A differential proteomics study of cerebrospinal fluid from individuals with Niemann-Pick disease,Type C1

Niemann-Pick, type C1 (NPC1) is a fatal, neurodegenerative disease, which belongs to the family of lysosomal diseases. In NPC1, endo/lysosomal accumulation of unesterified cholesterol and sphingolipids arise from improper intracellular trafficking resulting in multi-organ dysfunction. With the proximity between the brain and cerebrospinal fluid (CSF), performing differential proteomics provides a means to shed light to changes occurring in the brain. In this study, CSF samples obtained from NPC1 individuals and unaffected controls were used for protein biomarker identification. A subset of these individuals with NPC1 are being treated with miglustat, a glycosphingolipid synthesis inhibitor. Of the 300 identified proteins, 71 proteins were altered in individuals with NPC1 compared to controls including cathepsin D, and members of the complement family. Included are a report of 10 potential markers for monitoring therapeutic treatment. We observed that pro-neuropeptide Y (NPY) was significantly increased in NPC1 individuals relative to healthy controls; however, individuals treated with miglustat displayed levels comparable to healthy controls. In further investigation, NPY levels in a NPC1 mouse model corroborated our findings. We posit that NPY could be a potential therapeutic target for NPC1 due to its multiple roles in the central nervous system such as attenuating neuroinflammation and reducing excitotoxicity.     

GENETIC EVIDENCE FOR MULTIPLE ORIGINS OF DIOECY IN THE HAWAIIAN SHRUB WIKSTROEMIA (THYMELAEACEAE)

Dioecy is unusually common in the Hawaiian Islands, yet little is known about the evolutionary biology of this breeding system. A native shrub, Wikstroemia, has an unusually diverse array of breeding systems: two forms of dioecy, cryptic and morphological dioecy, as well as hermaphroditism (perfect flowers). The existence of two forms of dioecy is significant for three reasons: 1) the presence of cryptic unisexuals that are functionally unisexual, but retain the appearance of hermaphroditism in both sexes, is strong evidence for the ancestral status of hermaphroditism; 2) the production of nonfunctional pollen, by female cryptic unisexuals, is a new instance of a phenomenon which has previously been reported for a few other species; 3) the two forms of dioecy are morphological markers which are useful in hybridization studies for tracing the genetic basis of their inheritance. Crosses were made between cryptically unisexual individuals (C), between morphologically unisexual individuals (M), and between the two types of unisexuality. The offspring of crosses between individuals with the same sex type usually resulted in offspring with that sex type, but most of the progeny of between-sex type crosses were, unexpectedly, perfect-flowered hermaphrodites. These results show that genetic control of sex determination is not homologous in all populations, suggesting that dioecy has evolved at least twice in Hawaiian Wikstroemia. The genetic data further suggest that males are the heterozygous sex.     

Ovine fetal adaptations to chronically reduced urine flow: preservation of amniotic fluid volume

Mann, Stephanie E., Mark J. M. Nijland, and Michael G. Ross.Ovine fetal adaptations to chronically reduced urine flow: preservation of amniotic fluid volume. J. Appl.Physiol. 81(6): 2588-2594, 1996.Adequateamniotic fluid (AF) volume is maintained by a balance of fetal fluidproduction (lung liquid and urine) and resorption (swallowing andintramembranous flow). Because fetal urine is the principle source ofAF, alterations in urine flow and composition directly impact AFdynamics. Intra-amniotic 1-desamino-8-D-argininevasopressin (DDAVP) is rapidly absorbed into fetal plasma and induces amarked fetal urinary antidiuresis. To examine the effect ofintra-amniotic- DDAVP-induced fetal urinary responses on AF volume andcomposition, six chronically prepared ewes with singleton fetuses(gestation 128 ± 2 days) were studied for 72 h after a singleintra-amniotic DDAVP (50-µg) injection. After DDAVP, fetal urineosmolality significantly increased at 2 h (157 ± 13 to 253 ± 21 mosmol/kg) and remained elevated at 72 h (400 ± 13 mosmol/kg). Urinary sodium (33.0 ± 4.5 to 117.2 ± 9.7 meq/l)and chloride (26.0 ± 2.8 to 92.4 ± 8.1 meq/l) concentrations similarly increased. AF osmolality increased (285 ± 3 to 299 ± 4 mosmol/kgH₂O), although there was no change in fetalplasma osmolality (294 ± 2 mosmol/kg). Despite a 50% reductionin fetal urine flow (0.12 ± 0.03 to 0.05 ± 0.02 ml ^· kg^{1 ·} min¹at 2 h and 0.06 ± 0.01 ml ^· kg^{1 ·} min¹after 72 h), AF volume did not change (693 ± 226 to 679 ± 214 ml). There were no changes in fetal arterial blood pressures, pH,PCO₂, orPO₂ after DDAVP. We conclude the following. 1)Intra-amniotic DDAVP injection induces a prolonged decrease in fetalurine flow and increases in urine and AF osmolalities. 2) Despite decreased urine flow, AFvolume does not change. We speculate that, in response to DDAVP-inducedfetal oliguria, reversed intramembranous flow (from isotonic fetalplasma to hypertonic AF) preserves AF volume.

     

A PHYLOGENETIC ANALYSIS OF THE SYNUROPHYCEAE USING MOLECULAR DATA AND SCALE CASE MORPHOLOGY

The phylogeny of the Synurophyceae was investigated by parsimony analysis of scale case characters and small-sub unit (18S) ribosomal RNA (rRNA) sequence data. Analysis of 1 eustigmatophycean (outgroup), 3 chrysophycean, and 10 synurophycean 18S rRNA sequences corroborated the inference from ultrastructural information that the Synurophyceae is a monophyletic assemblage . Tessellaria vol-vocina, which had been tentatively proposed as a member of the Synurophyceae, was confirmed as the earliest lineage within the Synurophyceae by both the molecular analysis and an evaluation of published ultrastructural data. A second set of analyses investigated the relationships among Tessellaria volvocina, 6 Synura species, and 10 Mallomonas species/varieties, with particular reference to the validity of current classifications of the Synurophyceae and the characters upon which they are based. The molecular and scale case phylogenies were not totally resolved but were largely congruent. The data sets were combined to produce another phylogeny, which showed greater resolution. The combined phylogeny weakly supported our representatives of Synura and Mallomonas as monophyletic groupings and also upheld several of the sections within these genera that are recognized by current classifications. However, some changes to the classification and delineation of these genera are recommended and predicted. Both our 18S rRNA sequence and scale case data sets were more appropriate for examining the branching order among the more closely related text rather than resolving the deeper branching points of the synurophycean phylogeny .     

Genetic variability of plant characters in the partial inbreeder Collinsia heterophylla (Scrophulariaceae)

Quantitative genetic variability for six characters was estimated in four populations of the annual plant, Collinsia heterophylla, with estimated selling rates ranging from 0.36 to 0.68. Based on large samples of parental plants, all four populations showed significant genetic variation for two or more characters, and there was no sign that the more selfing populations had lower amounts of genetic variance or lower heritability values. Autogamous fruit set rates in the absence of pollinators also showed significant heritability in one population.     

Meningococcal pilin: a glycoprotein substituted with digalactosyl 2,4-diacetamido-2,4,6-trideoxyhexose

Neisseria meningitidis pili are filamentous protein structures that are essential adhesins in capsulate bacteria. Pili of adhesion variants of meningococcal strain C311 contain glycosyl residues on pilin (PilE), their major structural subunit. Despite the presence of three potential N -linked glycosylation sites, none appears to be occupied in these pilins. Instead, a novel O -linked trisaccharide substituent, not previously found as a constituent of glycoproteins, is present within a peptide spanning amino acid residues 45 to 73 of the PilE molecule. This structure contains a terminal 1-4-linked digalactose moiety covalently linked to a 2,4-diacetamido-2,4,6-trideoxyhexose sugar which is directly attached to pilin. Pilins derived from galactose epimerase ( galE ) mutants lack the digalactosyl moiety, but retain the diacetamidotrideoxyhexose substitution. Both parental (#3) pilins and those derived from a hyper-adherent variant (#16) contained identical sugar substitutions in this region of pilin, and galE mutants of #3 were similar to the parental phenotype in their adherence to host cells. These studies have confirmed our previous observations that meningococcal pili are glycosylated and provided the first structural evidence for the presence of covalently linked carbohydrate on pili. In addition, they have revealed a completely novel protein/saccharide linkage.     

Can a Ca2+ pump in the endoplasmic reticulum of the Lepidium root be the trigger for rapid changes in membrane potential after gravistimulation?

Since gravistimulation is followed by alterations in the external current symmetry (Behrens et al., 1982), the effect of gravistimulation on cellular membrane potential was investigated using conventional glass microelectrode techniques. The resting potential of statocytes in a vertically oriented root is approx. -118 mV. Upon gravistimulation, the membrane potential is temporarily depolarized (lag time = 2 s) to a potential of approx. -93 mV. This depolarization is only observed in statocytes located on the physically lower root flank while those on the corresponding upper flank become weakly hyperpolarized (approx. -13 mV). These results reflect altered ion fluxes across the plasma membrane. The perception of gravistimulus was suggested to result from a pressure of the amyloplasts on the distal endoplasmic reticulum (ER) of the statocytes (Sievers and Volkmann, 1972). A causal relationship between changes in ER-amyloplast interactions and the rapid alterations in plasma membrane potential described above is not known. A candidate for such an intracellular messenger is Ca2+. As a first step in establishing the validity of such an assumption, we have isolated ER membranes from roots. When incubated with micromolar concentrations of Ca2+, the vesicular membrane fraction accumulates Ca2+. The accumulation is ATP-dependent and -specific and is directly coupled to ATP hydrolysis since a protonophore shows no inhibitory effect. Thus, in analogy to the sarcoplasmic reticulum of muscle, regulation of an ER-localized Ca2+ compartment might be an important step in such complex processes as stimulus-transduction in gravitropism.     

Effects of prolonged omnilateral gravistimulation on the ultrastructure of statocytes and on the graviresponse of roots

Statocytes of vertically growing roots of Lepidium sativum L. exhibit a strict polarity: The nucleus is positioned near the proximal periclinal cell wall, amyloplasts are sedimented on a complex of rough endoplasmic reticulum (ER) consisting of parallel cisternae near the distal periclinal cell wall.When 24 h old, vertically grown roots are rotated for an additional 20 h on a horizontal clinostat, this polarity is destroyed. Furthermore, the prolonged omnilateral stimulation leads to a damage of the statocytes, which in some cases ends in the self-destruction of the sensitive cells. The different components of the ultrastructural respones of the statocytes are: Displacement of the nucleus; changes in amount and distribution of the ER; loss of amyloplast starch; confluence of lipid droplets to large aggregates: a considerable increase of the lytic compartment. In addition, even anticlinal cell walls may be lysed up to small stumps. As all these effects are clearly restricted to the statocytes, only these cells are able to respond to the continuously changing direction of the gravity vector, thus perceiving gravity as such.After being exposed horizontally, the graviresponse of rotated roots is delayed as compared to the controls. About 20% of the rotated roots do not respond (curve) at all, but grow perpendicular in relation to the gravity vector. Perception of gravity is inevitably correlated with the polarity and the integrity of the statocytes.Abbreviation ER endoplasmic reticulum A preliminary report was presented at the Fall Meeting of the German Society for Cell Biology in Salzburg, Austria, September 1979 (Hensel and Sievers 1979)This paper represents part of a dissertation (D 5) of W. H.     

Ultrastructure of gravity-perceiving cells in plant roots

The root cap is the gravity-perceiving organ of plant roots. The central statenchyma consists of polarly organized statocytes which are characterized by sedimentable amyloplasts and by a striking distal endoplasmic reticulum (e.r. complex). During the normal downwards orientated growth of the root, the amyloplasts are sedimented onto the e.r. complex. Some observations indicate that the amyloplasts stress the e.r. complex by their sedimentation. The stress possibly influences the structural and functional state of the e.r. membranes. Therefore, graviperception is probably a function of differential stress on the e.r. complexes. The amyloplasts are partly or totally separated from the e.r. complex under experimental conditions such as deviations of the roots from the perpendicular between 30 and 180 degrees, rotation of the roots on the horizontal klinostat at 2 rev/min and at 55-120 rev/min, and placing the roots into an electric field at 2000-3000 V/cm. These are important facts for an understanding of the nature of graviperception. Spacelab experiments may help to verify the interpretation of results obtained in the experiments in simulated weightlessness.