(2+) responses, reduced AE, and a strong subfertility phenotype. Surprisingly, AE depended on spatiotemporal information encoded by flux through CaV2.3, not merely the presence/amplitude of Ca(2+) waves. Using studies in both sperm and voltage clamp of Xenopus oocytes, we define a molecular mechanism for GM1/CaV2.3 regulatory interaction, requiring GM1's lipid and sugar components and CaV2.3's α1E and α2δ subunits. Our results provide a mechanistic understanding of membrane lipid regulation of Ca(2+) flux and therefore Ca(2+)-dependent cellular and developmental processes such as exocytosis and fertilization.

Adaptive regulation of membrane lipids and fluidity during thermal acclimation in Tetrahymena

The free-living eukaryotic protozoan Tetrahymena is a potentially useful model for the thermoadaptive membrane regulation because of easy growth in the axenic culture, systematic isolation of subcellular organelles, and quick response to temperature stress. Exposure of Tetrahymena cells to the cold temperature induces marked alterations in the lipid composition and the physical properties (fluidity) of various membranes. The increase in fatty acid unsaturation of membrane phospholipids is required to preserve the proper fluidity. In this homeoviscous adaptive response, acyl-CoA desaturase plays a pivotal role and its activity is regulated by induction of the enzyme via transcriptional activation.

Regulation of the plasma membrane during exposure to low temperatures in suspension-cultured cells from a cryophyte (Chorispora bungeana)

As the outermost boundary of the cell, the plasma membrane plays an important role in determining the stress resistance of organisms. To test this concept in a cryophyte, we analyzed alterations of several components in plasma membranes isolated from suspension-cultured cells of Chorispora bungeana Fisch. & C.A. Mey in response to treatment at 0 and -4 degrees C for 192 h. When compared with the controls growing at 25 degrees C, both the membrane permeability and fluidity showed recovery after the initial impairment. Linolenic acid and membrane lipid unsaturation increased by about 0.8-fold following cold treatments, although the kinetics of the increase varied with the temperatures examined. During the treatments, the plasma membrane H(+)-ATPase (EC 3.6.1.3) activity increased by 78.06% at 0 degrees C and 100.47% at -4 degrees C. However, the plasma membrane NADH oxidase (EC 1.6.99.3) activity only decreased when exposed to a lower temperature (-4 degrees C), and remained at 63.93% after being treated for 192 h. After the treatments, the physical properties of the plasma membranes of suspension-cultured cells, especially the -4 degrees C treated cells, were similar to those in the wild plants. These findings indicate that the specific mechanism of cold resistance of C. bungeana is tightly linked with the rapid and flexible regulation of membrane lipids and membrane-associated enzymes, which ensure the structural and functional integrity of the plasma membrane that is essential for withstanding low temperature.

A modeling framework describing the enzyme regulation of membrane lipids underlying gradient perception in Dictyostelium cells

Gradient perception describes the process by which information about the chemoattractant concentration field surrounding a cell is transformed into an internal signal which is responsible for directed cell motion. Recently, many important biochemical details in immobilized and mobile Dictyostelium cells have been uncovered regarding the roles of enzymes regulating phosphoinositide lipids on the cell membrane which are responsible for gradient perception. We report on a modeling framework that describes the relationship between the membrane concentration of the primary 3’phosphoinositide lipids and the enzymes which regulate them. The model takes the form of partial differential equations describing the membrane concentration of these lipids. Working within this framework, we describe mechanisms which can be responsible for spatial amplification of these lipids and which do not employ lipid–enzyme feedback. An analysis of a basic module underlying this process is also performed.

Membrane lipid regulation of the tumor suppressor phosphatase PTEN

The neutral sphingolipid ceramide has been implicated in the apoptotic death of cells by a number of different mechanisms and we now present evidence that one of its actions is to activate the tumor supressor protein PTEN (phosphatase and tensin homolog deleted from chromosome 10). PTEN is mutated in multiple advanced cancers, is absent from many tumor cells such as gliomas, and functions largely as a polyphosphoinositide phosphatase. PTEN with a red‐fluorescent protein (RFP) tag was overexpressed in both a human cell line derived from oligodendroglioma (HOG) and a rat pheochromocytoma cells line (PC12) by means of an inducible promoter system (tet‐OFF). Induction of the gene by removal of doxocycline showed a generalized (cytosolic) distribution of PTEN which was changed to a more membrane‐associated pattern following treatment with C2‐ceramide. Activation of the cell death pathway with either staurosporine, wortmannin or C2‐ceramide was facilitated by PTEN expression while lowering PTEN expression with antisense constructs partially blocked cell death. The staurosporine‐induced increase in ceramide was localized to the Raft (detergent‐resistant‐membrane fraction) where active PTEN is also found. We propose that formation of ceramide during apoptosis results in the translocation of PTEN to the plasma membrane, where it can hydrolyze the polyphosphoinositides which normally activate the key antiapoptotic protein, phospho‐Akt (protein kinase B). Since ceramide may also activate a Phospho‐Akt phosphatase, this may further explain the high pro‐apoptoic potential of ceramide.

Membrane lipid regulation in Acholeplasma laidlawii grown with saturated fatty acids. Biosynthesis of a triacylglucolipid forming reversed micelles

The membrane lipid composition in several strains of Acholeplasma laidlawii is regulated upon a change in the growth conditions. Monoglucosyldiacylglycerol (MGlcDAG) and diglucosyldiacylglycerol (DGlcDAG) are the most abundant lipids in the A. laidlawii membrane. A third glucolipid, 3-O-acyl-monoglucosyldiacylglycerol (MAMGlcDAG) is synthesized by strain A-EF22 when the membrane lipids contain large amounts of saturated acyl chains. The lipid regulation can be understood from a simple theoretical model, in which the cells strive to maintain a balance between the lipids constituting a bilayer and those forming reversed non-lamellar liquid crystalline phases. Thus, the physical chemistry of membrane lipids, in particular their ability to form different aggregate structures, constitutes the basis for the lipid regulation, and therefore an understanding of the phase equilibria of membrane lipids is crucial. MGlcDAG and MAMGlcDAG isolated from A. laidlawii strain A-EF22 membranes were studied mainly by 2H NMR, 1H NMR, and 1H NMR diffusion measurements. MAMGlcDAG, containing 96 mol % saturated acyl chains formed a gel/crystalline phase up to about 80 degrees C, where a transition occurred to a reversed micellar (L2) phase. This is an unexpected finding for a membrane lipid. However, this lipid homogeneously mixes with the other membrane lipids at physiological temperatures. Previous and new data on MGlcDAG show that the lamellar phase is stabilized when the length and the degree of unsaturation of the acyl chains are decreased. The physicochemical properties of MAMGlcDAG and MGlcDAG were compared and found to be of great significance for the physiological regulation of the lipids in the membrane. MAMGlcDAG is synthesized under conditions when the phase equilibria of MGlcDAG are shifted from a non-lamellar toward a lamellar phase. Apart from MAMGlcDAG, MGlcDAG is the major lipid in A. laidlawii strain A-EF22 which is able to form reversed aggregate structures. MAMGlcDAG probably assists MGlcDAG in maintaining an optimal molecular packing, or negative curvature, of the lipids in the membrane.