Manfred Ruddat

Isolation and Characterization of the Lipid Reserve Bodies, Spherosomes, from Layers of Wheat

Lipid reserve bodies (spherosomes) isolated from aleurone layers of wheat (Triticum aestivum L.) by combined differential and density gradient centrifugation were heterogeneous in size and density and distributed throughout continuous sucrose density gradients The lightest spherosomes sedimented as floating lipid (d < 10), and the heaviest spherosomes sedimented with a density near that for mitochondria (d > 1.18). While all classes of spherosomes were lipid-rich, the light spherosomes contained more triglycerides and the heavy spherosomes contained more phospholipids and protein Acid phosphatase activity of aqueous homogenates occurred mainly in the microsome-free supernatant with only trace levels of the enzyme associated with spherosomes Among the particulate fractions, only aleurone grains obtained by nonaqueous procedures or aleurone fragments had acid phosphatase activity in both biochemical and cytochemical assays. Cytochemical analysis showed that the acid phosphatase activity in the spherosome fractions was not due to spherosomes per se but to adhering membrane fragments. In biochemical studies, acid lipase and phospholipase D activities paralleled the distribution of acid phosphatase among the cell fractions, demonstrating an initial association with aleurone grains rather than with spherosomes Upon incubation with gibberellic acid (GA3), these enzymes disappeared from the aleurone grain fraction Our findings suggest that spherosomes of wheat aleurone are not lysosomal in nature but cellular repositories of lipids Aleurone grains appear to function as the main repository of hydrolytic enzymes in ungerminated caryopsis

Participation of gibberellin in the control of apical dominance in soybean and redwood.

SummaryLoss of apical dominance in soybeans and redwood was increased when the plants were treated with the growth retardant AMO-1618. Simultaneous application of gibberellin reduced the number of elongating buds and promoted growth of the first or second uppermost axillary bud, thus restoring apical dominance. It is concluded that gibberellin participates in the expression of apical dominance.

Interactions of Microbotryum violaceum (ustilago violacea) with its host plant Silene alba

Abstract Sex expression in Silene alba is determined genetically but can be changed in female plants upon infection with the heterobasidiomycete Microbotryum violaceum. This change is not caused by steroids, the classical plant growth regulators, nor by a diffusible morphogen produced and secreted by the fungus. Nor is the production of stamnes in genotypically female flowers caused by the transmission, incorporation and expression of a fungal plasmid since plants regenerated from diseased tissue of genotypically female S. alba did not yield stamen-producing flowers. Neither density gradient centrifugation nor agarose gel electrophoresis of endonuclease restricted DNA from M. violaceum revealed the presence of a plasmid. Southern blots of DNA from S. alba probed with labeled DNA of M. violaceum, however, indicated the presence of homologous, unique sequences absent in non-host plants. Since the same homologous sequences were identified in male and female S. alba, these DNA fragments are not homologous to the coding sequences for male sex expression in S. alba unless they represent genetic elements of the hypothetical gyndyioecious precursor. Two other aspects of the S. alba-M. violaceum interaction have yielded interesting results. M. violaceum grows as sporidia outside of the host, but as short hyphae in planta. The switch from sporidial to hyphal growth is mediated in vitro by hyphal growth factors (HGFs) isolated from aqueous host plant extracts as well as by α-tocopherol. In addition to changing the fungal growth form. HGFs may serve as host recognition factors. Siderophore mutants of M. violaceum that accumulated less rhodotorulic acid than wild type also showed reduced or no pathogenicity, indicating that siderophores are an important factor in the host-pathogen interaction.

Characterization of carotene accumulation inUstilago violacea using high-performance liquid chromatography

Quantitative analysis of carotene accumulation in white, pink, pumpkin, orange, and yellow haploid strains ofUstilago violacea by high-performance liquid chromatography indicated that specific patterns of carotene accumulation are primarily responsible for the white, pumpkin, orange, and yellow phenotypes. The yellow strains accumulated primarily β-zeacarotene and β-carotene. The white strains accumulated primarily the colorless carotene, phytoene, or did not accumulate any carotene at all. Carotene accumulation in pink haploid strains followed the same patterns as for the white, pumpkin, orange, or yellow strains. Pink diploid and disomic strains ofU. violacea with various parental combinations of the color mutations accumulated either cis-β-zeacarotene and β-carotene or only β-carotene. The pattern of carotene accumulation in conjunction with the available genetic information for the carotene loci inU. violacea was used as a basis for the construction of a new genetic model for carotene biosynthesis inU. violacea. The model employs three dehydrogenases and one cyclase for the synthesis of β-carotene from phytoene, and accounts for the carotene accumulation patterns of either cis-β-zeacarotene and β-carotene or lycopene, γ-carotene, and β-carotene.

Genetics of Ustilago violacea. XXXII. Genetic evidence for transposable elements

Crosses between Ustilago violacea mutant strains with different color phenotypes that were derived from the 1.A1 and 2.A2 laboratory strains yielded, as expected, bisectored teliospore colonies with the parental colors as well as the a-1 and the a-2 mating-types. Generally, wild teliospore collections usually produced sporidia of both mating-types, providing two-mating-type (TMT) strains. Occasionally, however, sporidia with only one mating-type allele, a-1 or a-2, were obtained from teliospores, providing one-mating-type (OMT) strains. Crosses between OMT and laboratory strains with different color phenotypes gave (1) bisectored teliospore colonies with the parental colors or colonies with a parental color and a nonparental color and (2) nonsectored colonies with the nonparental color or with the parental color. The frequencies for the occurrence of non-parental color ranged from 41% to 93%, depending on the strain. The yield of teliospore colonies was usually reduced for these crosses. In many of these teliospore colonies, morphologically-altered sporidia (MAS phenotype) were observed. The morphology and the size of the sporidia with the MAS phenotype differed from those of teliospore colonies of the crosses between the laboratory strains. In addition, these sporidia did not form conjugants. A cross involving the TMT strains C449 yielded the MAS phenotype as well as a high incidence of tetrad colonies with a nonparental color. The high degree of instability of the parental color phenotypes, and the high frequency of the appearance of nonparental color phenotypes as well as the appearance of the MAS phenotype, are in accord with the presence of active and inactive transposable elements in the OMT strains, TMT strains, and laboratory strains.

Characterization of the pigment in pink sporidial colonies of Ustilago violacea as cytochrome c

The pigment of a pink strain of Ustilago violacea 1.C429 was associated with the mitochondria isolated on a sucrose gradient. The extracted red pigment was identified as cytochrome c by spectral analysis and by oxidation with cytochrome c oxidase from rat liver mitochondria. The pink strain accumulated at least tenfold more cytochrome c than a white strain. Zn2+ enhanced cytochrome c production in a white strain by 75%.

Characterization of cytochrome c accumulation in white and pink strains ofUstilago violacea using low-temperature spectroscopy and amino acid analysis

The extent of cytochrome c accumulation in 46 pink and white strains ofUstilago violacea was determined using low-temperature spectroscopy. Pink strains accumulated approximately 14 times more cytochrome c than white strains. Cytochrome c was extracted and purified from two pink (2A2, 1.C429) and two white (15.10, 900-42.1) strains ofU. violacea and subjected to amino acid analysis. One pink (2A2) and one white (15.10) strain were genetically related; the others (900-42.1, 1.C429) were not. One white strain (900-42.1) contained spectrally distinct cytochrome c. Comparisons of the amino acid compositions of the cytochrome c from these four strains ofU. violacea using divergence calculations and computer-assisted cluster analysis indicated a high degree of relatedness for the two pink strains, a moderate degree of relatedness for the pink strains and white strain 15.10, and a low degree of relatedness for white strain 900-42.1 with the others. These results support the hypothesis that there are two distinct cytochrome c loci inU. violacea.

Allelopathic compounds, thelypterin A and B in the fern Thelypteris normalis

SummaryThe growth of Thelypteris normalis (C. Chr.) Moxley gametophytes is inhibited under T. normalis sporophytes. Competition for minerals, light, change in pH, or microbial inhibitors were experimentally eliminated as causes of the inhibition. This is the first demonstration of allelopathy between a sporophyte and gametophyte in a fern. Two inhibitors, thelypterin A and B, which were released from the roots of the Thelypteris sporophyte, were isolated and a bioassay for the inhibitors was devised. Thelypterin A gave an Ehrlich-positive reaction indicative of secondary aromatic amines and an ultraviolet absorption spectrum indicative of a heterocyclic ring. The inhibitors affected the growth of Thelypteris, Pteris and Phlebodium gametophytes.

Ultrastructure and Development of the Sexine in the Pollen Wall of Silene alba (Caryophyllaceae)

Sexine development in Silene alba (Caryophyllaceae) was investigated by light and electron microscopy. Earliest indications of wall pattern formation occur in the tetrad when a fibrous primexine arises near the microspore plasma membrane The protectum elaborated by the primexine seems to serve as a scaffold upon which, after tetrad dissolution, electron-dense particles from the intralocular cavity are deposited, forming the mature sexine Sporopollenin synthesis is associated with lamellae of unit membrane dimensions at the apertural pore regions. White-line lamellae probably contribute to further sporopollenin synthesis during final maturation stages Indirect evidence suggests that pattern control mechanisms may reside in microspore mother cells

Response of Conifers to Growth Retardants

Application of four growth retardants (B-995, AMO-1618, Phosfon-D, and CCC) to seedlings of Arizona cypress, coastal redwood, Douglas fir, and Coulter pine showed that: (a) growth of Arizona cypress and coastal redwood treated with B-995 or AMO-1618 was retarded, and this effect was partially reversed with a 10-ppm gibberellin spray; (b) Arizona cypress was stunted by Phosfon-D but was not affected by CCC; (c) results with Douglas fir were inconclusive because of bud dormancy on treated plants; (d) growth of Coulter pine was retarded by B-995 or Phosfon-D but was unaffected by AMO-1618 or CCC.