Antibody response to sulfolipids in experimental tuberculosis

Live Mycobacterium tuberculosis H₃₇Rv, when injected into guinea pigs, induced antibodies to sulfolipids whereas antibodies were not detected in animals injected with heat killed cells. The antibody titre was found to be related to the degree of infection. A significant decrease in the titre was noted after streptomycin treatment, suggesting that antibodies were produced in response to an increasing bacterial load that occurred as the infection progressed.     

On the role of sulfolipids in mammalian metabolism

Summary Sulfolipids of mammalian origin include sulfosphingolipids, sulfoglycerolipids and steroid sulfates. Sulfosphingolipids (sulfogalactosylceramide) may be involved in sodium transport, interaction of opiates with their receptors, activation of oxygen radical generating system, and blood coagulation Factor XII. Sulfoglycerolipids and steroid sulfates may be involved in spermatogenesis and sperm capacitation.     

Metabolism of sulfolipids in isolated renal tubules from rat

Proximal-rich tubules were prepared from rat kidneys by using collagenase treatment. The isolated rat renal tubules were compared with the intact kidney on the following characteristics. (1) Composition of the sulfoglycolipid. (2) Sulfoglycolipid metabolism based on incorporation of [35S]sulfate or some properties of sulfoglycolipid metabolism, including the activities of anabolic and catabolic enzymes. The results indicated following characteristics of the isolated renal tubules in comparison to the kidney in vivo. (1) The sulfoglycolipid compositions are qualitatively similar, except that the content of glucosyl sulfatide, Gg3Cer II3-sulfate, and GM4 was slightly higher in the isolated tubules. (2) The apparent half-lives (15-55 min) of sulfoglycolipids in the isolated tubules could indicate the existence of a rapid turnover pool of these lipids. (3) The sulfotransferase and sulfatase activities related to sulfoamphiphiles in the renal tubule were similar to those reported for the whole kidney. Based on the above criteria, we conclude that the isolated rat renal tubule should be a useful metabolic system for clarification of the short-term physiological events, up to 90 min, of proximal tubular sulfoglycolipids. By using the present system, we showed that biosynthesis of the renal total sulfoglycolipid was significantly elevated in rats deprived of water for 24 h.     

Identification of the sulfolipids in the non-photosynthetic diatom Nitzschia alba

The four major sulfolipids in the non-photosynthetic marine diatom, Nitzschia alba, were isolated in pure form and their structures were established spectrometrically and by identification of their hydrolysis products as (a) 24-methylene cholesterol sulfate, (b) 1-deoxyceramide-1-sulfonate, (c) phosphatidyl sulfocholine (a sulfonium analogue of phosphatidylcholine) and (d) sulfoquinovosyl diglyceride. The major characteristic fatty acids of the sulfolipids were: for the deoxyceramide sulfonate, 16 : 0 (26%) and 16 : 1-delta3-trans (64%); for the sulfonium analogue, 14 : 0 (30%), 18 : 1 (12%), 18 : 2 (8%), 20 : 5 (27%) and 22 : 6 (4%); and for the sulfoquinovosyl diglyceride (two species, respectively), 14 : 0 (9%, 22%), 16 : 0 (16%, 28%), 18 : 1 (8%, 22%), 20 : 5 (42%, 23%) and 22 : 6 (14%, 2%). Traces of lyso-derivatives of sulfoquinovosyl diglyceride and phosphatidyl sulfocholine were also detected. The deoxyceramide sulfonate and the phosphatidyl sulfocholine represent novel membrane lipid components not previously detected in other organisms. They may however have a widespread distribution in marine diatoms and perhaps in marine organisms generally.     

Effects of triiodothyronine on the synthesis of sulfolipids by oligodendrocyte-enriched glial cultures

Glial cultures were obtained from the brains of 1-week-old rats and were grown in a chemically defined, serum-free medium. We investigated the development of oligodendrocytes in these cultures and the synthesis of sulfolipids in the presence and absence of triiodothyronine (T3) in the medium: (1) In the presence of T3, the incorporation of [35S]sulfate into sulfolipids exhibited a developmental profile which is comparable to that found in the developing brain in vivo. A sharp peak of sulfolipid synthesis was observed at day 5 in vitro, which is equivalent to day 12 after birth. As observed in vivo, the percentage of label incorporated into sulfogalactosyldiradylglycerols decreased with time in culture. (2) Addition of T3 to the medium stimulated sulfolipid synthesis by oligodendrocytes in a dose-related manner (optimal T3 concentration, 30 nM). The hormone also enhanced the rates of cholesterogenesis and lipogenesis but to a lesser extent than sulfolipid synthesis. (3) The temporary omission of T3 from the medium resulted in lower rates of sulfolipid synthesis that could not be restored by readdition of T3. This inhibitory effect was most pronounced if the hormone was omitted from the medium on days 2 and 3 in culture. (4) Omission of T3 also resulted in the development of fewer oligodendrocytes in the cultures. Our results show that T3 is essential for the development of oligodendrocytes in our neurone-free culture system. They also indicate that the stimulation of myelination by thyroid hormones can, at least partially, be explained as a direct effect of T3 on oligodendrocytes, independent of an effect of T3 on neuronal growth.     

Studies on the biosynthesis of sulfolipids in the Diatom Nitzschia alba.

Labeling of sulfolipids in Nitzschia alba was studied after growth of the cells in media containing L-[35S]cystine, L-[35S], L-[35S]cysteine, L-[35S]-methionine or a mixture of L-[Me-3H]methionine and L-[35S]methionine, [35S]Cysteine or [35S]cystine labeled the deoxyceramide sulfonate and the sulfonium analog, phosphatidylsulfocholine (and its lyso derivative) but not the sterol sulfate nor the sulfoquinovosyl diglyceride; [35S]methionine labeled only the phosphatidylsulfocholine and its lyso derivative. With the [35S]- and [Me-3H]methionine mixture (3H/35S ratio 1.0) the phosphatidylsulfocholine had a 3H/35 S ratio of 1.5 indicating that both sulfonium methyl groups were derived from methionine. Probable biosynthetic pathways for these novel sulfolipids are discussed.     

Hormone-specific responses and biosynthesis of sulfolipids in cell lines derived from mammalian kidney

The established cell lines isolated from mammalian kidney were characterized by its receptor activities against hormones and the ability to synthesize sulfolipids localized in the renal tubule.The level of 3′: 5′-cyclic AMP in JTC-12.P3 (monkey kidney) cells increased in 2 min as much as 2.5–5-fold on activation with 1.0 unit/ml of bovine parathyroid hormone or 1.9 units/ml of synthetic parathyroid hormone (1–34) resulting in intracellular cyclic AMP concentration of more than 40 pmol/mg protein. Prostaglandin E₁ (14 μM) and isopropylnorepinephrine (10 μM) were also found to increase the concentration of cyclic AMP by more than 30- and 9-fold, respectively. Addition in medium of calcitonin, arginine vasopressin, adrenocorticotropic hormone and glucagon caused no significant changes of cyclic AMP level in the cell.In contrast, MDCK, a cell line isolated from canine kidney, reacted to arginine vasopressin, isopropylnorepinephrine and prostaglandin E₁ and only slightly to parathyroid hormone. MDBK cell line derived from bovine kidney or fibroblast cell lines from rat lung and guinea pig kidney did not react to any of the hormones specific to kidney, i.e. arginine vasopressin, calcitonin or parathyroid hormone in the presence of theophylline. However, in the presence of 2 mM isobutylmethylxanthine, small but significant elevation of cellular cyclic AMP levels in response to calcitonin, arginine vasopressin, isopropylnorepinephrine and prostaglandin E₁ was observed.The cell lines JTC-12, MDCK and MDBK, when incubated with H₂³⁵SO₄, incorporated the isotope into sulfolipids assigned as sulfatides and ceramide dihexoside sulfate or in MDCK also into cholesterol sulfate.The results suggested that JTC-12, MDCK and MDBK cell lines are epithelial origin and also JTC-12 and MDCK originated most probably from renal tubular cells of cortex and medulla, respectively.     

Hormone-specific responses and biosynthesis of sulfolipids in cell lines derived from mammalian kidney.

The established cell lines isolated from mammalian kidney were characterized by its receptor activities against hormones and the ability to synthesize sulfolipids localized in the renal tubule. The level of 3':5'-cyclic AMP in JTC-12.P3 (monkey kidney) cells increased in 2 min as much as 2.5-5-fold on activation with 1.0 unit/ml of bovine parathyroid hormone or 1.9 units/ml of synthetic parathyroid hormone (1-34) resulting in intracellular cyclic AMP concentration of more than 40 pmol/mg protein. Prostaglandin E1 (14 micronM) and isopropylnorepinephrine (10 micronM) were also found to increase the concentration of cyclic AMP by more than 30- and 9-fold, respectively. Addition in medium of calcitonin, arginine vasopressin, adrenocorticotropic hormone and glucagon caused no significant changes of cyclic AMP level in the cell. In contrast, MDCK, a cell line isolated from canine kidney, reacted to arginine vasopressin, isopropylnorepinephrine and prostaglandin E1 and only slightly to parathyroid hormone. MDBK cell line derived from bovine kidney or fibroblast cell lines from rat lung and guinea pig kidney did not react to any of the hormones specific to kidney, i.e. arginine vasopressin, calcitonin or parathyroid hormone in the presence of theophylline. However, in the presence of 2 mM isobutylmethylxanthine, small but significant elevation of cellular cyclic AMP levels in response to calcitonin, arginine vasopressin, isopropylnorepinephrine and prostaglandin E1 was observed. The cell lines JTC-12, MDCK and MDBK, when incubated with H235SO4, incorporated the isotope into sulfolipids assigned as sulfatides and ceramide dihexoside sulfate or in MDCK also into cholesterol sulfate. The results suggested that JTC-12, MDCK and MDBK cell lines are epithelial origin and also JTC-12 and MDCK originated most probably from renal tubular cells of cortex and medulla, respectively.     

Visualization method for sulfolipids and its application to the determination of nanomole quantities of lipid sulfur

A simple and sensitive visualization method for sulfolipids on a thin-layer chromatogram is described. By spraying with an acidic solution of azure A, a complex was formed between an anionic sulfolipid and a blue cationic compound. After the unbound dye was washed out by brief soaking in methanol, sulfolipids were visualized as clear dark-blue bands on a light-blue background. As little as 0.5 nmol could be detected. Sulfolipid-dye complex was estimated by densitometry or colorimetric measurement after extraction with chloroform/methanol. For the quantitative determination of sulfolipids having long sugar chains, it is necessary to treat thin-layer chromatography plates with acetic anhydride before color development. Of the other tissue lipids not containing sulfuric acid ester that were tested none were stained significantly. A linearity of quantitative determination was observed over the range of 1-8 nmol.     

The sulfolipids 2'-O-acyl-sulfoquinovosyldiacylglycerol and sulfoquinovosyldiacylglycerol are absent from a Chlamydomonas reinhardtii mutant deleted in SQD1

The biosynthesis of thylakoid lipids in eukaryotic photosynthetic organisms often involves enzymes in the endoplasmic reticulum (ER) and the chloroplast envelopes. Two pathways of thylakoid lipid biosynthesis, the ER and the plastid pathways, are present in parallel in many species, including Arabidopsis, but in other plants, e.g. grasses, only the ER pathway is active. The unicellular alga Chlamydomonas reinhardtii diverges from plants like Arabidopsis in a different way because its membranes do not contain phosphatidylcholine, and most thylakoid lipids are derived from the plastid pathway. Here, we describe an acylated derivative of sulfolipid, 2'-O-acyl-sulfoquinovosyldiacylglycerol (ASQD), which is present in C. reinhardtii. Although the fatty acids of sulfoquinovosyldiacylglycerol (SQDG) were mostly saturated, ASQD molecular species carried predominantly unsaturated fatty acids. Moreover, directly attached to the head group of ASQD was preferentially an 18-carbon fatty acid with four double bonds. High-throughput robotic screening led to the isolation of a plasmid disruption mutant of C. reinhardtii, designated Deltasqd1, which lacks ASQD as well as SQDG. In this mutant, the SQD1 ortholog was completely deleted and replaced by plasmid sequences. It is proposed that ASQD arises from the sugar nucleotide pathway of sulfolipid biosynthesis by acylation of the 2'-hydroxyl of the sulfoquinovosyl head group. At the physiological level, the mutant showed increased sensitivity to a diuron herbicide and reduced growth under phosphate limitation, suggesting a role for SQDG and/or ASQD in photosynthesis as conducted by C. reinhardtii, particularly under phosphate-limited conditions.     

Quantification and fatty acid profiles of sulfolipids in two halophytes and a glycophyte grown under different salt concentrations

This study was aimed at understanding the role of sulfolipids in salt tolerance mechanisms of the halophytes Aster tripolium L., Compositae, and Sesuvium portulacastrum L., Aizoaceae, and of the glycophyte Arabidopsis thaliana (L.) Heynh., Brassicaceae. In Aster and Sesuvium the sulfolipid contents increased significantly under salt stress conditions (517 mM or 864 mM). In Arabidopsis, changes in sulfolipid contents were not observed (NaCl up to 100 mM). The fatty acid profile of sulfoquinovosyldiacylglycerol (SQDG) in Aster was modified with increasing NaCl concentrations. LC-MS analyses of sulfolipids from Aster and Sesuvium revealed the presence of 18:3/18:3 and 16:0/18:3 molecules. Obviously, the function of sulfolipids during salt stress differs between halophytic species and between halophytes and glycophytes where sulfolipid accumulation was not observed.     

Suppression of the antibody response to sulfolipids in leprosy patients by 4,4′-diaminodiphenyl sulfone (DDS)

Antibodies to sulfolipids were demonstrated in patients suffering from lepromatous leprosy. The antibody titre was found to decrease gradually on treatment with DDS. This effect was maximum for patients undergoing treatment for more than 1 year.     

Determination of lipid-bound sulfate by ion chromatography and its application to quantification of sulfolipids from kidneys of various mammalian species

A variety of procedures have been developed for determining the sulfate ester content of various biomolecules. Ion chromatography (IC), that is, quantitation of ionic substances by ion conductimetry after separation by anion-exchange chromatography, has been increasingly utilized for the determination of inorganic sulfate in clinical and environmental samples. We adopted suppressed-mode IC to the determination of lipid- or glycolipid-bound sulfate released by acid hydrolysis and found that it has the advantage of increased precision for wide concentration ranges (30 pmol to approximately micromol) and lack of interference from other lipids. To minimize deterioration of the separation column, the lipophilic constituents in the acid hydrolysate were removed by a two-phase partition system of chloroform-methanol-water. The inorganic sulfate was quantitatively extracted into the aqueous phase by replacing water with an alkaline buffer. By this method, the concentration of sulfolipids was determined in the kidney of mammals with various body mass. Sulfolipids were more concentrated in the kidney of smaller animals, which have higher maximum urine concentrating activity per gram of body mass, supporting the hypothesis of the function of sulfolipids as an ion barrier on the luminal surface of renal tubules.     

Changes in content and fatty acid profiles of total lipids and sulfolipids in the halophyte Crithmum maritimum under salt stress

In the halophyte Crithmum maritimum, the sulfolipid content increased considerably in the presence of NaCl. There were no significant changes in the total fatty acid composition of sulfolipids during salt treatment, except for linoleic and linolenic acids. In comparison to the control plants, sulfolipids in NaCl-treated plants showed a decrease in the percentage of unsaturated fatty acid (C18:3), and a corresponding increase in the percentage of unsaturated fatty acids (C18:2). As a whole, the data reported in this work suggest that sulfolipds may be one important aspect of strategies involved in salt tolerance of this halophyte.     

Differentiation of mycobacterial species by investigation of petroleum ether-soluble sulfolipids using thin-layer chromatography after incubation with [35S]sulfate

The method of detecting petroleum ether-soluble sulfolipids by thin-layer chromatography after incubation with [35S]sulfate is useful for differentiation between mycobacterial species. Rapidly growing mycobacteria, including two subspecies of Mycobacterium chelonei, were differentiated by this method. Most species of slowly growing mycobacteria were characterized by the pattern of distribution of radioactive sulfolipids in the thin-layer chromatograms. Mycobacterium nonchromogenicum was clearly differentiated from Mycobacterium terrae and Mycobacterium triviale by the presence of a sulfolipid. The latter two did not contain any significant amount of sulfolipids. Mycobacterium avium contained a large amount of sulfolipids, whereas Mycobacterium scrofulaceum did not contain any detectable sulfolipid. Rapidly growing, scotochromogenic mycobacteria, except for Mycobacterium flavescens, did not contain any remarkable amounts of sulfolipids.     

Characterization of sulfolipids of Mycobacterium tuberculosis H37Rv by multiple-stage linear ion-trap high-resolution mass spectrometry with electrospray ionization reveals that the family of sulfolipid II predominates

Mycobacterium tuberculosis, the causative agent of tuberculosis, is unique among bacterial pathogens in that it contains a wide array of complex lipids and lipoglycans on its cell wall. Among them, the sulfated glycolipid, termed the sulfolipid, is thought to mediate specific host-pathogen interactions during infection. Sulfolipids (SLs), including sulfolipid I (SL-I) and sulfolipid II (SL-II), are 2,3,6,6'-tetraacyltrehalose 2'-sulfates. SL-I was identified as a family of homologous 2-palmitoyl(stearoyl)-3-phthioceranoyl-6,6'-bis(hydroxyphthioceranoy1)trehalose 2'-sulfates and was believed to be the principal sulfolipid of M. tuberculosis strain H37Rv. We cultured and extracted sulfolipids using various conditions, including those originally described, and employed high-resolution multiple-stage linear ion-trap mass spectrometry with electrospray ionization to characterize the structure of the principal SL. We revealed that SL-II, a family of homologous 2-stearoyl(palmitoyl)-3,6,6'-tris(hydroxyphthioceranoy1)trehalose 2'-sulfates, rather than SL-I is the principal sulfolipid class. We identified a great number of isomers resulting from permutation of the various hydroxyphthioceranoyl substituents at positions 6 and 6' of the trehalose backbone for each of the SL-II species in the entire family. We redefined the structure of this important lipid family that was misassigned using the traditional methods 40 years ago.     

植物G蛋白与植物防卫反应

近年来, 植物G蛋白(包括异三聚体G蛋白和小G蛋白)的存在及其信号调控途径已经成为人们研究细胞信号转导过程的热点问题。从多种植物细胞中相继分离克隆出多个与动物G蛋白同源的编码植物G蛋白的基因, 并且植物G蛋白的种类和数量有其独特性。植物G蛋白在植物细胞跨膜信号转导中发挥重要的作用, 参与多种生命活动的调控。本文主要综述了植物G蛋白参与植物防卫反应调节作用的研究进展。