The glycosphingolipid composition of the human hepatoma cell line,Hep-G2

The origin of plasma glycosphingolipids in normal individuals and the mechanisms by which tumor-associated glycosphingolipid antigens enter the plasma in patients with cancer are largely unknown. The Hep-G2 human hepatoma cell line retains many of the characteristics of differentiated hepatocytes including the ability to synthesize and secrete lipoproteins. Preliminary results indicated that newly synthesized Hep-G2 cell glycosphingolipids are coupled to the secreted lipoproteins. This suggests that this cell line may offer an interesting model for studying glycosphingolipid secretion, transfer, and shedding. We now report on the chemical and immunological characterization of Hep-G2 cell glycosphingolipids. Five major glycosphingolipids were purified and biochemically characterized: glycosylceramide, lactosyl ceramide, ceramide trihexoside, ganglioside GM3, and lactosyl sulfatide. Four additional minor components (3-fucosyl-lactosamine containing glycolipids, asialo GM2, galactosylgloboside, and ganglioside GM1) were identified using a combination of exoglycosidase digestion and immunostaining of thin-layer chromatography plates with specific carbohydrate binding proteins. This demonstrates that although this cell line synthesizes a limited number of major glycosphingolipids, it retains the ability to produce at least small amounts of structures in the lactoneo, globo, and ganglio series of glycosphingolipids. These studies show that it will be possible to investigate the mechanisms of secretion by Hep-G2 cells of different classes of these molecules such as neutral glycosphingolipids, gangliosides, and sulfatides.     

Plasmodium falciparum glycolipid synthesis: constant and variant molecules of isolates and of strains with differing knob and cytoadherence phenotype

Plasmodium falciparum-infected erythrocytes were metabolically labeled with tritiated glucosamine. Lipid extracts were analyzed by high-performance thin-layer chromatography to compare labeled molecules of eleven isolates from patients, six cytoadherent in vitro strains, and two knobbed and two knobless strains from Aotus monkeys. Up to nineteen labeled bands were identified. Glycolipid GL1, previously identified in Malayan Camp, was present in all isolates and strains. Other molecules, between CG and GM1 and between GM1 and GD1a, varied in mobility or presence. There was no apparent association between labeled molecules and the presence of knobs or the property of cytoadherence.     

<Emphasis Type="Italic">α</Emphasis>-Amylase inhibitor-1 gene from <Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> expressed in <Emphasis Type="Italic">Coffea arabica</Emphasis>plants inhibits α-amylases from the coffee berry borer pest

Background  

Coffee is an important crop and is crucial to the economy of many developing countries, generating around US70 billion per year. There are 115 species in the < i > Coffea < /i > genus, but only two, < i > C. arabica < /i > and < i > C. canephora < /i > , are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer ( < i > Hypotheneumus hampei < /i > ), is responsible for worldwide annual losses of around US70 billion per year. There are 115 species in the Coffea genus, but only two, C. arabica and C. canephora, are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer (Hypotheneumus hampei), is responsible for worldwide annual losses of around US500 million. The coffee berry borer exclusively damages the coffee berries, and it is mainly controlled by organochlorine insecticides that are both toxic and carcinogenic. Unfortunately, natural resistance in the genus Coffea to H. hampei has not been documented. To overcome these problems, biotechnological strategies can be used to introduce an α-amylase inhibitor gene (α-AI1), which confers resistance against the coffee berry borer insect-pest, into C. arabica plants.     

Characterization of galactosyl glycerolipids in the HT29 human colon carcinoma cell line.

Glycoglycerolipids constitute a family of glycolipids with apparently very restricted expression in human tissues. They have previously been detected only in the testis and the nervous system. In the present study, two glycoglycerolipids were isolated from the HT29 human colon carcinoma cell line. The glycoglycerolipids were structurally characterized as a monogalactosylglycerolipid (1-O-alkyl-2-O-acyl-3-O-(beta-galactosyl)-sn-glycerol) and a digalactosylglycerolipid (1-O-alkyl-2-O-acyl-3-O-(beta-galactosyl(1-4)alpha-galactosyl)-sn-glycerol) using NMR and mass spectrometry. This digalactosylglycerolipid has not previously been structurally characterized. When HT29 cells were allowed to differentiate into more enterocyte-like cells by culture in glucose-free medium, expression of both of these glycoglycerolipids was greatly diminished. The presence of glycoglycerolipids in a human colon carcinoma cell line indicates that expression of this family of glycolipids may not be as restricted as previously thought. Instead this class of glycolipids may serve as differentiation antigens in various normal tissues and in tumor development. The Galalpha1-4Gal epitope was previously identified as a receptor for bacterial adhesins and toxins. The finding that this epitope is also linked to a glycerolipid moiety opens up new possible roles for this carbohydrate receptor in intracellular signaling.     

Dynamic rearrangement of the filamentous actin network occurs during zoosporogenesis and encystment in the oomycete phytophthora cinnamomi

The organization of filamentous actin (F-actin) in living cells of the oomycete Phytophthora cinnamomi was determined during zoosporogenesis and zoospore encystment by microinjecting sporangia with fluorescently labeled phalloidin and observing resultant fluorescence by confocal microscopy. In multinucleate sporangia prior to the induction of cleavage, phalloidin labeling took the form of plaques which occurred mainly in the periphery of the sporangia. After induction of cleavage, phalloidin labeling showed that the plaques disappeared and that F-actin began to accumulate along the developing cleavage planes and around nuclei and water expulsion vacuoles. F-actin labeling was also observed near the plasma membrane in zoospores and young cysts but reverted to the plaque form in older cysts. Localization of F-actin close to the developing cleavage planes is consistent with the idea that actin microfilaments function in the positioning and expansion of the cleavage membranes. Observations of plaques of actin in living sporangia provide evidence that plaques are not aldehyde-induced fixation artifacts. Copyright 1998 Academic Press.     

Serglycin expression during monocytic differentiation of U937-1 cells

Serglycin is the major proteoglycan in most hematopoietic cells, including monocytes and macrophages. The monoblastic cell line U937-1 was used to study the expression of serglycin during proliferation and differentiation. In unstimulated proliferating U937-1 cells serglycin mRNA is nonconstitutively expressed. The level of serglycin mRNA was found to correlate with the synthesis of chondroitin sulfate proteoglycan (CSPG). The U937-1 cells were induced to differentiate into different types of macrophage-like cells by exposing the cells to PMA, RA, or VitD3. These inducers of differentiation affected the expression of serglycin mRNA in three different ways. The initial upregulation seen in the normally proliferating cells was not observed in PMA treated cells. In contrast, RA increased the initial upregulation, giving a reproducible six times increase in serglycin mRNA level from 4 to 24 h of incubation, compared to a four times increase in the control cells. VitD3 had no effect on the expression of serglycin mRNA. The incorporation of (35S)sulfate into CSPG decreased approximately 50% in all three differentiated cell types. Further, the (35S)CSPGs expressed were of larger size in PMA treated cells than controls, but smaller after RA treatment. This was due to the expression of CSPGs, with CS-chains of 25 and 5 kDa in PMA and RA treated cells, respectively, compared to 11 kDa in the controls. VitD3 had no significant effect on the size of CSPG produced. PMA treated cells secreted 75% of the (35S)PGs expressed, but the major portion was retained in cells treated with VitD3 or RA. The differences seen in serglycin mRNA levels, the macromolecular properties of serglycin and in the PG secretion patterns, suggest that serglycin may have different functions in different types of macrophages.