Regulation of the expression of the sn-glycerol-3-phosphate dehydrogenase gene in Drosophila melanogaster

P element-mediated transformation has been used to investigate the regulation of expression of the sn-glycerol-3-phosphate dehydrogenase gene of Drosophila melanogaster. A 13-kb construct containing the eight exons and associated introns, 5 kb of the 5' region, and 3 kb downstream from the structural gene produced normal levels of enzyme activity and rescued the poor viability of flies lacking the enzyme. All the regulatory elements essential for normal enzyme expression were located in a fragment that included the exons and introns and 1-kb upstream noncoding sequence. Deletions of the 1.6-kb second intron reduced activity to 25%. Transformants with fusion constructs between the sn-glycerol-3-phosphate dehydrogenase gene and the beta-galactosidase gene from E. coli revealed three elements that affected expression. A (CT)9 repeat element at the 5' end of the second intron increased expression in both larvae and adults, particularly at emergence. A second regulatory element, which includes a (CT)7 repeat, was located 5' to the TATA box and had similar effects on the gene's expression. A third, undefined, enhancer was located in the second intron, between 0.5 and 1.8 kb downstream of the translation initiation codon. This element increases enzyme activity to a similar extent in larvae and adults but has little effect when the enhancer at the 5' end of the intron is present.     

Maternal Nanos regulates zygotic gene expression in germline progenitors of Drosophila melanogaster

Maternal Nanos (Nos) protein is required for germline development in Drosophila embryos. Here we show that Nos regulates zygotic gene expression in the germline progenitors, or pole cells. In order to probe the gene expression in pole cells, we screened ten enhancer-trap lines which showed beta-gal expression in pole cells. All of these enhancer-trap markers were fully activated in pole cells after their migration to the embryonic gonads. In the pole cells lacking Nos, the expression of nine out of ten enhancer-trap markers was affected. Among nine markers, five (Type-A) were prematurely expressed in the pole cells during the course of their migration. The expression of other four markers (Type-B) initiated correctly after pole-cell migration, but their expression was significantly reduced. Thus, we conclude that the maternal Nos plays a dual role in zygotic gene regulation in pole cells: to define the stages of expression for Type-A markers, and to enhance expression for Type-B markers. Contrary to our results, "Heller and Steinmann-Zwicky (1998)" have recently reported that no premature expression of Type-A markers occurs in the pole cells of embryos derived from nos mutant females. This discrepancy is due to the difference in the nos mutant alleles used for these analyses. We used the much stronger allele, nosBN.     

Divergent and conserved features in the spatial expression of the Drosophila pseudoobscura esterase-5B gene and the esterase-6 gene of Drosophila melanogaster

The regulatory regions of homologous genes encoding esterase 6 (Est-6) of Drosophila melanogaster and esterase 5B (Est-5B) of Drosophila pseudoobscura show very little similarity. We have undertaken a comparative study of the pattern of expression directed by the Est-5B and Est-6 5'-flanking DNA to attempt to reveal conserved elements regulating tissue-specific expression in adults. Esterase regulatory sequences were linked to a lacZ reporter gene and transformed into D. melanogaster embryos. Est-5B, 5' upstream elements, give rise to a beta-galactosidase expression pattern that coincides with the wild-type expression of Est-5B in D. pseudoobscura. The expression patterns of the Est-5B/lacZ construct are different from those of a fusion gene containing the upstream region of Est-6. Common sites of expression for both kinds of constructs are the third segment of antenna, the maxillary palps, and salivary glands. In vitro deletion mutagenesis has shown that the two genes have a different organization of regulatory elements controlling expression in both the third segment of antenna and maxillary palps. The results suggest that the conservation of the expression pattern in genes that evolved from a common ancestor may not be accompanied by preservation of the corresponding cis-regulatory elements.     

Critical periods in the development of mutants in scute Drosophila melanogaster

Thermosensitive periods of the development of germs of separate pairs of macrochaetae were studied in two lines of mutation of scute Drosophila melanogaster (y/sc4B and cs7). The experiments have revealed the following three features which had not been observed earlier in similar investigations in other mutants of drosophila: 1) change of thermosensitivity in mutants sc7 in different periods of ontogenesis; 2) different sensitivity in mutants of both lines in different critical periods of the same pair of macrochaetae; 3) discrete sensitivity of separate macrochaetae in mutants of both lines at different stages of oogenesis when there are no germs of these organs yet. The change of penetrability of the mutation of scute for scuttellar macrochaetae in mutants y/sc4B caused by the 90-minute-long heating of larvae on the 4th day of their life (critical period of development) is inherited in 15 observed generations of the maternal line. One may consider these periods of the increased thermosensitivity to be tje periods of stepwise determination of germs of separate pairs of macrochaetae.     

Disruption of local retinoid-mediated gene expression accompanies abnormal development in the mammalian olfactory pathway

Glycine, a neutral amino acid has been studied for its ability to inhibit gastric secretion and to protect the gastric mucosa against chemically and stress-induced ulcers. Acid secretion studies were undertaken in pylorus-ligated rats with and without glycine treatment. Experimental gastric lesions were induced by hypothermic-restraint stress, indomethacin and necrotizing agents including 80% ethanol, 0.2 M sodium hydroxide and 0.6 M hydrochloric acid in rats. The level of nonprotein sulfhydryl compounds and gastric wall mucus were also measured in the glandular stomach of the rats following ethanol-induced gastric lesions. The results of this study demonstrate that glycine dose dependently reduced the gastric secretions in rats. Pretreatment with glycine significantly protected animals against stress-, indomethacin- and necrotizing agents induced gastric lesions. The antiulcer activity of glycine was associated with significant inhibition of ethanol-induced depletion of nonprotein sulfhydryls and gastric wall mucus. In conclusion, this study demonstrates that glycine possesses significant antiulcer and cytoprotective activity. However, further detailed studies are warranted to establish the mechanism(s) of action, and to determine its role in the prophylaxis and treatment of gastric ulcer disease.     

Spatio-temporal patterns of ensheathing cell differentiation in the rat olfactory system during development

An immunocytochemical approach with specific glial markers was used to investigate the temporal and spatial patterns of differentiation of ensheathing glia wrapping axon fascicles along the primary olfactory pathway of the rat during development. The two glial markers tested, the proteins S-100 and glial fibrillary acidic protein, are known to be expressed at different stages of maturation in glial cells. The S-100 protein was first weakly expressed in cells accompanying the olfactory axons at embryonic day 14 (E14), while a first faint glial fibrillary acidic protein staining was detected along the olfactory axons at E15 and along the vomeronasal nerves at E16. A strong S-100 immunoreactivity was already present from E16 onwards along the axon fascicles through their course in both the nasal mesenchyme and the subarachnoid space before entering the olfactory nerve layer of the olfactory bulb. A gradual increase in glial fibrillary acidic protein expression was observed along this part of the developing olfactory pathway from E16 up to E20, when an adult-like pattern of staining intensity was seen. By contrast, most of the ensheathing cells residing in the olfactory nerve layer exhibited some delay in their differentiation timing and also a noticeable delayed maturation. It was only from E20 onwards that a weak to moderate S-100 expression was detected in an increasing number of cells throughout this layer, and only few of them appeared weakly glial fibrillary acidic protein positive at postnatal days 1 and 5. The immunocytochemical data indicate that there is a proximodistal gradient of differentiation of ensheathing cells along the developing olfactory pathway. The prolonged immaturity of ensheathing cells in the olfactory nerve layer, which coincides with the formation of the first glomeruli, might facilitate the sorting out of olfactory axons leading to a radical reorganization of afferents before they end in specific glomeruli.     

DSP1 gene of Drosophila melanogaster encodes an HMG-domain protein that plays multiple roles in development

Historically, juvenile justice policy has oscillated between rehabilitative and punitive approaches to managing young offenders. Policy and practice in the 1970s and 1980s emphasized individual treatment for young offenders in nonsecure, community-based programs. An increase in violent youth crime during the past decade has renewed interest in punishing delinquent youths. Cyclic fluctuations in juvenile justice policy and their relationship to policy, practice, and youth crime are examined. Our analysis suggests that overall crime rates have remained relatively stable over the past three decades and are independent of prevailing juvenile justice policies. The findings support the need for targeted prevention efforts addressing the root causes of juvenile crime. Needed policy reforms, public education efforts, and practice approaches are outlined.     

Changes in the expression of alpha- and beta-myosin heavy chain gene during the development of renal hypertension in rats

The changes in the expression of cardiac alpha- and beta-myosin heavy chain (MHC) gene of the left ventricle were investigated in two-kidney, one-clip (2K1C) renal hypertensive rats. The results were as follows: (1) When blood pressure was increased, the left ventricle became hypertrophic, alpha-MHC gene expression was reduced and beta-MHC gene expression was increased in 2K1C renal hypertensive rats. (2) When the animal was treated with captopril, angiotensin converting enzyme inhibitor 4 W after operation and then 8 W with removal of the ischemic kidney, the blood pressure was decreased with attendant regression of left ventricular hypertrophy, while the increase in beta-MHC mRNA level was attenuated and the inhibition of alpha-MHC mRNA level was reduced. The above results suggest that the rise in arteral pressure is an important factor in the left ventricular hypertrophy and the MHC gene switch. Renin angiotension system may be involved in the cardiac hypertrophic and MHC gene switch during the development and maintenance of 2K1C renal hypertension.     

A photographic study of development in the living embryo of Drosophila melanogaster

The changes which can be seen occurring during the development of a living embryo of Drosophila melanogaster are described in detail, and represented photographically as a series of developmental stages. This provides an easy, but accurate technique for selecting eggs at precise developmental stages for experiments.     

The su(Hw) protein insulates expression of the Drosophila melanogaster white gene from chromosomal position-effects

Mutations in the suppressor of Hairy-wing [su(Hw)] locus reverse the phenotype of a number of tissue-specific mutations caused by insertion of a gypsy retrotransposon. The su(Hw) gene encodes a zinc finger protein which binds to a 430 bp region of gypsy shown to be both necessary and sufficient for its mutagenic effects. su(Hw) protein causes mutations by inactivation of enhancer elements only when a su(Hw) binding region is located between these regulatory sequences and a promoter. To understand the molecular basis of enhancer inactivation, we tested the effects of su(Hw) protein on expression of the mini-white gene. We find that su(Hw) protein stabilizes mini-white gene expression from chromosomal position-effects in euchromatic locations by inactivating negative and positive regulatory elements present in flanking DNA. Furthermore, the su(Hw) protein partially protects transposon insertions from the negative effects of heterochromatin. To explain our current results, we propose that su(Hw) protein alters the organization of chromatin by creating a new boundary in a pre-existing domain of higher order chromatin structure. This separates enhancers and silencers distal to the su(Hw) binding region into an independent unit of gene activity, thereby causing their inactivation.     

A study of penetrance of Drosophila melanogaster forked gene

Group I intron endonuclease I-CreI is encoded by an open reading frame contained within a self-splicing intron in the Chlamydomonas reinhardtii chloroplast 23S rRNA gene. I-CreI initiates the lateral transfer or homing of this intron by specifically recognizing and cleaving a pseudopalindromic 19-24 bp homing site in chloroplast 23S rRNA genes that lack the intron. The gene encoding this enzyme has been subcloned, and the protein product has been purified and crystallized. The crystals belong to space group P321, with unit cell dimensions a = b = 78.2 A, c = 67.4 A. The crystal unit cell is consistent with an asymmetric unit consisting of the enzyme monomer. The specific volume of this unit cell is 3.3 A3/Da. The crystals diffract to at least 3.0 A resolution after flash-cooling, when using a rotating anode x-ray source and an RAXIS image plate detector.     

Gene-enzyme alcohol dehydrogenase system and adaptability in Drosophila melanogaster

As follows from the experiments on the genotypic related populations of Drosophila melanogaster with different frequency of ADH allozymes selection for postponement of ageing and resistance to hypothermia lead to saturation of population with S-allozyme ADH, and genotypic adaptation to ethanol-to increasing of frequency of F-allozyme. It is supposed that genotypic adaptation is realized by selection of specimens with the most favourable alleles of genes. At the same time ontogenetic adaptation is accompanied by biochemical modification of existing allozymes.     

Cell-specific differential expression of Na(+)-channel beta 1-subunit mRNA in the olfactory system during postnatal development and after denervation

Activity-dependent mechanisms have been implicated in olfactory system development but, although such activity requires ion channels, few reports have described their expression in the olfactory system. We investigated the developmental and denervation-induced regulation of the Na(+)-channel beta 1 subunit (Na beta 1) in rat olfactory bulb (OB) and piriform cortex (PC). In situ hybridization shows that Na beta 1 mRNA expression is upregulated developmentally, but with different time courses in mitral, tufted, and pyramidal cells. In mitral cells, label was detected at postnatal day 4 (P4) and gradually increased to P14. Tufted cells were devoid of Na beta 1 mRNA before P14, when most cells expressed adult levels. In pyramidal cells of PC, Na beta 1 expression was not detectable clearly until P14, with maximal expression at P28. To examine the regulation of Na beta 1 mRNA, we surgically deafferented the OB at P30 and compared the effects on Na beta 1 with those for Na(+)-channel alpha-subunit (Na alpha) mRNAs. Within 5 d of surgery, the Na beta 1 and Na alpha II signals within tufted cells disappeared almost completely. Na beta 1 and Na alpha II expression was decreased in mitral cells to low-to-moderate levels. In pyramidal cells, Na beta 1 mRNA expression was decreased moderately without significant changes in Na alpha II mRNA. Deafferentation had no detectable effects on Na alpha I or III mRNAs in either OB or PC. These data indicate that Na beta 1 mRNA is expressed differentially in subpopulations of cells in the olfactory system during development and after deafferentation and suggest that the expression of Na beta 1 is regulated independently of Na alpha mRNAs via cell-specific and pathway-specific mechanisms.     

The dynamics of neurogenic signalling underlying bristle development in Drosophila melanogaster

We have examined expression of the neurogenic gene, Delta (Dl), and the regulatory relationships between the Delta-Notch signalling pathway and the proneural gene, achaete, during microchaeta development in Drosophila. Delta is expressed in all microchaeta proneural cells and microchaeta sensory organ precursors (SOPs) and is expressed dynamically in SOP progeny. We find that Delta expression in microchaeta proneural cells is detected prior to the onset of achaete expression and arises normally in the absence of achaete/scute function, indicating that initial Delta expression in the notum is not dependent on proneural gene function. Activation of the Delta-Notch pathway results in loss of Delta protein accumulation, suggesting that Delta expression is regulated, in part, by Delta-Notch signalling activity. We find that Delta signalling is required for correct delineation of early proneural gene expression in developing nota. Within microchaeta proneural stripes, we demonstrate that Delta-Notch signalling prohibits adoption of the SOP fate by repressing expression of proneural genes.     

Effect of hexachlorocyclohexane on hsp26 expression in transgenic Drosophila melanogaster

The type and amount of lipophilic antioxidants in plasma of hyperlipidemic patients are of great importance since they play a central role in preventing deleterious oxidation of blood lipids and proteins. Isolation and quantitation of lipophilic antioxidants from hyperlipidemic plasma samples meet great obstacles because of increased levels of various intermediary lipid products. This study was designed to develop a rapid and efficient extraction and separation procedure for simultaneous analysis of ubiquinone-9 and -10 as well as alpha-, delta-, and gamma-tocopherol isomers. The levels of ubiquinone-10, alpha- and gamma-tocopherol were analyzed in human plasma samples using high-performance liquid chromatography. Lipid extraction was performed by petroleum ether/methanol/water. After phase separation, ubiquinone was reduced to ubiquinol by sodium borohydride and the lipids were separated on a C18 column. A binary gradient with solvents containing lithium perchlorate was used, and an electrochemical detector was employed for quantitation. This procedure was also efficient for the analysis of antioxidant lipids in samples containing a large number of accumulated and interfering lipid intermediates. Thus, the procedure described here is useful for efficient and rapid quantitation of ubiquinones and tocopherols in human plasma samples, especially those originating from hyperlipidemic patients.