Deficiencies of the müllerian ducts induced by norethindrone in the female chick embryo

Norethindrone produces two effects on müllerian ducts (MD) of female chick embryos. It induces the loss of the lower end of both ducts, as a result of a stop in their development, before 8 days. After 12 days NET causes regressions of the upper part of the MD particularly of the oviduct. NET like estrogens are the only known substances which present these both properties.     

Identification of novel TGF-β regulated genes with pro-migratory roles

Transforming growth factor-β (TGF-β) signaling plays fundamental roles in the development and homeostasis of somatic cells. Dysregulated TGF-β signaling contributes to cancer progression and relapse to therapies by inducing epithelial-to-mesenchymal transition (EMT), enriching cancer stem cells, and promoting immunosuppression. Although many TGF-β-regulated genes have been identified, only a few datasets were obtained by next-generation sequencing. In this study, we performed RNA-sequencing analysis of MCF10A cells and identified 1166 genes that were upregulated and 861 genes that were downregulated by TGF-β. Gene set enrichment analysis revealed that focal adhesion and metabolic pathways were the top enriched pathways of the up- and downregulated genes, respectively. Genes in these pathways also possess significant predictive value for renal cancers. Moreover, we confirmed that TGF-β induced expression of MICAL1 and 2, and the histone demethylase, KDM7A, and revealed their regulatory roles on TGF-β-induced cell migration. We also show a critical effect of KDM7A in regulating the acetylation of H3K27 on TGF-β-induced genes. In sum, this study identified novel effectors that mediate the pro-migratory role of TGF-β signaling, paving the way for future studies that investigate the function of MICAL family members in cancer and the novel epigenetic mechanisms downstream TGF-β signaling.     

Endogenous β-d-galactoside binding lectin during the expansion of the yolk sac in the developing chick embryo

Summary A lectin with an affinity for -d-galactoside-containing saccharides is present in the developing yolk sac from the chick embryo at stages from 2 to 7 days of incubation. This activity is present in the area vitellina (less differentiated) and the area vasculosa (more differentiated). In both areas, lectin activity increases significantly during the spreading of the yolk sac up to 5 days of incubation. At all of the stages studied lectin activity was significantly higher in the area vasculosa, as compared to the area vitellina.Lectins were purified by affinity chromatography and examined by SDS-PAGE. Under reducing conditions two components are evident. A more prominent band of subunit molecular weight of 14,200±100 for the area vitellina and 13,700±300 for the area vasculosa and a second band with molecular weight of about 68,000±700 and 68,000±1,200 for the area vitellina and area vasculosa respectively, were observed. The -d-galactoside-binding lectin appears to be similar if not identical to that of the early chick blastoderm.     

Detailed analysis of the δ-crystallin mRNA-expressing region in early development of the chick pituitary gland

Although δ-crystallin (δ-crys), also known as lens protein, is transiently expressed in Rathke's pouch (RP) of the chick embryo, detailed temporal and spatial expression patterns have been obscure. In this study, to understand the relationship between the δ-crys mRNA-expressing region and RP formation, we examined the embryonic expression pattern of δ-crys mRNA in the primordium of the adenohypophysis. δ-crys mRNA expression was initially found at stage 15 anterior to the foregut and posterior to the invaginated oral ectoderm. After RP formation, the δ-crys mRNA was expressed in the post-ventral region of RP and the anterior region of RP. δ-crys mRNA expression was then restricted to the cephalic lobe of the pituitary gland. From stage 20, the δ-crys and alpha-glycoprotein subunit (αGSU) mRNA-expressing regions were almost completely overlapping. The αGSU mRNA-expressing region is thought to be the primordium of the pars tuberalis, and these regions were overlapped with the Lhx3 mRNA-expressing region. The intensity of δ-crys mRNA expression gradually decreased with development and completely disappeared by stage 34. These results suggest that the embryonic chick pituitary gland consists of two different regions labeled with δ-crys and Lhx3.     

Identification of differentially expressed genes in the zebrafish hypothalamic–pituitary axis

The vertebrate hypothalamic–pituitary axis (HP) is the main link between the central nervous system and endocrine system. Although several signal pathways and regulatory genes have been implicated in adenohypophysis ontogenesis, little is known about hypothalamic–neurohypophysial development or when the HP matures and becomes functional. To identify markers of the HP, we constructed subtractive cDNA libraries between adult zebrafish hypothalamus and pituitary. We identified previously published genes, ESTs and novel zebrafish genes, some of which were predicted by genomic database analysis. We also analyzed expression patterns of these genes and found that several are expressed in the embryonic and larval hypothalamus, neurohypophysis, and/or adenohypophysis. Expression at these stages makes these genes useful markers to study HP maturation and function.     

Identification ofStreptomyces violaceoruber Tü22 genes involved in the biosynthesis of granaticin

A 50 kb region of DNA fromStreptomyces violaceoruber Tü22, containing genes encoding proteins involved in the biosynthesis of granaticin, was isolated. The DNA sequence of a 7.3 kb fragment from this region, located approximately 10 kb from the genes that encode the polyketide synthetase responsible for formation of the benzoisochromane quinone skeleton, revealed five open reading frames (ORF1-ORF5). The deduced amino acid sequence of GraE, encoded by ORF2, shows 60.8% identity (75.2% similarity) to a dTDP-glucose dehydratase (StrE) fromStreptomyces griseus. Cultures ofEscherichia coli containing plasmids with ORF2, on a 2.1 kbBamHI fragment, were able to catalyze the formation of dTDP-4-keto-6-deoxy-d-glucose from dTDP-glucose at 5 times the rate of control cultures, confirming that ORF2 encodes a dTDP-glucose dehydratase. The amino acid sequence encoded by ORF3 (GraD) is 51.4% identical (69.9% similar) to that of StrD, a dTDP-glucose synthase fromStreptomyces griseus. The amino acid sequence encoded by ORF4 shares similarities with proteins that confer resistance to tetracycline and methylenomycin, and is suggested to be involved in transporting granaticin out of the cells by an active efflux mechanism.     

How do avian embryos breathe? Oxygen transport in the blood of early chick embryos

1. Chick embryos with primary circulation, up to about 3 days of development, show no hemoglobin-mediated transport of oxygen. 2. In embryos with secondary circulation, between 3 and 6 days of incubation, the vascular area acts as the respiratory organ. Its efficiency in the oxygen uptake is less than that of the chorioallantois of later embryos. On the contrary, oxygen release to the tissues is highly efficient. 3. A full efficient hematic uptake of oxygen is reached at about the 6th incubation day, when chorioallantois acts as the embryonic respiratory organ. 4. The different respiratory mechanisms of developing chick embryo are closely related to the functional properties of the various hemoglobins which are produced during the embryonic life.     

Identification of novel α-gliadin genes

Ten novel α-gliadin genes (Gli-ta, Gli-turg1, Gli-turg2, Gli-turg3, Gli-turg4, Gli-turg5, Gli-turg6, Gli-cs1, Gli-cs2, and Gli-cs3) with unique characteristics were isolated from wheat (Triticum aestivumL.), among which Gli-cs1, Gli-cs2, Gli-cs3, and Gli-turg6 were pseudogenes. Gli-cs3 and nine other sequences were much larger and smaller, respectively, than the typical α-gliadins. This variation was caused by insertion or deletion of the unique domain I and a polyglutamine region, possibly the result of illegitimate recombination. Consequently, Gli-cs3 contained 10 cysteine residues, whereas there were 2 cysteine residues only in the other nine sequences. Gli-ta/Gli-ta-like α-gliadin genes are normally expressed during the development of seeds. SDS-PAGE analysis showed that in-vitro-expressed Gli-ta could form intermolecular disulphide bonds and could be chain extenders. A protein band similar in size to Gli-ta has been observed in seed extracts, and mass spectrometry results confirm that the band contains small molecular mass α-gliadins, which is a characteristic of the novel α-gliadins. Mass spectrometry results also indicated that the two cysteine residues of Gli-ta/Gli-ta-like proteins participated in the formation of intermolecular disulphide bonds in vivo.     

Identification of differentially expressed genes involved in the regression and development of the chicken Müllerian duct

Müllerian ducts of male chickens undergo regression around day 12 of incubation, but the underlining mechanisms remain unclear. The purpose of this study was to identify factors that contribute to regression of the Müllerian duct in the chicken. We first employed annealing control primer-based RT-PCR to screen candidate genes differentially expressed in the Müllerian ducts between male and female. Four differentially expressed genes (MSX2, GAL10, VCP and PLCH1) were partially sequenced. The expression of mRNA of the latter genes and MSX1 in the male and female Müllerian ducts were compared at 7.5, 8 and 9 days of incubation using semi-quantitative RT-PCR. The results indicated that both MSX1 and MSX2 mRNA was highly expressed in the male Müllerian duct at day 9 of incubation, whereas, PLCH1 mRNA was lower in the male duct at day 9 of incubation compared to that of the female duct. Although VCP mRNA was expressed in both left and right female Müllerian ducts, no expression was detected in the male duct. Whole mount in situ hybridyzation analysis showed that the expression of MSX1 and MSX2 mRNA were localized specifically in the mesenchymal cells of the male Müllerian duct at day 9 of incubation. In contrast, VCP mRNA expression was observed in both mesenchymal and epithelial cells of the female Müllerian duct but not detected in the male duct. These results suggest that both up-regulation of MSX1 and MSX2 mRNA expression is involved in the regression of the Müllerian duct in male chicken embryo, whereas VCP expression is involved in development of the female duct.     

Identification of Streptomyces violaceoruber Tü22 genes involved in the biosynthesis of granaticin

A 50 kb region of DNA fromStreptomyces violaceoruber Tü22, containing genes encoding proteins involved in the biosynthesis of granaticin, was isolated. The DNA sequence of a 7.3 kb fragment from this region, located approximately 10 kb from the genes that encode the polyketide synthetase responsible for formation of the benzoisochromane quinone skeleton, revealed five open reading frames (ORF1-ORF5). The deduced amino acid sequence of GraE, encoded by ORF2, shows 60.8% identity (75.2% similarity) to a dTDP-glucose dehydratase (StrE) fromStreptomyces griseus. Cultures ofEscherichia coli containing plasmids with ORF2, on a 2.1 kbBamHI fragment, were able to catalyze the formation of dTDP-4-keto-6-deoxy-d-glucose from dTDP-glucose at 5 times the rate of control cultures, confirming that ORF2 encodes a dTDP-glucose dehydratase. The amino acid sequence encoded by ORF3 (GraD) is 51.4% identical (69.9% similar) to that of StrD, a dTDP-glucose synthase fromStreptomyces griseus. The amino acid sequence encoded by ORF4 shares similarities with proteins that confer resistance to tetracycline and methylenomycin, and is suggested to be involved in transporting granaticin out of the cells by an active efflux mechanism. Dedicated to Professor Satoshi Ōmura, a pioneer in the field of antibiotics, on the occasion of his 60th birthday     

Toxicological implications of a commercial formulation of deltamethrin (Decis®) in developing chick embryo

A deltamethrin containing insecticide formulation (Decis®) was evaluated for its toxic potential in developing chick embryos. For the present study, three water emulsified concentrations of Decis® (12.5 mg L^?1, 25 mg L^?1, and 50 mg L^?1) were used. Fertilized eggs of Gallus domesticus were immersed in these three concentrations of the insecticide for 60 min at 37°C on day 0 of incubation and kept for incubation till embryonic day 7. Recovered embryos were evaluated for teratogenic and biochemical changes. The results revealed that administration of Decis® at its lower concentrations (12.5 mg L^?1 and 25 mg L^?1) did not show any significant teratological changes but the significant number of abnormal survivors was observed at 50 mg L^?1 of dose concentration when compared with vehicle-treated control. Among biochemical changes, total glycogen and RNA contents of embryos was significantly decreased at 25 mg L^?1 and 50 mg L^?1 of Decis® concentrations. Similarly, significant alteration (p ≤ .05) was observed in alanine transaminase activity at 50 mg L^?1 concentration of Decis®. Thus, the present study concluded that the no-effect-level for developmental toxicity for Decis® is below the concentration of 25 mg L^?1 under standard laboratory conditions.     

Identification of two Drosophila TGF-β family members in the grasshopper Schistocerca americana

Intercellular signaling molecules of the transforming growth factor- (TGF-) superfamily are required for pattern formation in many multicellular organisms. The decapentaplegic (dpp) gene of Drosophila melanogaster has several developmental roles. To improve our understanding of the evolutionary diversification of this large family we identified dpp in the grasshopper Schistocerca americana. S. americana diverged from D. melanogaster approximately 350 million years ago, utilizes a distinct developmental program, and has a 60-fold-larger genome than D. melanogaster. Our analyses indicate a single dpp locus in D. melanogaster and S. americana, suggesting that dpp copy number does not correlate with increasing genome size. Another TGF- superfamily member, the D. melanogaster gene 60A, is also present in only one copy in each species. Comparison of homologous sequences from D. melanogaster, S. americana, and H. sapiens, representing roughly 900 million years of evolutionary distance, reveals significant constraint on sequence divergence for both dpp and 60A. In the signaling portion of the dpp protein, the amino acid identity between these species exceeds 74%. Our results for the TGF- superfamily are consistent with current hypotheses describing gene duplication and diversification as a frequent response to high levels of selective pressure on individual family members.