Phosphatidylinositol synthase of Tetrahymena: inositol isomers as substrates in phosphatidylinositol biosynthesis and headgroup exchange reactions.

Phosphatidylinositol (PtdIns) synthase in microsomal fractions derived from Tetrahymena vorax was studied to determine its activity requirements. The suitability of inositol isomers as substrates for the synthase and in headgroup exchange reactions also was investigated. Tetrahymena PtdIn synthase activity was optimum in the presence of 2 mM MgCl2 plus 2 mM MnCl2, a pH of 7.

Subcellular distribution of Plp-40, a lipoprotein in a serotype A strain of Pasteurella multocida.

A 40-kDa lipoprotein (Plp-40) is expressed by serotype A strains of Pasteurella multocida in amounts which correlate with the amount of capsular material present. We hypothesized that Plp-40 is exposed at the outer surface of the outer membrane (OM) of the cell and is associated with the serotype A exopolysaccharide material. The objectives of the present study were to confirm the lipoprotein nature of Plp-40 and to determine its subcellular location.

Phospholipids of the differentiating species of Tetrahymena.

The whole-cell phospholipid composition of the six known polymorphic species of Tetrahymena has been examined by [(3)H]acetate and [(3)H]myristic acid radiolabeling, and by gas-liquid chromatography of total phospholipid-bound fatty acids. Five of the polymorphic species contained similar phospholipid profiles following radiolabeling in that phosphatidylethanolamine (PE) was the predominant phospholipid; however, in cells of Tetrahymena patula LFF, aminoethylphosphonolipid was present in amounts nearly equal to PE. Tetrahymena patula LFF contained an unusually large percentage of sphingolipid (16.

Sodium-dependent transport of [3H](1D)chiro-inositol by Tetrahymena.

The transport characteristics of (1D)chiro-inositol by the ciliate Tetrahymena were examined in competition studies employing [3H](1D)chiro-inositol. (1D)chiro-Inositol transport was competed by unlabeled (1D)chiro-inositol, myo-inositol, scyllo-inositol, and D-glucose in a concentration-dependent manner. Conversely, (1D)chiro-inositol competed for [3H]myo- and [3H]scyllo-inositol transport.

Identification of the inositol isomers present in Tetrahymena.

The inositol isomer composition of phosphoinositides, polyphosphoinositols, phosphatidylinositol-linked glycans, and glycosyl phosphatidylinositol-anchored proteins of logarithmic phase Tetrahymena vorax was determined by GC-MS analysis of trimethylsilylimadazole derivatives. The most abundant inositol found was the myo-isomer; however, appreciable percentages of scylloinositol were present in the free inositol pool, phosphatidylinositol-linked glycan fraction, and glycosyl phosphatidylinositol-anchored protein fraction. Trace quantities of chiro- and neo-inositols also were present.

Prevalence of a novel capsule-associated lipoprotein among pasteurellaceae pathogenic in animals.

Capsular serotype A strains of Pasteurella multocida of avian origin express a 40-kDa lipoprotein (Plp-40) thought to attach the extracellular polysaccharide to the cell surface. The objective of the present study was to assess the prevalence of Plp-40 in P. multocida strains of disparate serotypes and host origins, as well as other pathogenic members of the family Pasteurellaceae.

Phenotype switching in polymorphic Tetrahymena: a single-cell Jekyll and Hyde.

For nearly half a century, phenotype switching in the group of polymorphic species of the ciliate genus Tetrahymena has been the subject of investigations of the underlying mechanisms, the accompanying biochemical and structural changes, and the evolution of polymorphic survival strategy. Beginning with the pioneering systematic studies by Furgason in 1940 of hymenostome ciliates, the experimental approach rapidly expanded to include investigations of growth, nutrition, physiology, morphology, and morphogenesis in the polymorphic species. Recently, with progress in elucidation of the novel signaling ligand and identification of elements of the subsequent signal transduction cascade, in addition to the growing catalog of intracellular events associated with differentiation in these unicellular eukaryotes, we have begun to address the mechanistic basis of polymorphism.

A complex of iron and nucleic acid catabolites is a signal that triggers differentiation in a freshwater protozoan.

The polymorphic ciliated protozoan Tetrahymena vorax can undergo differentiation from the microstomal form, which normally feeds on bacteria and other particulate matter, into the macrostomal cell type, which is capable of ingesting prey ciliates. The process is triggered by exposure of the microstome to an inducer contained in stomatin, an exudate of the prey. To establish the identity of the signal, stomatin was fractionated by combinations of cation exchange, HPLC, and TLC, and the fractions were assayed for biological activity.

The medical department in military operations other than war. Part II: Medical Civic Assistance Program in Southeast Asia.

It is a short trip by air from Bangkok, Thailand, to Phnom Penh, Cambodia, but a dramatic contrast. A Medical Civic Assistance Program (MEDCAP) visit was conducted in conjunction with Exercise Flash Canoe 97, the first U.S.

The medical department in military operations other than war. Part I. Planning for deployment.

Many military deployments are "military operations other than war" (MOOTW), a spectrum of assignments less than all-out combat. The corresponding medical support requirement differs from conventional military medical combat support and also from customary civilian medical practice. Hence, medical planners will use different doctrine and planning tools than are used in civilian facilities or on field training exercises when tasked for MOOTW activities.

Sodium-dependent uptake of [3H]scyllo-inositol by Tetrahymena: incorporation into phosphatidylinositol, phosphatidylinositol-linked glycans, and polyphosphoinositols.

[3H]Scyllo-inositol was taken up by Tetrahymena cells through a sodium-dependent pathway wherein unlabeled scyllo- and myo-inositol competed for uptake. d-Glucose was a competitor of [3H]myo-inositol uptake, but did not appear to compete for [3H]scyllo-inositol uptake. Transport of [3H]scyllo- and [3H]myo-inositol was inhibited when sodium was removed from the labeling buffer and by phlorizin, an inhibitor of sodium-dependent transporters.

Derivation of extracellular polysaccharide-deficient variants from a serotype A strain of Pasteurella multocida.

The production of serotype A extracellular polysaccharide is thought to be associated with expression of an approximately 40-kDa lipoprotein (P1p-40) present on the outer surface of Pasteurella multocida strains of avian origin. The tendency of certain strains to undergo colonial dissociation concomitantly with serial passaging on laboratory growth media was exploited to derive two variant strains exhibiting the capsule-deficient phenotype from a heavily capsulated parental strain. Assessments of colonial consistency, iridescence, gentian violet binding, and hyaluronidase sensitivity were consistent with cellular observations indicating little or no capsulation of derivative strains.

Evidence for early signaling events in stomatin-induced differentiation of Tetrahymena vorax.

The mechanism of stomatin-induced differentiation of Tetrahymena vorax was investigated by in vivo protease degradation of cell surface proteins, the direct measurement of products formed from the activation of phospholipase C, and the use of an array of signal transduction inhibitors/activators. The data indicate that a surface-exposed protein is required for stomatin to signal the cells to differentiate and that the cells are committed to the differentiation pathway within two hours after exposure to stomatin. Analysis of radiolabeled polyphosphoinositols and inositol lipids from control and stomatin-treated populations in the presence of 10 mM LiCl were consistent with a rapid activation of phospholipase C.

Relationship between serotype A encapsulation and a 40-kDa lipoprotein in Pasteurella multocida.

Eleven serotype A encapsulated and nonencapsulated strains of Pasteurella multocida were examined with regard to lipoprotein content. Relative amounts of an approximately 40-kDa lipoprotein (Plp-40) were found to correlate directly with the degree of encapsulation in that heavily encapsulated strains exhibited the greatest amounts, while nonencapsulated strains possessed little or no Plp-40.

Polysaccharide inducible outer membrane proteins of Bacteroides xylanolyticus X5-1.

Little is known about how bacteria degrade structural polysaccharides or the regulatory systems that control this degradation. Bacteroides xylanolyticus X5-1 is a Gram-negative, anaerobic bacterium that can grow on structural polysaccharides such as xylan and pectin. In order to determine the response of this organism to specific substrates,B.

Effects of dichloroisoproterenol on macromolecular synthesis and differentiation in Tetrahymena vorax.

The effect of dichloroisoproterenol on macrostomal cell formation in Tetrahymena vorax was examined and the drug was found to be 50% inhibitory at a concentration of 88 microM. Cellular uptake and incorporation of a variety of radiolabelled precursors was monitored in the presence of dichloroisoproterenol. The results demonstrate a strong, concentration-dependent inhibitory effect on RNA and protein biosynthesis, with a lesser inhibition observed for lipid biosynthesis.

Military medicine in operations other than war. Part II: Humanitarian relief missions for Naval Reserve fleet hospitals.

The U.S. military has a long tradition of providing emergency humanitarian assistance to civilian populations overwhelmed by natural disasters or civil strife.

Military medicine in operations other than war. Part I: Use of deployable medical systems facilities to assist in urban crises and enhance reserve medical training.

Navy medicine is challenged to provide ongoing and realistic training for medical personnel who will be deployed to provide battlefield care. Current events in this country suggest that the need for realistic training could be met while simultaneously providing support to civilian communities with great need. Deployable Medical Systems hospital modules could be set up in areas where the medical care system is overwhelmed and provide real patient management on the deployment platform.

Cytodifferentiation in Tetrahymena vorax is linked to glycosyl-phosphatidylinositol-anchored protein assembly.

The role of glycosyl-PtdIns (GPI)-anchored proteins in the cytodifferentiation of Tetrahymena vorax was examined. Labelling of cells with [3H]myristate or [3H]palmitate followed by electrophoresis showed an array of proteins carrying covalently bound lipids. Electrophoresis of protein from cells labelled with the GPI-anchor components [3H]Ins and [14C]ethanolamine revealed three polypeptides on fluorograms which have apparent molecular masses of approx.

Phosphatidylinositol-linked glycans and phosphatidylinositol-anchored proteins of Tetrahymena mimbres.

The insoluble residue from Tetrahymena mimbres cells that had been preincubated in vivo for 2 h with [3H]myristic acid and then exhaustively delipidated with organic solvents retained radioactivity, principally in material which migrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 10-14 kDa. This material was extractable from the delipidated cell residue with organic solvents known to solubilize phosphatidylinositol glycans (PI glycans). The same material could also be labeled with [3H]inositol, [14C]glucosamine, and [3H] ethanolamine.

Phosphatidylinositol glycan formation and utilization by the ciliate Tetrahymena mimbres.

Ryals et al. (Ryals, P.E.

Lipid modification during cytodifferentiation of Tetrahymena vorax. Whole cell phospholipids and triacylglycerols of microstomal and macrostomal phenotypes.

Microstomal cells of the ciliate Tetrahymena vorax V2S can be induced to undergo cytodifferentiation to form an alternate phenotype known as the macrostomal cell; however, sublines of T. vorax exist that respond differently to methods that induce macrostomal cell formation. The phospholipid- and triacylglycerol-bound fatty acid compositions of microstomal and macrostomal cells of a high-transforming subline (designated 3-C) were determined and compared to similar data from cells of a low-transforming subline (designated Ala).

Fatty Acid Acylated Proteins of the Halotolerant Alga Dunaliella salina.

The unicellular, wall-less alga Dunaliella salina has been shown to contain an array of proteins modified by the covalent attachment of fatty acids. Myristic acid (14:0) comprised approximately 80% by weight of the protein-linked acyl groups in samples derived from cells cultured in medium containing 1.7 molar NaCl and 93% in samples from cells grown in medium containing 3.

Protein acylation in Tetrahymena.

Examination of exhaustively delipidated Tetrahymena mimbres cells by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the presence of several protein bands containing covalently linked fatty acids. Palmitic (16:0) and stearic (18:0) acids together accounted for approximately 90% of the protein-linked acyl chains, with myristic acid (14:0) comprising most of the remainder. Each of these three fatty acids was present mainly in alkali-stable linkage, indicating that unlike most other systems examined, fatty acids are attached to proteins of Tetrahymena principally by amide bonds.

Alterations of the composition and size of the free fatty acid pool of Tetrahymena responding to low-temperature stress.

Cells of Tetrahymena mimbres (formerly T. pyriformis NT-1) in midlogarithmic growth under isothermal conditions (at 39 degrees C) contained a very small, compositionally discrete pool of free fatty acids, principally (60.6% of the total free fatty acid mass) palmitic and stearic acids.