Unique Sterol Research Articles

A receptor kinase senses sterol by coupling with elicitins in auxotrophic Phytophthora

Sterols are vital nutrients and signals for eukaryotic organisms. Mammalian cells are known to sense and respond to sterol status changes to maintain them within strict limits, a process associated with various human diseases. However, this process is not understood in oomycete pathogens, most of which are sterol auxotrophic and must obtain sterols from host plants. Here, we report that Phytophthora sojae SSRK1 (sterol-sensing receptor kinase 1) detects host sterols by coupling with elicitins, thereby controlling signaling and sterol absorption. Sterols are recruited by extracellular soluble elicitins, and these complexes then bind to SSRK1 to form trimolecular complexes. These complexes subsequently trigger downstream calcium influx, activation of mitogen-activated protein kinase, and transcriptome reprogramming through the receptor's kinase activity. Our data demonstrate a unique sterol sensing pathway where elicitins and a transmembrane receptor kinase SSRK1 act as coreceptors for extracellular sterols. These findings also portray a sterol-based war between oomycetes and plants, providing a unique perspective on how a pathogen detects host signals during infection.

Unique sterol metabolite shifts in inflammatory bowel disease and primary sclerosing cholangitis

Inflammatory bowel disease (IBD) triggers chronic intestinal inflammation and is linked to primary sclerosing cholangitis (PSC). Cholesterol homeostasis, tightly regulated under normal conditions, becomes disrupted in both inflammation and chronic liver disease. We analyzed fecal and serum levels of cholesterol synthesis precursors, oxysterols, and phytosterols in 87 patients with IBD (81 for serum analysis) including patients with Crohn's disease (CD) and ulcerative colitis (UC), 11 patients with PSC, 21 patients with PSC-IBD (18 for serum analysis), and 16 healthy controls (17 for serum analysis). Cholesterol was analysed by flow injection analysis on a high-resolution hybrid quadrupole-Orbitrap mass spectrometer and further serum sterols and all fecal sterols were analysed by a gas chromatograph mass spectrometer. Serum levels of lanosterol, 7-dehydrocholesterol, 7-beta-hydroxycholesterol, 27-hydroxycholesterol, and the plant sterols campesterol, stigmasterol, and sitosterol were similar across control and patient groups. Notably, serum lathosterol was elevated in CD patients compared to those with UC, PSC, PSC-IBD, and healthy controls. All other serum and fecal sterols showed no differences between CD and UC. Cholesterol synthesis precursors in serum, serum cholesterol levels, and both serum and fecal plant sterol levels decreased with increasing IBD severity. Consequently, serum cholesterol, campesterol, sitosterol, and fecal 5-beta sitostanol and 5-alpha sitostanol were negatively correlated with C-reactive protein and fecal calprotectin. The conversion of cholesterol to coprostanol in feces was impaired in IBD, PSC, and PSC-IBD, independent of bowel inflammation severity or liver disease extent. Patients with PSC, and to a lesser extent PSC-IBD, had elevated serum plant sterol levels, positively correlating with liver disease markers. In conclusion, in patients with IBD, cholesterol biosynthetic precursors, serum cholesterol levels, and fecal plant sterols decrease with intestinal inflammation. An inverse association of serum plant sterols with intestinal inflammation was observed in patients with IBD and a direct association of serum phytosterols with liver injury in patients with PSC. The conversion of fecal cholesterol to coprostanol was impaired in all patient cohorts. IBD and PSC alter serum sterol levels differently, whereas changes in fecal sterols are not disease specific and are moderate.

Inhibition of SC4MOL and HSD17B7 shifts cellular sterol composition and promotes oligodendrocyte formation.

While the cholesterol biosynthesis pathway has been extensively studied, recent work has forged new links between inhibition of specific sterol pathway enzymes, accumulation of their unique sterol substrates, and biological areas as diverse as cancer, immunology, and neurodegenerative disease. We recently reported that dozens of small molecules enhance formation of oligodendrocytes, a glial cell type lost in multiple sclerosis, by inhibiting CYP51, Sterol 14-reductase, or EBP and inducing cellular accumulation of their 8,9-unsaturated sterol substrates. Several adjacent pathway enzymes also have 8,9-unsaturated sterol substrates but have not yet been evaluated as potential targets for oligodendrocyte formation or in many other biological contexts, in part due to a lack of available small-molecule probes. Here, we show that genetic suppression of SC4MOL or HSD17B7 increases the formation of oligodendrocytes. Additionally, we have identified and optimized multiple potent new series of SC4MOL and HSD17B7 inhibitors and shown that these small molecules enhance oligodendrocyte formation. SC4MOL inhibitor CW4142 induced accumulation of SC4MOL's sterol substrates in mouse brain and represents an in vivo probe of SC4MOL activity. Mechanistically, the cellular accumulation of these 8,9-unsaturated sterols represents a central driver of enhanced oligodendrocyte formation, as exogenous addition of purified SC4MOL and HSD17B7 substrates but not their 8,9-saturated analogs promotes OPC differentiation. Our work validates SC4MOL and HSD17B7 as novel targets for promoting oligodendrocyte formation, underlines a broad role for 8,9-unsaturated sterols as enhancers of oligodendrocyte formation, and establishes the first high-quality small molecules targeting SC4MOL and HSD17B7 as novel tools for probing diverse areas of biology.

C4-monomethylsterol β-glucoside and its synthase in Aurantiochytrium limacinum mh0186.

Thraustochytrids, unicellular marine protists, synthesize polyunsaturated fatty acids (PUFAs) and PUFA-containing phospholipids; however, little is known about their glycolipids and their associated metabolism. Here, we report two glycolipids (GL-A, B) and their synthases in Aurantiochytrium limacinum mh0186. Two glycolipids were purified from A. limacinum mh0186, and they were determined by gas chromatography, mass spectrometry and 2D nuclear magnetic resonance to be 3-O-β-D-glucopyranosyl-stigmasta-5,7,22-triene (GL-A) and 3-O-β-D-glucopyranosyl-4α-methyl-stigmasta-7,22-diene (GL-B), both of which are sterol β-glucosides (β-SGs); the structure of GL-B has not been reported thus far. Seven candidate genes responsible for the synthesis of these β-SGs were extracted from the draft genome database of A. limacinum using the yeast sterol β-glucosyltransferase (SGT; EC 2.4.1.173) sequence as a query. Expression analysis using Saccharomyces cerevisiae revealed that two gene products (AlSGT-1 and 2) catalyze the transfer of glucose from uridine diphosphate (UDP)-glucose to sterols, generating sterylglucosides (SGs). Compared to AlSGT-1, AlSGT-2 exhibited wide specificity for sterols and used C4-monomethylsterol to synthesize GL-B. The disruption of alsgt-2 but not alsgt-1 in strain mh0186 resulted in a decrease in the total SG and an almost complete loss of GL-B, indicating that AlSGT-2 is responsible for the synthesis of β-SGs in A. limacinum mh0186, especially GL-B, which possesses a unique sterol structure.

The unique sterol biosynthesis pathway of three model diatoms consists of a conserved core and diversified endpoints

Diatoms produce a wide diversity of sterols among different species, the biosynthesis and conservation of which is not yet fully understood. To investigate the conservation and divergence of sterol biosynthesis pathways among diatoms, we performed comparative metabolic profiling and transcriptomics for a centric diatom (Thalassiosira pseudonana), a pennate diatom (Phaeodactylum tricornutum) and a chaetocerid (Chaetoceros muelleri) in response to inhibitors of enzymes involved in sterol biosynthesis. These three model diatoms, which are representative of distinct clades, share a unique core phytosterol biosynthesis pathway that relies on a terbinafine-insensitive alternative squalene epoxidase and the cyclization of 2,3-epoxysqualene into cycloartenol by a conserved oxidosqualene cyclase. Lineage-specific divergence in the synthesis of sterol precursors was found in the species analyzed. Cholesterol synthesis in diatoms seems to occur via cycloartenol rather than lanosterol. The diversification of natural sterols produced by each species appears to occur downstream of all experimentally targeted enzymes, suggesting adaptive specialization in terminal synthesis pathways.

Sterol Characteristics in Silkworm Brain and Various Tissues Characterized by Precise Sterol Profiling Using LC-MS/MS.

Sterols, especially cholesterol (Chl), are fundamental for animal survival. Insects lacking the ability to synthesize Chl are sterol auxotrophic animals and utilize dietary Chl and phytosterols to survive. The sterols obtained from a diet are distributed to the tissues; however, sterol homeostasis in insect tissues remains to be elucidated. This study sought to understand the sterol characteristics of insect tissues through detailed sterol quantification and statistics. The combination of sterol quantification using liquid chromatography tandem mass spectrometry (LC-MS/MS) and principal component analysis (PCA) revealed tissue-specific sterol characteristics in the silkworm, Bombyx mori, a phytophagous insect. We found that insect tissues have tissue-intrinsic sterol profiles. The brain has a unique sterol composition as compared to other tissues—high concentration of Chl and less accumulation of phytosterols. Other tissues also have intrinsic sterol characteristics, which when defined by dietary sterols or Chl metabolites, indicate preference for a sterol and consistently manage their own sterol homeostasis. Though most tissues never change sterol profiles during development, the brain drastically changes its sterol profile at the wandering stage, indicating that it could alter sterol composition in preparation for metamorphosis. These results suggest the existence of tissue- and sterol-specific systems for sterol homeostasis in insects.

Synthesis and potent cytotoxic activity of a novel diosgenin derivative and its phytosomes against lung cancer cells.

Diosgenin (Di), a steroidal sapogenin derived from plants, has been shown to exert anticancer effects in preclinical studies. Using Di as a starting material, various Di derivatives were designed and synthesized, aiming to discover new steroid-based antitumor agents. In this work, we synthesized several Di derivatives and screened FZU-0021-194-P2 (P2), which showed more potent cytotoxic activities against human non-small-cell lung cancer A549 and PC9 cells. Considering that Di has a unique sterol structure similarly to cholesterol, P2 phytosomes (P2Ps) were prepared to further improve the water solubility of P2. The P2Ps exhibited a particle size of 53.6 ± 0.3 nm with oval shape and a zeta potential of −4.0 ± 0.7 mV. P2Ps could inhibit the proliferation of lung cancer cells more efficiently than Di phytosomes after 72 h of incubation time by inducing cell cycle arrest and apoptosis. The results indicated that P2Ps could be a promising anticancer formulation for non-small-cell lung cancer.

Profiling and Metabolism of Sterols in the Weaver Ant Genus Oecophylla

Sterols are essential to insects because they are vital for many biochemical processes, nevertheless insects cannot synthesize sterols but have to acquire them through their diet. Studies of sterols in ants are sparse and here the sterols of the weaver ant genus Oecophylla are identified for the first time. The sterol profile and the dietary sterols provided to a laboratory Oecophylla longinoda colony were analyzed. Most sterols originated from the diet, except one, which was probably formed via dealkylation in the ants and two sterols of fungal origin, which likely originate from hitherto unidentified endosymbionts responsible for supplying these two compounds. The sterol profile of a wild Oecophylla smaragdina colony was also investigated. Remarkable qualitative similarities were established between the two species despite the differences in diet, species, and origin. This may reflect a common sterol need/aversion in the weaver ants. Additionally, each individual caste of both species displayed unique sterol profiles.

Characterization of unusual sterols and long chain diols, triols, keto-ols and n-alkenols in El Junco Lake, Galápagos

A variety of lipid biomarkers were identified in sediments from El Junco Lake, Galápagos and their sources investigated for potential use in paleoclimate applications. A series of unusual sterols was also found, including 4α-methylgorgostanol, reported in only four species of dinoflagellates to date. We also tentatively assigned 22,23-methylene-4α-methyl-24-ethylcholest-5-en-3β-ol, the mass spectrum of which matched a sterol found in resting cysts of the dinoflagellate Peridinium umbonatum. In addition, we identified the novel sterol 4α,22,23,24-tetramethyl-5α-cholest-22E-en-3β-ol. Based on the unique sterol distribution, we hypothesize that a dinoflagellate from the genus Peridinium was the primary source of dinosterol and the novel sterols throughout the sediment record. The source specificity and abundance throughout the 3.7m of recovered sediment make dinosterol an excellent target for hydrogen isotope analysis for use as a paleohydrological proxy in future studies. The abundant C30 and C32 1,ω20-diols and keto-ols, C29 9,10-diol and C29 1,ω9,ω10-triol likely derive from the ferns Azolla microphylla and Cyathea weatherbyana, while sources of the C30 1,ω16-diol and keto-ol, C32 1,ω18-diol and keto-ol, and the C30–C32n-alken-1-ols are likely limited to aquatic microalgae. Due to their source specificity, these diol, triol, keto-ol, and n-alkenol biomarkers present further tools for studying past environmental and climatic change.

GC-MS Analysis of Some Bioactive Constituents from the Marine Sponge Spongosorites halichondriodes

Sponges have proved to be a rich source of sterols with unusual structural features 1, 2. They are also a rich source of structurally unusual and interesting fatty acids and steryl esters. To analyze and extract phylogenetic information from chemical characters, the 3-hydroxysterol composition of Halichondria lutea, H. magniconulosa, Topsentia ophiraphidites, and Petromica ciocalyptoides (family Halichondriidae) were studied. Three out of four Halichondriidae species were related by a unique sterol property. The diversity of sterols found in the studied species of Halichondrida confirms the usual pattern in Porifera 3. A close relationship between Topsentia and Petromica has already been drawn from morphology 4 and from similarity in secondary metabolite composition 5. In contrast, the two studied species of Halichondria came out separately in the analysis, not sharing a sterol synapomorphy, further evidencing the fact that this genus has been mainly defined by primitive characters 4, 6 and that it is polyphyletic 7. It is of special interest that, although some sponge species provide the greatest structural diversity of membrane sterols in comparison with any other animals, many of the studied sponges have sterol compositions resembling those of animals belonging to other taxa. In another study, D. australiensis contained 7 -, 5 -, 5,7 -, 5,7,9 (11)-sterols, and cholest-7-ene-3-ol was shown to be a main sterol. The free sterols from A. mauritiana proved to be stanols and 7 -series compounds. Haliclona sp. contained 5 -sterols with cholesterol as a main constituent. C. major and D. aceratus contained 5 -sterols, and clionasterol was a main sterol. T. labirintica was shown to contain 3-hydroxymethyl-A-nor-sterols. 8. The Caribbean sponge Neofibularia nolitangere showed comparable amounts of 24-methylene-4-methyl-5-cholest-8-ene-3-ol, which is very unusual among sponge sterols, and lesser quantities of two new polyoxygenated sterols, (24S)-24-ethyl-5-cholest-8-ene-5,6,7-triol and (24S)-24-ethyl-5-cholest-8(14)-ene-5,6,7triol 9. In our search for active compounds in Spongosorites halichondriodes, a marine sponge of class Demospongiae and family Halichondriidae, we have found that polyhydroxy steryl esters and cholesterol are predominant in this species. Spongosorites sp. is in almost all instances found in association with Vermetid molluscs and possesses potent bioactive compounds. Interestingly, it has also been observed that organisms that are conspicuously colored and live in habitats with limited or no ambient light often possess interesting secondary metabolites (Dercitus sp. and Spongosorites sp.) Seven monoindole derivatives were isolated from the MeOH extract of the marine sponge Spongosorites sp. by bioactivity-guided fractionation 10. Also, four bis (indole) alkaloids of the topsentin class, including two new brominated compounds, have been isolated from the sponge Spongosorites genitrix collected from Jaeju Island, Korea 11. These compounds also exhibited moderate cytotoxicity against cancer cell lines at concentrations between 1.1 and > 20 g/mL 12. Spongosorites halichondriodes has been studied and investigated for its antibacterial and antifungal activity. It has shown prominent antimicrobial activity in all the extracts along with cytotoxic activity in ethyl acetate extract. In the current work, the data collected from the GC-MS of the butanol extract was analyzed, and spectral matches were searched by comparing the parent ion fragmentation pattern m/z with the Mass Bank Database, Japan using Quick peak Search. The separation chromatogram of the crude extract of butanol and ethyl acetate showed the presence of similar peaks; butanol was selected for MS because of major peaks. Identification of Compounds by GC-MS. The fatty acid composition of phospholipids from marine sponges of the Colombian Caribbean Sea of the order Halichondrida (Halichondria magniconulosa, Halichondria lutea, Petromica ciocalyptoides, Axinyssa ambrosia, Didiscus oxeata, and Dragmaxia undata) has already been studied. Structure elucidation was accomplished by means of gas chromatography retention parameters and GC-MS of eight new fatty acids and their methyl esters and N-acylpyrrolidide derivatives (i.e., 5-methyl-6-octadecenoic, 16-methyl-11-heptadecenoic, 8-methyl-4-tetracosenoic,

Microbial Symbionts Shape the Sterol Profile of the Xylem-Feeding Woodwasp, Sirex noctilio

The symbiotic fungus Amylostereum areolatum is essential for growth and development of larvae of the invasive woodwasp, Sirex noctilio. In the nutrient poor xylem of pine trees, upon which Sirex feeds, it is unknown whether Amylostereum facilitates survival directly through consumption (mycetophagy) and/or indirectly through digestion of recalcitrant plant polymers (external rumen hypothesis). We tested these alternative hypotheses for Amylostereum involvement in Sirex foraging using the innate dependency of all insects on dietary sources of sterol and the unique sterols indicative of fungi and plants. We tested alternative hypotheses by using GC-MS to quantify concentrations of free and bound sterol pools from multiple life-stages of Sirex, food sources, and waste products in red pine (Pinus resinosa). Cholesterol was the primary sterol found in all life-stages of Sirex. However, cholesterol was not found in significant quantities in either plant or fungal resources. Ergosterol was the most prevalent sterol in Amylostereum but was not detectable in either wood or insect tissue (<0.001μg/g). Phytosterols were ubiquitous in both pine xylem and Sirex. Therefore, dealkylation of phytosterols (sitosterol and campesterol) is the most likely pathway to meet dietary demand for cholesterol in Sirex. Ergosterol concentrations from fungal-infested wood demonstrated low fungal biomass, which suggests mycetophagy is not the primary source of sterol or bulk nutrition for Sirex. Our findings suggest there is a potentially greater importance for fungal enzymes, including the external digestion of recalcitrant plant polymers (e.g., lignin and cellulose), shaping this insect-fungal symbiosis.

Chemical Composition of Iran′s Pistacia atlantica Cold‐Pressed Oil

The lipid fraction of Pistacia atlantica seeds was extracted for the first time by means of cold‐press technique and analyzed for its chemical composition. The fatty acids, sterols, triacylglycerols (TAG), tocopherols, polyphenols, and pigments were identified and their concentrations were determined by means of reversed‐phase high‐performance liquid chromatography (RP‐HPLC) and gas chromatography (GC). Because of its high content of unsaturated fatty acids, it might prove to be of value in diets and it may be used as edible cooking or salad oils or for margarine manufacture. Pistacia atlantica seed oil has the unique sterols and tocopherols content providing source of natural antioxidants. The main triacylglycerols were SLL + PLO, SOL + POO, OOLn + PLL, OOO, and SOO. This paper examined the phenolic fraction of Pistacia atlantica seed oil. Moreover, caffeic acid followed by cinnamic acid, pinoresinol, vanillin, p‐Coumaric acid, ferulic acid, and o‐Coumaric acid was also determined. This paper presents the first investigation of chlorophyll′s and carotene′s composition in Pistacia atlantica seed oil. Furthermore, pheophytin a was the major component, followed by luteoxanthin, neoxanthin, violaxanthin, lutein, lutein isomers, chlorophyll a, chlorophyll a′, and pheophytin a′ were also determined.

Steryl glycosides and acylated steryl glycosides in plant foods reflect unique sterol patterns

AbstractGlycosylated plant sterols (steryl glycosides, SG and acylated steryl glycosides, ASG) are minor glycolipids found in a variety of plants at different levels and in different compositions. They have until now been far less studied than other sterol conjugates, though they possess at least the same potential health benefits as free or esterified plant sterols. Additionally, due to the more hydrophilic conjugated part, they may possess further beneficial properties in forming water‐soluble structures. We studied the content and sterol profiles of glycosylated sterols in various plant foods to be able to better understand their levels in human diets, as well as to identify sources with high levels and/or unique sterol profile for possible extraction for ingredients. Glycosylated sterols contributed up to almost 60% of total sterols in e.g. potatoes, indicating that they indeed are an important group of sterols in plants. The ratio of ASG and SG varied from 0.4 to 3.6, indicating significant differences in the proportions of these two conjugates in various plants. Furthermore, extremely large differences were observed in the sterol profiles not only between but also within different plants, which indicates that the sterol profile of the glycosylated sterols does not always reflect the total sterol composition in the same plant tissue.Practical applications: Glycosylated sterols have hydrophilic properties owing to the carbohydrate moiety of the conjugate. As a consequence, they could potentially be introduced to water‐based functional foods (unlike the hydrophobic free sterols and sterol esters in current applications). Various plant materials or side streams of food processes were shown to be good sources of these glycosylated plant sterols, and could be used as sources for extraction of glycosylated sterols for food ingredients. Furthermore, the detailed compositional information reported here will provide a basis to select also the desired sterol composition for the product. Finally, the data presented in this paper is invaluable for estimating the contribution of different foods as sources of glycosylated sterols in the diet.See commentary by Jiménez‐Escrig [p. 615–616]

Sterols of the Green-Pigmented, Aberrant Plastid Dinoflagellate, Lepidodinium chlorophorum (Dinophyceae)

Lepidodinium chlorophorum is a green-pigmented dinoflagellate with an aberrant, tertiary plastid of chlorophyte ancestry rather than the typical red algal, secondary endosymbiont found in the vast majority of photosynthetic dinoflagellates. To date, only one published study exists on the galactolipids of L. chlorophorum, with nothing known about other lipid classes, including sterols. Our objectives were to examine the sterol composition of L. chlorophorum to determine if it produces any unique sterols with the potential to serve as biomarkers, and to compare it to members of the Chlorophyceae to determine if it has inherited any signature green algal sterols from its chlorophyte-derived endosymbiont. We have found that L. chlorophorum produces 6 sterols, all with a 4α-methyl substituent and none of which are known to occur in the Chlorophyceae. Rather, the sterols produced by L. chlorophorum place it within a group of dinoflagellates that have the common dinoflagellate sterols, dinosterol and dinostanol, as part of their sterol composition.

ChemInform Abstract: Structure of Paxisterol, a Sterol from Penicillium paxilli.

The complete structure of paxisterol 1, produced by a Penicillium species, was determined by means of spectral and chemical studies of the parent compound and several of its derivatives. Paxisterol is a unique sterol, having an acetal structure at C-18. Application of 2D-INADEQUATE technique in 13C NMR spectral analysis was useful in this structure determination, where all carbon-carbon bonds in the molecule were demonstrated. Absolute stereochemistry was confirmed by the CD spectrum of 3-keto derivative 10 of compound 1 to be the same as that of cholesterol. An interesting acetal migration reaction was observed during the chemical conversions.

Changes in fatty acids and sterols during batch growth of Pavlova viridis in photobioreactor

Changes in the composition of fatty acids and sterols of Pavlova viridis cultured in an air-lift photobioreactor were studied using gas chromatography-mass spectrometry (GC-MS). The results show that radical changes in fatty acid and sterol contents and compositions occurred during growth phase transitions: the total lipid increased along with the culture age, from 166.4 mg g−1 (late exponential phase) to 232.7 mg g−1 (linear phase), and increased further to be 235.1 mg g−1 in the stationary phase. Polyunsaturated fatty acids (PUFAs), especially eicosapentaenoic acid (EPA), decreased along with the culture time, PUFAs, and EPA contents maximized in the late exponential phase to become 46.2 mg g−1 and 22.1 mg g−1 respectively; there was no significant change in docosahexaenoic acid (DHA) content during the whole growth phase, although it reached the peak in the linear phase with 3.5 mg g−1. As for the sterols, two unique sterols with two hydroxyl groups, termed pavlovols, were observed. 4α,24-Dimethylcholestan-3β,4β-diol, one of the pavlovols, increased almost 2-fold from the late exponential phase (2.5 mg g−1) to the stationary phase (4.3 mg g−1). On the contrary, the contents of stigmasterol and sitosterol decreased with culture age, with the maximum content of 2.4 mg g−1 and 3.1 mg g−1, both obtained in the late exponential phase, respectively. The results indicate that growth phase control could be used as a methodology to optimize the total lipid, EPA, PUFA, and sterol contents with the potential for both aquaculture feeds and nutraceutical applications, especially for further research into unique pavlovols.

Sneaking under the toxin surveillance radar: parasitism and sterol content

Parasitic dinoflagellates of the genus Amoebophrya infect and kill bloom-forming dinoflagellates, including the toxic species Karlodinium micrum. Strains of K. micrum produce cytotoxic compounds (KmTX) that render cell membranes permeable to a range of small ions and molecules, resulting in cell death through osmotic lysis. Membrane sterol composition appears to play a role in the sensitivity of different algal species to the membrane-disrupting effects of KmTX. This sterol specificity also appears to be responsible for the apparent immunity of K. micrum to its own toxins. K. micrum has a unique sterol profile, shared only by the congeneric dinoflagellates Karenia brevis and K. mikimotoi, dominated by (24S)-4α-methyl-5α-ergosta-8(14), 22-dien-3β-ol (72% by weight). This sterol has recently been named gymnodinosterol. Analysis of the sterol content in Amoebophrya sp. infecting K. micrum showed gymnodinosterol also to be dominant (62%). This was not simply a reflection of retaining host lipid content because K. micrum contains octadecapentaenoic acid (18:5n3), largely in galactolipids of the chloroplast, whereas Amoebophrya sp. contained little to no 18:5n3. By having a sterol content similar to its host, Amoebophrya sp. is able to avoid cell lysis caused by the cytotoxic compounds produced by the host.

Fatty acid and sterol composition of two evolutionarily closely related dinoflagellate morphospecies from cold Scandinavian brackish and freshwaters

Little is known about the lipid composition of cold-adapted dinoflagellates. Here, we investigate Peridinium aciculiferum and Scrippsiella hangoei, two cold-adapted dinoflagellate morphospecies that present significant differences in general morphology, physiology and habitat, but which share identical ribosomal DNA (rDNA) sequences, indicating a recent evolutionary divergence between them. Despite their present taxonomic names, it is now known that these two genetically isolated morphospecies are evolutionarily related to the so-called Pfiesteria-like species. We have found that chloroplast-associated glycolipids from P. aciculiferum and S. hangoei contained a higher relative percentage of two polyunsaturated C18 fatty acids (octadecapentaenoic [18:5(n-3)] and octadecatetraenoic [18:4(n-3)] acids), and a lower relative percentage of hexadecanoic acid (16:0) than typically observed in previously examined warm-adapted dinoflagellate species. This suggests that these two fatty acids provided the requisite membrane fluidity for chloroplasts under a cold growth condition in these two organisms. A similar increase in 18:5(n-3) and 18:4(n-3) over warm-adapted species was also found in the phospholipids and triglycerides. P. aciculiferum and S. hangoei were also examined for the production of unique sterols that may serve as chemical biomarkers. Both were found to have a very similar sterol composition, consisting of two common dinoflagellate sterols, cholestanol and dinostanol, as dominant sterols, along with a number of other common dinoflagellate sterols as minor sterols. This demonstrated the absence of a specific steroidal biomarker but, more importantly, showed that these two morphospecies still produce very similar sterols despite having diverged in several other traits.

Transcription factor sterol regulatory element binding protein 2 regulates scavenger receptor Cla-1 gene expression.

The human scavenger receptor class B type I (Cla-1) plays a key role in cellular cholesterol movement in facilitating transport of cholesterol between cells and lipoproteins. Indirect evidence has suggested that Cla-1 gene expression is under the feedback control of cellular cholesterol content. To define the molecular mechanisms underlying such putative regulation, we evaluated whether Cla-1 is a target gene of the sterol regulatory element binding protein (SREBP) transcription factor family. Transient transfections demonstrated that SREBP factors induce Cla-1 promoter activity and that SREBP-2 is a more potent inducer than the SREBP-1a isoform. The 5'-deletion analysis of 3 kb of the 5'-flanking sequence of the Cla-1 gene, combined with site-directed mutagenesis and electrophoretic mobility shift assay, allowed identification of a unique sterol responsive element. SREBP-mediated Cla-1 regulation was confirmed in stably transfected human embryonic kidney 293 cells expressing the active form of SREBP-2 at incremental levels. In these cell lines, Cla-1 mRNA and protein levels were increased in direct proportion to the level of SREBP-2 expression. These findings provide evidence that SREBP-2, a key regulator of cellular cholesterol uptake through modulation of the expression of the low-density lipoprotein receptor gene, may influence cellular cholesterol homeostasis via regulation of Cla-1 gene expression.

A link between sterol biosynthesis, the cell wall, and cellulose in Arabidopsis.

A crucial role for sterols in plant growth and development is underscored by the identification of three Arabidopsis sterol biosynthesis mutants that exhibit embryonic defects: fackel (fk), hydra1 (hyd1), and sterol methyltransferase 1/cephalopod (smt1/cph). We have taken a dual approach of sterol profiling and ultrastructural analysis to investigate the primary defects underlying the mutant phenotypes. Comprehensive gas chromatography GC-MS analysis of hyd1 in comparison to fk reveals an abnormal accumulation of unique sterol intermediates in each case. Sterol profiling of the fk hyd1 double mutant provides genetic evidence that FK C-14 reductase acts upstream of HYD1 C-8,7 isomerase. Despite distinct differences in sterol profiles, fk and hyd1 as well as smt1/cph share ultrastructural features such as incomplete cell walls and aberrant cell wall thickenings in embryonic and post-embryonic tissues. The common defects are coupled with ectopic callose and lignin deposits. We show that all three mutants exhibit a deficiency in cellulose, but are not reduced in pectin and sugars of the cell wall and cytosol. The sterol biosynthesis inhibitors 15-azasterol and fenpropimorph also cause cell wall gaps in dividing root cells and a reduction in bulk cellulose, corroborating that the cell wall abnormalities are due to altered sterol composition. Our results demonstrate that sterols are crucial for cellulose synthesis in the building of the plant cell wall.