Special secretory cells in the soybean seed coat

Summary The seed coat of soybean (Glycine max L. Merr.) is of physiological interest for synthesis and transport of amino acids and photosynthates during embryo development. A transmission and scanning electron microscopic study to elucidate the structure of the seed coat disclosed a specialized convex area (antipit) appressed to a concave pit in the center of the abaxial surface of the cotyledon. The antipit, which lies on the inner surface of the seed coat at a medial point in the anterior to posterior direction of the seed, contained specialized secretory cells bounded by loose multi-layered cell walls. These cells were rectangular in the developing seed, varied in length, and contributed directly to the convex morphology of the antipit seen on the ventral surface of the seed coat. At maturity these cells assumed the shape of a cone, extending from the aleurone layer in a perpendicular array. The aleurone and cone cells contained numerous Golgi apparatus, laminated rough endoplasmic reticulum, secretory vesicles, and amyloplasts. Secretory vesicles arose directly from tubules of fenestrated trans cisternae of the Golgi apparatus. Mitochondria were clustered with the amyloplasts; stacks of lamellar cisternae of rough endoplasmic reticulum were associated with the nucleus and Golgi apparatus. The cellular contents, the interconnections by plasmodesmata, and the close physical association with the cotyledon suggested that the aleurone and cone cells may be involved in symplastic transport of nutrients for use by the developing embryo.This paper is dedicated to the memory of my parents, Joseph and Theresa Yaklich, who by their example taught me the value of work and the enjoyment of simple things.     

Golgi apparatus respond to the antitumor sulfonylurea N-(4-methylphenylsulfonyl)-N′-(4-chlorophenyl)urea (LY 181984) in sulfonylurea-susceptible but not in resistant cell lines

Summary Cell lines susceptible or resistant to the active antitumor sulfonylurea [N-(4-methylphenylsulfonyl)-N-(4-chlorophenyl)-urea] (LY 181984) were treated with 100 M sulfonylurea for 1 or 3 h followed by monensin for 1 h. With cell lines where growth was inhibited by the active sulfonylurea, swollen Golgi apparatus cisternae following treatment were fewer and smaller than in untreated cells. Overall the volume of monensin-responsive trans cisternae was reduced by about 50% to 75% in cells lines where the antitumor sulfonylurea was growth inhibitory. The swelling response was unaffected by sulfonylurea in sulfonylurea-unresponsive cells. The antitumor-inactive sulfonylurea [N-(4-methylphenylsulfonyl)-N-(phenyl)urea] (LY 181985) was without effect on cisternal swelling with both susceptible and resistant cell lines. The results suggest a response of the trans Golgi apparatus to the active antitumor sulfonylurea that resulted in reduced acidification of the trans Golgi apparatus cisternae. This response appears to be restricted to susceptible cell lines where growth was inhibited by the active antitumor sulfonylurea but not in resistant cell lines where growth was unaffected by the active antitumor sulfonylurea.     

Hormone- and growth factor-stimulated NADH oxidase

An NADH oxidase activity of animal and plant plasma membrane is described that is stimulated by hormones and growth factors. In plasma membranes of cancer cells and tissues, the activity appears to be constitutively activated and no longer hormone responsive. With drugs that inhibit the activity, cells are unable to grow although growth inhibition may be more related to a failure of the cells to enlarge than to a direct inhibition of mitosis. The hormone-stimulated activity in plasma membranes of plants and the constitutively activated NADH oxidase in tumor cell plasma membranes is inhibited by thiol reagents whereas the basal activity is not. These findings point to a thiol involvement in the action of the activated form of the oxidase. NADH oxidase oxidation by Golgi apparatus of rat liver is inhibited by brefeldin A plus GDP. Brefeldin A is a macrolide antibiotic inhibitor of membrane trafficking. A model is presented where the NADH oxidase functions as a thiol-disulfide oxidoreductase activity involved in the formation and breakage of disulfide bonds. The thiol-disulfide interchange is postulated as being associated with physical membrane displacement as encountered in cell enlargement or in vesicle budding. The model, although speculative, does provide a basis for further experimentation to probe a potential function for this enzyme system which, under certain conditions, exhibits a hormone- and growth factor-stimulated oxidation of NADH.     

Diversity of Xiphinema americanum-group Species and Hierarchical Cluster Analysis of Morphometrics

Of the 39 species composing the Xiphinema americanum group, 14 were described originally from North America and two others have been reported from this region. Many species are very similar morphologically and can be distinguished only by a difficult comparison of various combinations of some morphometric characters. Study of morphometrics of 49 populations, including the type populations of the 39 species attributed to this group, by principal component analysis and hierarchical cluster analysis placed the populations into five subgroups, proposed here as the X. brevicolle subgroup (seven species), the X. americanum subgroup (17 species), the X. taylori subgroup (two species), the X. pachtaicum subgroup (eight species), and the X. lambertii subgroup (five species).     

Golgi apparatus isolation and use in cell-free systems

Summary Recent advances in understanding the molecular mechanisms of membrane traffic to and through the Golgi apparatus have been predicated in large measure on the use of permeabilized animal cells, and on completely cell-free systems. These systems have included those addressing inter-Golgi apparatus membrane traffic, endoplasmic reticulum to Golgi apparatus traffic, and endocytotic events. Development of cell-free systems depends on the use of isolated fractions. Specificity is often achieved by using a compartment-specific assay so that the fractions employed can be very crude. More recently cell-free systems also have evolved which employ highly purified and well-characterized cell fractions. The latter may be utilized in the absence of a compartment-specific assay but may require employment of compartment-specific assays for validation. Central to development of cell-free systems for membrane analysis has been the availability of isolated Golgi apparatus, first from plants and later from animal tissues and cells. A major advantage of cell-free systems is that they are most clearly amenable to the investigation of molecular mechanisms of membrane trafficking.Dedicated to Hilton H. Mollenhauer on the occasion of his retirement     

The protein content and morphogenesis of zymogen granules

When zymogen granules, the secretion granules of pancreatic acinar cells, fill, secretory product is accumulated in immature granules, condensing vacuoles. Mature granules are formed when this product (protein) condenses into an osmotically inactive aggregate and, bulk water is expelled. This hypothesis for granule morphogenesis has two elements. The first is that immature granules are precursors to mature granules. The second is that a particular maturational event, condensation, which involves the aggregation of protein, takes place. These hypotheses lead to two straightforward predictions. One, that condensing vacuoles on average, should contain less protein than filled or mature granules. And two, that, due to condensation, mature granules should contain protein at a common concentration. In the current work, both of these predictions were tested using measurements of the protein content of individual granules acquired by X-ray microscopy. Neither prediction was affirmed by the experimental results. First, there was no distinguishable difference in the distribution of protein between immature and mature granules. Second, the protein concentration of mature granules varied widely between preparations, although granules from the same preparation had similar concentrations. From the data we conclude that: 1) mature granules and condensing vacuoles are different, though not necessarily unrelated, types of secretory vesicle, and not two forms of the same object; 2) as such, condensing vacuoles are not precursors to mature granules; 3) all granules do not contain protein at one particular concentration when full, or mature; 4) granule maturation does not involve a condensation step; 5) concentration is not determined by such physical limits as the space available for protein packing or condensation; and 6) the amount of protein contained is physiologically regulated.     

Resonance Raman resolution of a-, b- and c-type cytochromes in membrane vesicles of alkalophilic bateria

Resonance Raman spectroscopy has been used to obtain complete spectra of each individual cytochrome type — a, b and c — in the reduced state within membrane vesicle preparations from two species of obligately alkalophilic bacteria: Bacillus alcalophilus and Bacillus firmus RAB. The vibrational spectra, in the range 250–1700 cm^?1, were obtained with tunable dye laser excitation in the wavelength range 550–600 nm tuned to resonance with the appropriate reduced alpha band maximum for the cytochrome type of interest. The spectra reveal details which serve to characterize the specific type of cytochrome as well as to confirm the similarity of the heme prosthetic group to previously well-characterized cytochromes of the the a- b- or c-type. Preliminary evidence in support of heterogeneity of b-type, and possibly a-type cytochromes, or of heme-heme interaction within the membrane is presented.     

Axonal transport of muscarinic receptors in vesicles containing noradrenaline and dopamine-β-hydroxylase

Presynaptic muscarinic receptors labeled with [³H]dexetimide and noradrenaline in dog splenic nerves accumulated proximally to a ligature at the same rate of axonal transport. After fractionation by differential centrifugation, specific [³H]quinuclidinyl benzilate or [³H]dexetimide binding revealed a distribution profile similar to that of dopamine-β-hydroxylase and noradrenaline. Subfractionation by density gradient centrifugation showed two peaks of muscarinic receptors; the peak of density 1.17 contained noradrenaline and dopamine-β-hydroxylase whereas that of density 1.14 was devoid of noradrenaline. Therefore the foregoing experiments provide evidence that presynaptic muscarinic receptors are transported in sympathetic nerves in synaptic vesicles which are similar to those containing noradrenaline and dopamine-β-hydroxylase. This suggests a possible coexistence of receptor and neurotransmitter in the same vesicle.     

Studies on the damage to Escherichia coli cell membrane caused by different rates of freeze-thawing

Freeze-thawing of Escherichia coli cells caused a release of cell membrane components such as protein, phospholipids and lipopolysaccharides. A greater amount of release and a lesser extent of cell survival were seen in slow freeze-thawing than in rapid freeze-thawing. Several dehydrogenases in the cells were also freed. The mode of release was also dependent on the rate of freeze-thawing.The materials released by slow freeze-thawing were found to be mostly composed of outer membrane components, whereas the materials released by rapid freeze-thawing contained cytoplasmic as well as outer membrane components. The chemical composition of these fragments differed significantly from that of the original membranes. The relative content of cytoplasmic membrane-bound enzymes in these fragments also differed from that of the cytoplasmic membrane.The fragmentation was assumed to have resulted mainly from the crystallization of external water. In slow freeze-thawing, it was considered that the phase separation of the membrane phospholipid bilayer increased the possibility of outer membrane fragmentation. Rapid freeze-thawing caused cytoplasmic membrane damage to the cells as well as to the outer membrane. In rapid freeze-thawing, the effect of phase separation appeared to be small because of rapid passage through the transition temperatures.The presence of 10% glycerol completely inhibited the release of cellular materials and enzymes. Cell survival was maintained at a high level in the glycerol-treated samples whether freeze-thawed slowly or rapidly.     

Inverted pyramidal neurons and their axons in the neocortex of reeler mutant mice

Summary Inverted pyramidal neurons are very abundant in the cerebral cortex of the adult reeler mutant mouse. Two types of inverted pyramid are found in rapid Golgi impregnations. In the first type the axon starts from the base of the cell body and bends towards the white matter. In the second type, which is more common, the axon emerges from the apical dendritic tree and descends directly towards the white matter.Despite its abnormal topography, the site of origin of the axon in pyramids of the second type displays a normal differentiation, when analysed with the electron microscopic Golgi technique, suggesting that the ectopic initial axon segment is able to fulfil its normal functions.