The partial purification and characterisation of gibberellin 2β-hydroxylases from seeds of Pisum sativum

The gibberellin (GA) 2-hydroxylases in mature and immature seeds of Pisum sativum have been partially purified and characterised. The enzymes are unstable when stored below pH 7.0 or in the absence of a thiol reagent. The optimum assay pH is between 7.4 and 7.8 and activity is dependent upon the presence of -ketoglutarate, Fe²⁺ and ascorbate. The 2-hydroxylase activities for GA₁, GA₄, GA₉ and GA₂₀ are chromatographically inseparable and correspond to a protein of M_r 44000. The rate of GA 2-hydroxylation varies according to substrate and some evidence indicates that the 2-hydroxylase activities for GA₁ and GA₄ and for GA₉ and GA₂₀ may reside in different proteins. During pea seed maturation, the specific activity of the enzyme(s) increases dramatically and reaches a maximum at a time when endogenous GA₉, GA₂₀, GA₂₉ and GA₅₁ are also at their greatest concentration. This correlation is not the result of substrate induction of enzyme activity. Since the GA 2-hydroxylases operate at maximal rate at low substrate concentrations they are incapable of rapidly 2-hydroxylating excessive quantities of (exogenously applied) GA₁ or GA₂₀. On the basis of the kinetic parameters of the GA 2-hydroxylase activities, a generalised model is discussed for the control of the steady-state levels of bioactive hormone under normal physiological conditions.Abbreviations DTE dithioerythritol - EDTA ethylenediaminetetraacetic acid - GA_n gibberellin A_n - HPLC high-performance liquid chromatography - HSS high-speed supernatant - LSS low-speed supernatant - PMSF phenylmethane sulphonyl fluoride     

Metabolism of gibberellins in a cell-free system from immature seeds of Phaseolus vulgaris L.

The biosynthetic steps from gibberellin A₁₂-aldehyde (GA₁₂-aldehyde) to C₁₉-GAs were studied by means of a cell-free system from the embryos of immature Phaseolus vulgaris seeds. Stable-isotope-labeled GAs were used as substrates and the products were identified by gas chromatography-mass spectrometry. Gibberellin A₁₂-aldehyde was converted to GA₄ via non-hydroxylated intermediates and to GA₁ via 13-hydroxylated intermediates. 13-Hydroxylation took place at the beginning of the pathway by the conversion of GA₁₂-aldehyde to GA₅₃-aldehyde. The conversion of GA₂₀ to GA₅ and GA₆ was also shown but no 2-hydroxylating activity was found. Endogenous GAs from embryos and testas of 17-dold seeds were re-examined by gas chromatography-selected ion monitoring using stable-isotopelabeled GAs as internal standards. Gibberellins A₉, A₁₂, A₁₅, A₁₉, A₂₃, A₂₄, and A₅₃ were identified for the first time in P. vulgaris, in addition to GA₁, GA₄, GA₅, GA₆, GA₈, GA₁₇, GA₂₀, GA₂₉, GA₃₇, GA₃₈ and GA₄₄, which were previously known to occur in this species. The levels of all GAs, except the 2-hydroxylated ones, were greater in the embryos than in the testas. Conversely, the contents of GA₈ and GA₂₉, both 2-hydroxylated, were much higher in the testas than in the embryos.Abbreviations GA_n gibberellin A_n - GC-MS gas chromatography-mass spectrometry - GC-SIM gas chromatography-selected ion monitoring - HPLC high-performance liquid chromatography - TLC thin-layer chromatography - m/z ion of mass     

The cytoskeleton of Cobaea seed hairs:

The cell wall of Cobaea scandens seed hairs developed in a characteristic sequence, with the deposition of a cellulose thread onto a pectic swelling layer was the final event. The cellulose thread was intracellularly accompanied by a band of 10–18 microtubules. During the formation of the swelling layer the microtubules were homogeneously distributed; they ran circumferentially normal to the cell axis. When cellulose-thread formation started, the microtubules became arranged in a helical band. The density of the microtubules varied during the different phases of development. The highest density was observed before cellulosethread formation and ranged from 6–15 m·m^-2. The length of the microtubules, 20–30 m, was determined by direct measurements, as well as estimated from the total microtubular length in a given area and the counted free ends. With the indirect immunofluorescence technique the microtubules of the band stained inhomogeneously. Those which were located at the edges of the band fluoresced more intensely than those of the central part. Attempts to visualize actin filaments in the hair cells with rhodaminyl-conjugated phalloidin resulted in a homogeneous staining of the area of the microtubular band, indicating that actin filaments may be present in this region. Though, in thin sections and dry-cleaved cells, filamentous structures were observed between the microtubules, caution is expressed that the observed fluorescence was, indeed, due to actin filaments. The role of the filamentous structures is discussed with respect to formation and maintenance of the microtubular band. Microtubules apparently did not cross coated pits which were visualized in the plasma membrane through the dry-cleaving technique.Abbreviations IFT indirect immunofluorescence technique - RP rhodaminyl-conjugated phalloidin - SEM scanning electron microscopy     

Toxic tetranortriterpenes of the fruit of Melia azedarach

Four new tetranortriterpenes, meliatoxins A₁, A₂, B₁ and B₂ have been isolated and identified as toxic constituents of the fruit of Melia azedarach L. var. australasica. Toxicity and pathological results confirm that the meliatoxins are responsible for most but not all of the symptoms resulting from the ingestion of whole fruit.     

Genetic specificity of nitrogen nutrition in leguminous plants

Summary Mineral nitrogen did not increase grain yield and seed protein levels ofVicia faba L. andLupinus luteus L. in field trials and pot experiments. Fixed N₂ was substituted by mineral nitrogen in these cases because of inhibition of N₂ fixation by mineral nitrogen. Contrary to these results mineral nitrogen increased grain yields and seed protein amounts ofLupinus albus L.,Pisum sativum L., andGlycine max. (L.) Merr. The nitrogen effect was caused at an early stage by saving energy due to inhibition of N₂ fixation (measurement of gas exchange by means of IRGA). In case of the N application after flowering grain, yields and seed protein levels increased because the mineral N was an additional nitrogen source for plants. At this stage the plants had ceased fixing atmospheric nitrogen. The high sink activity of growing fruits induced a lack of assimilates in nodules (determined by means of¹⁴CO₂ application). The N effect was therefore the consequence of the lower assimilate pool for supplying root nodules in these plants in comparison withVicia faba L. andLupinus luteus L. Hence it follows that response to mineral nitrogen can be a criterion for discovering more effective Rhizobium-host combinations.     

Establishment of microscale vegetation pattern in maritime chaparral after fire

We quantified microscale pattern in vegetation and seed assemblages along a 24 m transect before and for two years following a controlled burn in chamise (Adenostoma fasciculatum) chaparral in central coastal California. Our objective was to document scale-dependent correlation between pre-burn seed assemblages, pre-burn canopy cover, microtopography, soil temperatures during burning, post-burn seed assemblages and post-burn vegetation. Scale-dependent correlations among pre- and post-burn seed densities, maximum soil temperatures during burning, microtopography and post-burn vegetation were measured based on two-term local covariance analysis.Seed distribution varied among species prior to fire, with seeds of some annual species concentrated in gap areas. Maximum soil temperatures during burning ranged from less than 50 °C to 225 °C, and were generally lowest in gaps in the pre-burn canopy. These gaps were associated with local topographic depressions. After burning, readily germinable seeds were concentrated in or near gaps in the pre-burn canopy. Germination of different species was variously enhanced, diminished or unchanged by the passage of fire. Post-burn vegetation was very patchy, with some areas nearly devoid of seedlings and other areas, especially pre-burn canopy gaps, supporting numerous seedlings. Seedling recruitment patterns in the second year were generally highly correlated with patterns in pre-burn seed banks and first year vegetation. Although many species exhibited similar recruitment patterns, several different mechanisms may have been responsible for the origin of those patterns.     

Contribution of buried seeds to revegetation after eruptions of the volcano Usu,Northern Japan

Soon after the 1977–78 eruptions of the volcano Usu, there were created many gullies in which former topsoil, i.e., well-developed soil accumulated before the eruptions, was eroded in the crater basin, whereas the outside of the gully was covered with thick volcanic deposits. The short-dispersal-seed plants were the most abundant in the inside of gully where the former topsoil was exposed, however, they have not immigrated from external environments. The germination tests of the seeds extracted from the former topsoil demonstrated that viable seeds were buried at 1683.3/m² for 9 years after the eruptions. At least, 12 herbaceous species, e.g.,Rumex obtusifolius, Geum macrophyllum var.sachalinense andPoa annua, were derived from the buried seeds in the former topsoil. Their contribution to revegetation was estimated to be 40.0% in the inside of gully where the former topsoil was exposed in 1983, suggesting that buried seeds are one of the most important seed source for revegetation even in the case of a volcano which produced pumice and ash by eruption.     

Response of pondcypress growth rates to sewage effluent application

Cores were collected from dominant pondcypress trees growing in a swamp that had received sewage effluent for 7 yr and a nearby control swamp to determine the combined effects of changes in nutrient supply and hydrologic regime on tree growth. The cores were used to measure two indices of tree growth: basal area increment (BAI) and relative basal area increment (RBAI, which accounts for differences in growth due to the size of teh tree) between 1970–1983 while one swamp remained untreated and the other received weekly additions of sewage effluent from 1974–1981. Throughout the whole period, the mean BAI and RBAI of pond-cypress trees in the untreated swamp remained unchanged, ranging between 5.55–6.38 cm² yr^–1 and 1.09–1.27% yr^–1, respectively. In contrast, trees in the treated swamp increased their BAI approximately two-fold from 7.40 cm² yr^–1 prior to treatment to 14.83 cm² yr^–1 after the onset of treatment and maintained this rate of growth in the 2 yr period after cessation of treatment. Relative basal area increment showed a similar response, but the proportional increase due to treatment was less (1.5-fold factor) than for BAI. The response of pondcypress trees to the sewage effluent differed depending upon whether the trees were located in the deep or shallow water zones. Trees in the deep zone of the treated swamp had lower BAIs and RBAIs than those in the shallow zone during the treatment period, whereas in pre- and post-treatment periods growth indices were equal in both zones. No significant differences in growth between deep and shallow zones were observed during all three time periods in the control swamp.     

Evolutionary origin of aminoglycoside phosphotransferase resistance genes

Summary The protein sequences of seven 3-aminoglycoside phosphotransferases falling into the six identified types and three 6-aminoglycoside phosphotransferases were analyzed to give a rooted phylogenetic tree. This tree supports the origin of these groups of enzymes in an ancestor closely related to the actinomycetes, and that horizontal transfer of the resistance genes occurred, possibly via transposons. The implications for genetic engineering of a novel antibiotic are discussed.     

Ontogeny–a way forward for systematics, a way backward for phylogeny

In the history of biology, the term 'evolution' has carried a dual meaning, viz. ontogeny (the unfolding of the germ) versus phylogeny (descent with modification). A problem in modern biology is the question of whether it is ontogeny which creates phylogeny, or whether it is phylogeny which moulds ontogeny. The paper explores the relationship of ontogeny to phylogeny in the context of 'pattern cladism'. The conclusion is that the analysis of ontogeny provides a direct method for classification ('a way forward for systematics'), which is a logical prerequisite for a phylogenetic interpretation of ontogenetic sequences ('a way backward for phylogeny'). The ontogenetic process of growth, subdivision and differentiation is related to the 'morphogenetic tree theory' on the basis of Von Baer's "laws of individual development". This conceptual relation shows that ontogeny creates phylogeny in an upward direction within the morphogenetic tree, whereas phylogeny (by means of natural selection) moulds ontogeny in a downward direction. A conflict originates with the conventions of Linnaean classification if ontogenetic divergence is proposed as a causal agent in the origin of higher taxa. It is proposed to solve this conflict by viewing individual organisms (or reproductive communities) not as constituents, but as representatives of higher taxa.