Immunolocalization of alkaloids and X-ray microanalysis of elements in lupin seeds

Summary Immunolocalization of alkaloids in lupin seeds (Lupinus spp.) has been performed by cryofixation and conventional methods. Alkaloids were localized in the protein bodies of the cotyledon cells. Some immunogold particles in the walls of these cells were also observed. There were no differences in the sites of localization between the two mentioned methods. X-ray microanalysis of elements showed the presence of P, Mg, S, and K in the protein bodies of cotyledon cells in lupin seeds. The role of K⁺ in alkaloids transport is discussed.     

Roles of Vibrio fischeri and Nonsymbiotic Bacteria in the Dynamics of Mucus Secretion during Symbiont Colonization of the Euprymna scolopes Light Organ

During light organ colonization of the squid Euprymna scolopes by Vibrio fischeri, host-derived mucus provides a surface upon which environmental V. fischeri forms a biofilm and aggregates prior to colonization. In this study we defined the temporal and spatial characteristics of this process. Although permanent colonization is specific to certain strains of V. fischeri, confocal microscopy analyses revealed that light organ crypt spaces took up nonspecific bacteria and particles that were less than 2 μm in diameter during the first hour after hatching. However, within 2 h after inoculation, these cells or particles were not detectable, and further entry by nonspecific bacteria or particles appeared to be blocked. Exposure to environmental gram-negative or -positive bacteria or bacterial peptidoglycan caused the cells of the organ's superficial ciliated epithelium to release dense mucin stores at 1 to 2 h after hatching that were used to form the substrate upon which V. fischeri formed a biofilm and aggregated. Whereas the uncolonized organ surface continued to shed mucus, within 48 h of symbiont colonization mucus shedding ceased and the formation of bacterial aggregations was no longer observed. Eliminating the symbiont from the crypts with antibiotics restored the ability of the ciliated fields to secrete mucus and aggregate bacteria. While colonization by V. fischeri inhibited mucus secretion by the surface epithelium, secretion of host-derived mucus was induced in the crypt spaces. Together, these data indicate that although initiation of mucus secretion from the superficial epithelium is nonspecific, the inhibition of mucus secretion in these cells and the concomitant induction of secretion in the crypt cells are specific to natural colonization by V. fischeri.     

Comparison of Models Describing Light Dependence of N2 Fixation in Heterocystous Cyanobacteria

The abilities of four models to describe nitrogenase light-response curves were compared, using the heterocystous cyanobacterium Nodularia spumigena and a cyanobacterial bloom from the Baltic Sea as examples. All tested models gave a good fit of the data, and the rectangular hyperbola model is recommended for fitting nitrogenase-light response curves. This model describes an enzymatic process, while the others are empirical. It was possible to convert the process parameters between the four models and compare N₂ fixation with photosynthesis. The physiological meanings of the process parameters are discussed and compared to those of photosynthesis.     

Stigmatic Self-Incompatibility and Mating Patterns in Trillium grandiflorum and Trillium erectum(Melanthiaceae)

Post-pollination processes governing mating patterns in Trillium,a well-known genus of insect-pollinated woodland herbs, arepoorly understood. Mechanisms influencing outcrossing were investigatedin T. grandiflorum and T. erectum, two widespread species nativeto eastern North America. In southern Ontario, Canada, the twospecies are often sympatric; they flower in early May, and arepollinated by different assemblages of insects. Controlled cross-and self-pollinations and structural observations of pollengermination and pollen tube growth were conducted to determinewhether the two species possess a self-incompatibility (SI)system and, if so, the specific site(s) of self-rejection. Controlledpollinations indicated that both species set significantly moreseeds from cross-pollination than self-pollination, implicatingthe action of SI. This was confirmed by structural studies whichdemonstrated that self-recognition and rejection reactions occurredon dry-type stigmatic papillae. Observations of pollen hydrationrevealed that self-rejection was rapid, being initiated within10 min of pollination and prior to pollen tube emergence. Finalself-rejection resulted in failure of pollen tube growth atthe base of stigmatic papillae. SI was expressed more weaklyin T. erectum and thereby resulted in considerable self-seedset in some individuals . Estimates of outcrossing rates usingallozyme markers indicated that T. erectum displayed a mixed-matingsystem whereas T. grandiflorum was more highly outcrossed. Structuralstudies of pollen traits indicated that the two species differedwith respect to the size of grains and their aggregation withimplications for pollen dispersal and mating. The ecologicaland evolutionary implications of the variable expression ofSI in Trillium are discussed. Copyright 2001 Annals of BotanyCompany Trillium grandiflorum, Trillium erectum, self-incompatibility, mating     

The Aspergillus nidulans carnitine carrier encoded by the acuH gene is exclusively located in the mitochondria

The location of the Aspergillus nidulans carnitine/acyl-carnitine carrier (ACUH) was studied. ACUH with a His-tag at its N-terminus was over-expressed in Escherichia coli and purified by Ni(2+) affinity chromatography. The purified protein was utilised to raise polyclonal antibodies which were characterised by Western blotting. For localisation studies A. nidulans T1 strain, that contains the acuH gene under control of the strong promoter alcA(p), was derived. Results obtained demonstrate the exclusively mitochondrial localisation of ACUH and therefore exclude the targeting of the acuH gene product to the peroxisomal membrane.     

Contrasting influences of glucuronidation and O-methylation of epicatechin on hydrogen peroxide-induced cell death in neurons and fibroblasts.

The purpose of this study was to examine the comparative mechanisms by which the dietary form of the flavonoid epicatechin and its predominant in vivo metabolite, epicatechin glucuronide, influence oxidative stress-induced cell death in fibroblasts and neurons. The results demonstrate the contrasting influences of in vivo glucuronidation and methylation on the bioactivity of epicatechin.     

Characterization of the structure and dynamics of amyloidogenic variants of human lysozyme by NMR spectroscopy

The structures and dynamics of the native states of two mutational variants of human lysozyme, I56T and D67H, both associated with non-neuropathic systemic amyloidosis, have been investigated by NMR spectroscopy. The (1)H and (15)N main-chain amide chemical shifts of the I56T variant are very similar to those of the wild-type protein, but those of the D67H variant are greatly altered for 28 residues in the beta-domain. This finding is consistent with the X-ray crystallographic analysis, which shows that the structure of this variant is significantly altered from that of the wild-type protein in this region. The (1)H-(15)N heteronuclear NOE values show that, with the exception of V121, every residue in the wild-type and I56T proteins is located in tightly packed structures characteristic of the native states of most proteins. In contrast, D67H has a region of substantially increased mobility as shown by a dramatic decrease in heteronuclear NOE values of residues near the site of mutation. Despite this unusual flexibility, the D67H variant has no greater propensity to form amyloid fibrils in vivo or in vitro than has I56T. This finding indicates that it is the increased ability of the variants to access partially folded conformations, rather than intrinsic changes in their native state properties, that is the origin of their amyloidogenicity.