Heterotopic expression of the Xl-Fli transcription factor during Xenopus embryogenesis: modification of cell adhesion and engagement in the apoptotic pathway.

In the Xenopus laevis embryo, the overexpression of the Xl-FLI protein, a transcription factor of the ETS family, provokes severe developmental anomalies, which affect anteroposterior and dorsoventral polarities, optic cup formation, head cartilage morphogenesis, and erythrocyte differentiation. It has been proposed that these effects could be correlated to modifications of cell adhesion properties and/or to an increased engagement of cells in the apoptotic pathway during early development (Remy et al., Int. J. Dev. Biol. 40, 577-589, 1996). To address these questions, we have first analyzed the behavior of cells overexpressing the protein in both aggregation and adhesion assays. We observe perturbations of cell-cell interactions as well as perturbations of cell adhesion and spreading on fibronectin and extracellular matrix (ECM). Second, we have analyzed apoptosis of cells overexpressing the Xl-FLI protein, by testing DNA fragmentation, caspase-3 activity and by performing TUNEL assay. We show that Xl-Fli overexpression results in the appearance of hallmarks of apoptosis, including exclusion of cells from the interior of the embryo, internucleosomal fragmentation of DNA and dose-dependent induction of caspase-3, resulting in the hydrolysis of poly(ADP-ribose) polymerase. In addition, a dominant-negative mutation of BMPs receptors decreases the effects of Xl-Fli overexpression, suggesting that a modification of the BMP signalling could be responsible for increased apoptosis. The latter appears to affect predominantly ventral and ventrolateral regions of the embryo.

Immunocytochemical localization of Bombyx-PTTH-like molecules in neurosecretory cells of the brain of the migratory locust, Locusta migratoria. A comparison with neuroparsin and insulin-related peptide.

Using a monoclonal antibody directed against a synthetic pentadecapeptide corresponding to the N-terminus of the prothoracicotropic hormone (PTTH) of Bombyx mori, we report the presence of immunoreactive molecules in a large number of median neurosecretory cells of the pars intercerebralis of the migratory locust, Locusta migratoria. These cells correspond to the A1 cell type which we show to contain also neuroparsins, a family of predominant neurohormones of the migratory locust. In contrast, PTTH-like molecules are absent from A2 cells of the pars intercerebralis which contain Locusta insulin-related peptide (LIRP). Developmental studies show the presence of PTTH-related substances in neurosecretory cells of Locusta migratoria from late embryogenesis to adult development, including ageing vitellogenic female adults.

cDNAs from neurosecretory cells of brains of Locusta migratoria (Insecta, Orthoptera) encoding a novel member of the superfamily of insulins.

From neurohaemal lobes of corpora cardiaca of Locusta migratoria a 5-kDa peptide has been isolated and its sequence established [see the accompanying paper, by Hietter et al. (1990) Eur. J. Biochem. 187, 241-247]. We have designed oligonucleotide probes from the peptide sequence of this molecule and screened a library prepared from mRNA of the neurosecretory cell region of the brain of this insect. Several positive cDNAs were isolated, the combined nucleotide sequences of which predict a large precursor of 145 residues (15770 Da) containing the newly isolated 5-kDa peptide. The peptide is flanked by regions homologous to the A and B chains of the superfamily of insulins. The overall organization of the precursor is as follows: signal peptide/domain homologous to the B chain of insulins/C (connecting)-peptide (corresponding to the newly isolated 5-kDa peptide)/domain homologous to the A chain of insulins. The numbers and relative positions of the cysteines of the Locusta peptide are equivalent to those of the other members of the insulin superfamily and most of the hydrophobic core residues are conserved.

Isolation and structure of two novel 6-kDa dimeric peptides from the corpora cardiaca of the insect Locusta migratoria. Molecular mass determination by mass spectrometry.

We have isolated two major 6-kDa peptides from extracts of corpora cardiaca of adult females of Locusta migratoria. These peptides have been characterized by peptide sequencing and liquid secondary-ion mass spectrometry. They are structurally related dimers, one (6278.5 Da) being a homodimer (A-A chains), the other (6280.5 Da) being a heterodimer (A-B chains). A 60% similarity exists between the A and B chains. Both peptides have been chemically synthesized and the synthetic compounds appeared to be identical to the native ones. Polyclonal antibodies raised against each of these peptides demonstrated that they were contained within the secretory granules of the intrinsic cells of the glandular lobes of the corpora cardiaca. The physiological significance of these two peptides is unknown but, using the synthetic peptides, we are currently probing their biological role.

[Ultrastructural study of the normal development of the oocytes and of the follicle cells in Locusta migratoria (L.) (Orthoptera) (author's transl)]. / Etude ultrastructurale du développement normal des ovocytes et des cellules folliculaires chez l'orthoptère Locusta migratoria (L.).

Vitellogenesis in Locusta migratoria is studied by electron microscopy, and may be divided into two major periods: a. the first stage of growth (oocyte length from 0,8 mm to 1,8 mm) is characterized by protein synthesis and storage in the rER, by transfer of material of nucleolar origin into the oocyte perinuclear cytoplasm as well as by pinocytotic activity giving rise to granulo-membranous structures. At this stage the apical part of the follicle cells differenciates as attested by mitochondrial accumulation and microvilli development; b. the second period of vitellogenesis (oocyte length from 1,8 to 6,8 mm) is characterized by accumulation of hemolymphatic material which partly separates the follicle cells and progressively detaches the follicular epithelium from the oocyte. The intense uptake of this material by oocyte leads to the formation of typical yolk globules. The material of nucleolus origin disappears from the cytoplasm at the beginning of this stage. The secretory activity of the follicular epithelium starts by the end of this second period (oocyte length 4 mm), leading vitelline membrane formation, and intensifies when the chorion is formed. The secretory material is excreted in soluble form. In the vicinity of the oocyte, this material is submitted to a series of complex modifications: it precipitates again in granular form, progressively unites and finally leads to the "scale-like" structures which are characteristic of the chorion.

The follicle cell epithelium of maturing ovaries of Locusta migratoria: a new biosynthetic tissue for ecdysone.

Follicle cells of maturing ovaries of Locusta migratoria are demonstrated to synthesize the moulting hormone ecdysone (2beta,3beta,14alpha,22R,25-pentahydroxy-5beta-cholest-7-en-6-one). Studies of secretory kinetics under in vitro conditions show that the intensity of hormone secretion is strictly dependent on the stage of maturation of the excised ovaries.