Viability loss of neem (Azadirachta indica) seeds associated with membrane phase behaviour.

Storage of neem (Azadirachta indica) seeds is difficult because of their sensitivity to chilling stress at moisture contents (MC) > or =10% or imbibitional stress below 10% MC. The hypothesis was tested that an elevated gel-to-liquid crystalline phase transition temperature (Tm) of membranes is responsible for this storage behaviour. To this end a spin probe technique, Fourier transform infrared microspectroscopy, and electron microscopy were used. The in situ Tm of hydrated membranes was between 10 degrees C and 15 degrees C, coinciding with the critical minimum temperature for germination. During storage, viability of fresh embryos was lost within two weeks at 5 degrees C, but remained high at 25 degrees C. The loss of viability coincided with an increased leakage of K+ from the embryos upon imbibition and with an increased proportion of cells with injured plasma membranes. Freeze-fracture replicas of plasma membranes from chilled, hydrated axes showed lateral phase separation and signs of the inverted hexagonal phase. Dehydrated embryos were sensitive to soaking in water, particularly at low temperatures, but fresh embryos were not. After soaking dry embryos at 5 degrees C (4 h) plus 1 d of further incubation at 25 degrees C, the axis cells were structurally disorganized and did not become turgid. In contrast, cells had a healthy appearance and were turgid after soaking at 35 degrees C. Imbibitional stress was associated with the loss of plasma membrane integrity in a limited number of cells, which expanded during further incubation of the embryos at 25 degrees C. It is suggested that the injuries brought about by storage or imbibition at sub-optimal temperatures in tropical seeds whose membranes have a high intrinsic Tm (10-15 degrees C), are caused by gel phase formation.     

EVOLUTION OF VEGETATIVE INCOMPATIBILITY IN FILAMENTOUS ASCOMYCETES. I. DETERMINISTIC MODELS

In ascomycetes vegetative incompatibility can prevent the somatic exchange of genetic material between conspecifics. It must occur frequently in natural populations, since in all species studied many vegetative compatibility groups (VCGs) are found. Using a population-genetic approach, this paper explores two possible selective explanations for the evolution of vegetative incompatibility in asexual fungi: selection by a nuclear parasitic gene, and selection by a harmful cytoplasmic element. In a deterministic model, assuming a random spatial distribution of VCGs in an infinitely sized population, it is found that neither of these forms of frequency-dependent selection can explain the large number of VCGs found in nature. The selective pressure for more VCGs disappears once a limited number of VCGs exist, because the frequency of compatible interactions decreases when the number of VCGs increases. In comparing the two selective explanations, selection by a cytoplasmic element seems a more plausible explanation than selection by a nuclear gene.     

On the asymmetry of sex: Evolution of mating types in isogamous populations

This paper is an analysis using population genetic models of the problem of the evolution of two mating types from an original situation in which all gametes are alike functionally (i.e. each gamete can fuse with any other gamete). It is shown that inbreeding depression due to fusion of identical gametes from the same clone is unlikely to play a significant role in this evolution. Evolution towards unipolarity in gamete recognition or adhesion (implying the existence of two mating types) requires a more than twofold disadvantage for bipolarity. However, the evolution of mating types is greatly facilitated if a pheromonal gamete attraction mechanism is already present. The models finally predict the evolution of a mating type “super gene” determining the various functions associated with mating.     

Reconstitution and fusogenic properties of Sendai virus envelopes

Sendai virus membranes were reconstituted by detergent dialysis, using the non-ionic detergents Triton X-100 and octyl glucoside. Membrane reassembly was determined by measuring the surface-density-dependent efficiency of resonance energy transfer between two fluorescent phospholipid analogues, which were co-reconstituted with the viral envelopes. The functional incorporation of the viral proteins was established by monitoring the ability of the reconstitution products to fuse with erythrocyte membranes, utilizing assays based on either resonance energy transfer or on relief of fluorescence selfquenching. The persistent adherence of residual Triton X-100 with the reconstituted membrane was revealed by an artificial detergent-effect on the resonance energy transfer efficiency and the occurrence of hemolysis of human erythrocytes under conditions where fusion does not occur. Properly reconstituted Sendai virus envelopes were obtained with octyl glucoside. The fusion activity of the viral envelopes was dependent on the initial concentration of octyl glucoside used to disrupt the virus and the rate of detergent removal. Rapid removal of detergent by dialysis against large volumes of dialysis buffer (ratio 1:850) or by gel filtration produced reconstituted membranes capable of inducing hemagglutination but significant fusion activity was not detected. By decreasing the volume ratio of dialysate versus dialysis buffer to 1:250 or 1:25, fusogenic viral envelopes were obtained. The initial fusion kinetics of the reconstituted viral membrane and the parent virus were different in that both the onset and the initial rate of fusion of the reconstituted membranes were faster, whereas the extents to which both particles eventually fused with the target membrane were similar. The differences in the initial fusion kinetics lead us to suggest that the details of the fusion mechanism between Sendai virus and the target membrane involve factors other than the mere presence of glycoproteins F and HN in the viral bilayer. Finally, the results also indicate that determination of the viral fusion activity in a direct manner, rather than by an indirect assay, such as hemolysis, is imperative for a proper evaluation of the functional properties retained upon viral reconstitution.     

A Diurnal Rhythm in Brown Adipose Tissue Causes Rapid Clearance and Combustion of Plasma Lipids at Wakening

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The dynamics of pAL2-1 homologous linear plasmids in Podospora anserina

A natural population of recently isolated Podospora anserina strains was screened for homologues of the linear longevity-inducing plasmid pAL2-1. Of the 78 wild-type isolates, 14 hybridised with a pAL2-1 specific probe, half of which contained a single plasmid and the other half multiple plasmid copies (plasmid family). All strains except one plasmid-containing strain, senesced normally. However, no inserted plasmid sequences were detected in the mitochondrial DNA, as was the case for the longevity-inducing pAL2-1 plasmid. Occasional loss of plasmids and of repeated plasmid sequences occurred during sexual transfer. Plasmid transmission was equally efficient for mono- and dikaryotic spores and was independent of the genetic background of the strains. Furthermore, horizontal transfer experiments showed that the linear plasmid could easily infect plasmid-free strains. Horizontal transfer was even observed between strains showing a clear vegetative incompatibility response (barrage). The linear plasmids are inherited maternally; however, paternal transmission was observed in crosses between confronted vegetative-incompatible strains. Paternal transmission of the plasmid was never observed using isolated spermatia for fertilisation, showing that mitochondrial plasmids can only gain access to maternal sexual reproductive structures following horizontal transfer. These findings have implications for both the function of vegetative incompatibility in fungi and for the mechanism of maintenance of linear plasmids. Received: 13 November 1997 / Accepted: 17 February 1998     

Storage behavior of Typha latifolia pollen at low water contents: Interpretation on the basis of water activity and glass concepts

Germplasm must be stored under optimal conditions to maximize longevity and efficiently maintain genetic resources. In order to identify optimal storage conditions, we investigated the effects of temperature (−5 to 45°C) and water content (<0.17 g H₂O g⁻¹ dry weight) on longevity of Typha latifolia L. pollen. Longevity was highest at water contents corresponding to storage relative humidity (RH) of 11‐15% which corresponded to the shoulder of water sorption isotherms. Also coinciding with this shoulder were abrupt changes in heat capacity of water present in the pollen. Consistent with changes in isotherms with temperature and the concept of critical RH for storage, optimum water contents increased with decreasing temperature. An attempt was made to explain the aging behavior according to the glass concept. The water content‐temperature combinations of optimal storage were found to be below the glass transition curve, indicating that optimum storage conditions are achieved when intracellular glasses are present. We also found a change in activation energy of aging in Arrhenius plots around T_g, demonstrating a change in aging kinetics when the glassy state is lost. We concluL that T_g curves cannot be used solely to predict precise conditions of optimum storage, but might be useful for predictions of storage longevity above optimum water contents. The data imply that too much drying reduces longevity and should be avoided, particularly when cryogenic storage is considered.