Insect tissue culture systems: models for study of hormonal control of development

Summary The regulation of growth and development of insects is under endocrine control and involves both juvenile hormones and ecdysteroids. Neuropeptides are master regulators which control the secretion of these hormones. Most experiments in insect endocrinology have been conducted in vivo, but tissue culture methodology is playing an increasing role due to the great interest in simpler model systems for the study of complex processes that occur in vivo. The availability of appropriate media has allowed the culture of a variety of insect organs and cell lines of defined origin which have kept certain properties of the parent tissues. Tissue culture approaches have been useful for studying hormonal control of morphogenetic processes. Cell lines are particularly suited to the study of hormonally regulated mechanisms of macromolecular biosynthesis and gene expression. Thus, the value of in vitro analysis in studies of regulation of hormone production is now recognized. Results obtained from tissue culture allow more precise definition of the hormonal requirements of insect cells and tissues for growth and differentiation and might make possible the discovery of new growth regulators.     

Influence of red irradiation and of removal of integuments on the metabolism of (±) [2-14C]-abscisic acid by Lactuca sativa Grand Rapids achenes

Achenes of Lactuca saliva L. cv. Grand Rapids, imbibed for 6 h in water or in a 10 µ M solution of non-radioactive abscisic acid (ABA), were cultivated on (2-¹⁴C]-ABA (10 µ M ) for 40 to 90 h. Red irradiation (660 ± 2.5 nm, 5 min, 2 W m ^-2) or removal of integuments were carried out before transfer to (2-¹⁴C]-ABA. When both treatments were applied, irradiation preceded removal of integuments. Imbibition and culture took place in darkness at 24°C. Two acidic diethyl ether phases, which contained the free acids (free phase) and the acids released after mild alkaline hydrolysis, respectively, were isolated. They were analyzed by thin layer chromatography (TLC). as well as the remaining aqueous phase.
Both red irradiation and removal of integuments led to increased [2-¹⁴C|-ABA uptake. Application of ABA during imbibition partly limited the stimulating effect of red irradiation on radioactive ABA uptake. Red irradiation stimulated [2-¹⁴C|-ABA metabolism by achenes, favouring the formation of the polar compound found in the remaining aqueous phase. Removal of the integuments stimulated metabolism notably, leading to an increase of the radioactivity in the remaining aqueous phase. This treatment also induced the appearance of new metabolites in the free phase (compound believed to be 7'-hydroxy-ABA) as well as in the remaining aqueous phase. The glucose ester of ABA was the only representative compound of the ester phase. Irrespective of the experimental conditions, there was no classical oxidative metabolism indicating that oxygen was not the limiting factor.     

The unusual tegumental tissues of the Lunaria annua (Brassicaceae) seed: A developmental study using light and electron microscopy

With light, fluorescence, transmission electron, and environmental scanning electron microscopy we studied the development of the Lunaria annua L. (Brassicaceae) seeds in order to reveal basic anatomical information about the unusual tissues of these seeds. In particular the seed tegument tissues possess complex morphological aspects that are relevant to the biology and ecology of this plant. A sclerenchymatic tissue as the innermost layer of the teguments apparently offers robust protection for the embryo, yet is organized to be flexible. This tissue likely controls the passage of water from the tegumental layers towards the embryo. We report here the presence of tannins in the pre-sclerenchymatic layer of the unripe seed. The inner tegument also houses a spongy tissue with wide intercellular spaces. This tissue could impart buoyancy to the seeds, which possibly might be required for water transport. The structural features could indicate that Lunaria may have evolved in a Mediterranean environment, which is characterized by a long dry season, but with a large amount of rainfall concentrated in short periods. Probably, not only the typical enlarged and flattened fruits of Lunaria can easily float and be dispersed away from the mother plant, but also the seeds have this dispersal peculiarity after release from the silicules.     

Selective forces in the emergence of the seed habit

The evolution of the seed is one of the major events in the history of land plants. In this paper, we consider the suite of characters that define the seed habit, and discuss the probable selective pressures that produced each character. Our major conclusion is that most characters are a direct consequence of the origin of heterospory and of natural selection for propagules with larger food reserves.
Seeds are traditionally defined by the possession of integuments. However, some heterosporous pteridophytes possess integument-like structures. Therefore, integuments cannot explain the evolutionary success of seed plants. Rather, we believe that the decisive character in this success is related to pollination. Seed plants differ from other heterosporous lineages in the capture of microspores before dispersal of the 'megaspore'. Modern gymnosperms all possess mechanisms whereby the maternal sporophyte withholds resources from potential propagules that have not been pollinated and/or fertilized. This represents an increase in efficiency over Pteridophytic reproduction. Wind-pollination means the propagule is vulnerable to pathogens that mimic pollen, and pathogen pressures may have contributed to some seed characters.     

Angiosperm ovules: diversity, development, evolution

Background

Ovules as developmental precursors of seeds are organs of central importance in angiosperm flowers and can be traced back in evolution to the earliest seed plants. Angiosperm ovules are diverse in their position in the ovary, nucellus thickness, number and thickness of integuments, degree and direction of curvature, and histological differentiations. There is a large body of literature on this diversity, and various views on its evolution have been proposed over the course of time. Most recently evo–devo studies have been concentrated on molecular developmental genetics in ovules of model plants.

Scope

The present review provides a synthetic treatment of several aspects of the sporophytic part of ovule diversity, development and evolution, based on extensive research on the vast original literature and on experience from my own comparative studies in a broad range of angiosperm clades.

Conclusions

In angiosperms the presence of an outer integument appears to be instrumental for ovule curvature, as indicated from studies on ovule diversity through the major clades of angiosperms, molecular developmental genetics in model species, abnormal ovules in a broad range of angiosperms, and comparison with gymnosperms with curved ovules. Lobation of integuments is not an atavism indicating evolution from telomes, but simply a morphogenetic constraint from the necessity of closure of the micropyle. Ovule shape is partly dependent on locule architecture, which is especially indicated by the occurrence of orthotropous ovules. Some ovule features are even more conservative than earlier assumed and thus of special interest in angiosperm macrosystematics.     

Deposition and localization of lipid polyester in developing seeds of Brassica napus and Arabidopsis thaliana

Mature seeds of Arabidopsis thaliana and Brassica napus contain complex mixtures of aliphatic monomers derived from non-extractable lipid polyesters. Most of the monomers are deposited in the seed coat, and their compositions suggest the presence of both cutin and suberin layers. The location of these polyesters within the seed coat, and their contributions to permeability of the seed coat and other functional properties are unknown. Polyester deposition was followed over Brassica seed development and distinct temporal patterns of monomer accumulation were observed. Octadecadiene-1,18-dioate, the major leaf cutin monomer, was transiently deposited. In contrast, the saturated dicarboxylates maintained a constant level during seed desiccation, whereas the fatty alcohols and saturated omega-hydroxy fatty acids continually increased. Dissection and analysis of Brassica seed coats showed that suberization is not specific to the chalaza. Analysis of the Arabidopsis ap2-7 mutant suggested that suberin monomers are preferentially associated with the outer integument. Several Arabidopsis knockout mutant lines for genes involved in polyester biosynthesis (att1, fatB and gpat5) were examined for seed monomer load and composition. The variance in polyester monomers of these mutants is correlated with dye penetration assays. Furthermore, stable transgenic plants expressing promoter::YFP fusions showed ATT1 promoter activity in the inner integument, whereas GPAT5 promoter is active in the outer integument. Together, the Arabidopsis data indicated that there is a suberized layer associated with the outer integument and a cutin-like polyester layer associated with the inner seed coat.     

Passive vectoring of entomopathogenic fungus Beauveria bassiana among the wax moth Galleria mellonella larvae by the ectoparasitoid Habrobracon hebetor females

Females of the ectoparasitoid Habrobracon hebetor attack and envenomate numerous host individuals during oviposition. The vectoring of the entomopathogenic fungus Beauveria bassiana during the adhesion stage by ectoparasitoid females among the wax moth larvae Galleria mellonella was explored under laboratory conditions. Vectoring occurred both from infected parasitoids to wax moth larvae and from infected to healthy wax moth larvae by parasitoids. The efficacy of vectoring in both cases was dose dependent. Parasitoid females were unable to recognize infected larvae in a labyrinth test. In addition, the presence of H. hebetor females significantly (1.5–13 fold) increased the mycoses level in clusters of G. mellonella, with 40% of the larvae infected with fungal conidia. Envenomation by H. hebetor increased conidia germination on the cuticles of the wax moth larvae by 4.4 fold. An enhanced germination rate (2 fold) was registered in the n‐hexane epicuticular extract of envenomated larvae compared to that of healthy larvae. Both envenomation and mycoses enhanced the phenoloxidase (PO) activity in the integument of G. mellonella and, in contrast, decreased the encapsulation rate in hemolymphs. We hypothesize that changes in the integument property and inhibition of cellular immunity provide the highest infection efficacy of entomopathogenic fungi with H. hebetor.