Biomedical subjects

M Bownes

Publications and source records attributed to M Bownes.

At least 19 recordsLinked to original sources

PubMed2006

E75A and E75B have opposite effects on the apoptosis/development choice of the Drosophila egg chamber.

The number of Drosophila egg chambers is controlled by the nutritional status of the female. There is a developmental checkpoint at stage 8, which is controlled by BR-C in the follicle cells along with ecdysteroid. During this period, developmental decision is made in each egg chamber to determine if it will develop or die. During nutritional shortage, inducing apoptosis in the nurse cells of stages 8 and 9 egg chambers reduces the number of egg chambers. We show that ecdysone response genes E75A and E75B are involved in inducing or suppressing apoptosis. It is thus possible that the E75 isoforms A and B are involved in the decision to develop or die in oogenesis. We have established part of the pathway by which ecdysone response genes control apoptosis of the nurse cells and hence select between degeneration or development of individual egg chambers at stages 8 and 9.

Animals↗

PubMed2005

Effects of dopamine on juvenile hormone metabolism and fitness in Drosophila virilis.

The effects of dopamine (DA) on juvenile hormone (JH) metabolism and fitness (estimated as fecundity and viability levels under heat stress (38 degrees C)) in Drosophila virilis have been studied. An increase of DA level obtained by feeding with DA reduced fitness of wild-type (wt) flies under stress, and decreased JH degradation in young wt females while increasing it in sexually mature wt females. A decrease in DA levels resulted from 3-iodo-tyrosine treatment and caused a decrease in JH degradation in sexually mature wt and heat sensitive (hs) mutant females (DA level in hs females is twice as high in wt females). A dramatic decrease in viability under stress and fecundity under normal conditions in wt, but not hs, females was observed. 3-iodo-tyrosine treatment also reduced the number of oocytes at stages 8-14, delayed oocyte transition to stage 10 and resulted in the accumulation of mature eggs in wt females. It delayed maturation of wt, but not hs, males as well, but did not affect their fertility. This advances our understanding of the regulation of JH metabolism by DA in Drosophila and suggests a crucial role for the basal DA level in fitness.

Age Factors↗

PubMed2005

A microarray analysis of genes involved in relating egg production to nutritional intake in Drosophila melanogaster.

Egg chambers of Drosophila are reabsorbed under conditions of nutritional shortage by inducing apoptosis at stages 8 and 9, midway through oogenesis. Nutritional shortage leads to an increase in ecdysone concentration in flies. Apoptosis at stage 8/9 is also induced by 20-hydroxyecdysone injection into the females maintained with adequate nutrition. The expression pattern in the ovary of some ecdysone response genes, E75A, BR-C, is different according to the nutritional environment and the overexpression of these genes induces apoptosis. Apoptosis is suppressed by Juvenile hormone analog treatment of females under nutritional shortage. We predict nutritional and stress response genes control hormone levels and the increase in ecdysone concentration in the flies following starvation induces the ovarian apoptosis. We therefore used a microarray approach to identify the genes involved in receiving the nutritional signal from the environment and translating it in the ovary, thus initiating and executing apoptosis.

Animal Feed↗

PubMed2005

Nutritional status affects 20-hydroxyecdysone concentration and progression of oogenesis in Drosophila melanogaster.

Drosophila egg production depends upon the nutritional available to females. When food is in short supply, oogenesis is arrested and apoptosis of the nurse cells is induced at mid-oogenesis via a mechanism that is probably controlled by ecdysteroid hormone. We have shown that expression of some ecdysone-response genes is correlated with apoptosis of egg chambers. Moreover, ecdysteroid injection and application of juvenile hormone induces and suppresses the apoptosis, respectively. In this study, we investigated which tissues show increases in the concentration of ecdysteroids under nutritional shortage to begin to link together nutrient intake, hormone regulation and the choice between egg development or apoptosis made within egg chambers. We measured ecdysteroid levels in the whole body, ovaries and haemolymph samples by RIA and found that the concentration of ecdysteroid increased in all samples. This contributes to the idea that nutritional shortage leads to a rapid high ecdysteroid concentration within the fly and that the high concentration induces apoptosis. Low concentrations of ecdysteroid are essential for normal oogenesis. We suggest there is threshold concentration in the egg chambers and that apoptosis at mid-oogenesis is induced when the ecdysteroid levels exceed the threshold. Starvation causes the ovary to retain the ecdysteroid it produces, thus enabling individual egg chambers to undergo apoptosis and thus control the number of eggs produced in relation to food intake.

Animal Nutritional Physiological Phenomena↗

PubMed2004

The role of juvenile hormone in the control of reproductive function in Drosophila virilis under nutritional stress.

To investigate the role of juvenile hormone (JH) in the control of Drosophila reproduction under stress, JH degradation and reproduction were studied under nutritional stress and JH treatment in Drosophila virilis females of wild type (wt) and a heat stress (hs) mutant: this mutant does not respond to heat stress by alterations in JH metabolism and has decreased JH level and fertility under normal conditions. One day of starvation results in a decrease of JH degradation, a delay in oocyte maturation, degradation of early vitellogenic egg chambers, accumulation of mature oocytes and a 24 h oviposition arrest in both wt and hs females. A fertility decrease was observed in both wt and hs females 24 h following the end of starvation. JH treatment leads to a decrease of JH degradation and an arrest of oviposition for 24 h in fed females. JH treatment prior to starvation seems to protect some oocytes from resorption: in JH-treated wt females, fertility increases rapidly following the end of starvation. The dynamics of JH degradation and fertility are similar following starvation and JH treatment. The role of JH in the accumulation of mature oocytes and the delay of oviposition under stress are discussed.

Animals↗

PubMed2003

Stress response in a juvenile hormone-deficient Drosophila melanogaster mutant apterous56f.

The apterous56f (ap56f) mutation leads to increases in juvenile hormone (JH) degradation levels and JH-esterase makes a greater contribution to the increase than JH-epoxide hydrolase. Dopamine levels in ap56f females, but not males, are higher than in wild-type. JH treatment of ap56f and wild-type females decreases their dopamine levels. ap56f females, but not males, produce less progeny. Survival under heat stress is dramatically decreased in ap56f females, but not males. ap56f flies show a stress reaction, as judged by changes in tyrosine decarboxylase and JH-hydrolysing activities, dopamine levels and fertility, but its intensity in the mutant females, but not males, differs significantly from wild-type. Thus, the ap56f mutation causes dramatic changes in female, but not male, metabolism and fitness.

Animals↗

PubMed2003

Heat stress affects oogenesis differently in wild-type Drosophila virilis and a mutant with altered juvenile hormone and 20-hydroxyecdysone levels.

The link between reproduction and environmental signals is poorly understood at the physiological, genetic and molecular levels. We describe a mutant strain of Drosophila virilis that has altered responses to heat stress. Heat stress in wild-type females results in oocyte maturation delays, degradation of early vitellogenic egg chambers, inhibition of yolk protein gene expression in follicle cells and accumulation of mature oocytes. The mutant females have increased levels of ecdysteroids and decreased juvenile hormone degradation, and show all of the heat-stress-induced reproductive effects observed in wild-type flies, without exposure to heat stress. During oogenesis in mutant females following heat stress there is an increase in early vitellogenic oocyte degradation and some degradation of late egg chambers. 20-Hydroxyecdysone levels, but not juvenile hormone degradation, change following heat stress in mutant females.

Animals↗

PubMed2002

Drosophila yolk protein produced in E. coli is accumulated by mosquito ovaries.

Despite similar functions, the yolk proteins of the higher dipteran flies and the vitellogenins found in other insects are unrelated at the sequence level and have evolved from different genes. Both are selectively endocytosed into the ovary via receptors belonging to the LDLR receptor subfamily. We cloned the Drosophila yp1 gene into an E. coli expression vector and showed that the yolk protein produced by E. coli is taken up into ovaries of both Drosophila melanogaster and the malaria mosquito Anopheles gambiae, which normally uses vitellogenin.

Animals↗

PubMed2001

The Drosophila RGS protein Loco is required for dorsal/ventral axis formation of the egg and embryo, and nurse cell dumping.

The loco gene encodes members of a family of RGS proteins responsible for the negative regulation of G-protein signalling. At least two transcripts of loco are expressed in oogenesis, loco-c2 is observed in the anterior-dorsal follicle cells and is downstream of the epidermal growth factor receptor signalling pathway, initiated in the oocyte. loco-c3 is a new transcript of loco, which is expressed in the nurse cells from stage 6 onwards. Analysis of newly generated mutants and antisense technology enabled us to establish that disrupting loco in follicle cells results in ventralized eggs, while disrupting loco in nurse cells results in short eggs, due to defective dumping of the nurse cell cytoplasm into the oocyte.

Amino Acid Sequence↗

PubMed2000

The Mirror transcription factor links signalling pathways in Drosophila oogenesis.

Many genetic cascades are conserved in evolution, yet they trigger different responses and hence determine different cell fates at specific times and positions in development. At stage 10 of oogenesis, mirror is expressed in anterior-dorsal follicle cells, and we show that this is dependent upon the Gurken signal from the oocyte. The fringe gene is expressed in a complementary pattern in posterior-ventral follicle cells at the same stage. Ectopic expression of mirror represses fringe expression, thus linking the epidermal growth factor receptor (EGFR) signalling pathway to the Fringe signalling pathway via Mirror. The EGFR pathway also triggers the cascade that leads to dorsal-ventral axis determination in the embryo. We used twist as an embryonic marker for ventral cells. Ectopic expression of mirror in the follicle cells during oogenesis ultimately represses twist expression in the embryo, and leads to similar phenotypes to the ectopic expression of the activated form of EGFR. Thus, mirror also controls the Toll signalling pathway, leading to Dorsal nuclear transport. In summary, we show that the Mirror homeodomain protein provides a link that coordinates the Gurken/EGFR signalling pathway (initiated in the oocyte) with the Fringe/Notch/Delta pathway (in follicle cells). This coordination is required for epithelial morphogenesis, and for producing the signal in ventral follicle cells that determines the dorsal/ventral axis of the embryo.

Animals↗

PubMed2000

A putative sodium-dependent inorganic phosphate co-transporter from Drosophila melanogaster.

We have isolated and sequenced a cDNA encoding a predicted 524 amino acid protein from a Drosophila melanogaster ovarian library. Sequence comparisons suggest that this protein encodes a sodium-dependent inorganic phosphate co-transporter similar to a sequence isolated from a rat brain library. In situ hybridisation to messenger RNA in ovaries shows strong expression in germarium at stage 2 of oogenesis. Expression is then weak in follicle cells until stage 10, when high transcript levels are seen in the nurse cells and transferred to the oocyte. This presumably reflects functions in oogenesis and the production of stored mRNAs for use in embryogenesis.

Amino Acid Sequence↗

PubMed2000

Mouse staufen genes are expressed in germ cells during oogenesis and spermatogenesis.

The Drosophila melanogaster staufen gene encodes an RNA binding protein (Dm Stau) required for the localization and translational repression of mRNAs within the Drosophila oocyte. In mammals translational repression is important for normal spermatogenesis in males and storage of mRNAs in the oocytes of females. In the present study we identified two mouse cDNA expressed sequence tags (ESTs), encoding proteins with significant homology to Dm Stau and used these firstly to screen a mouse kidney cDNA library and secondly to determine whether staufen mRNAs are expressed in the ovaries and testes of mice and rats. Sequence analysis of the cDNAs revealed that they originated from two different genes. Using Northern blots of RNAs from kidneys, ovaries and testes, both cDNAs hybridized to mRNA species of approximately 3 kb in all three tissues. On sections of mouse ovaries, staufen mRNA was localized specifically to oocytes. On sections of mouse testes, staufen mRNA was expressed in spermatocytes found in seminiferous tubules at stages VI-XII of the spermatogenic cycle. In conclusion, we have shown that the mammalian homologues of Dm stau are expressed in germ cells in both male and female mice, consistent with a role for these RNA binding proteins in mammalian gametogenesis.

Amino Acid Sequence↗

PubMed2000

Expression of fringe is down regulated by Gurken/Epidermal Growth Factor Receptor signalling and is required for the morphogenesis of ovarian follicle cells.

Signalling by the Gurken/Epidermal Growth Factor Receptor (Grk/EGFR) pathway is involved in epithelial cell fate decision, morphogenesis and axis establishment in Drosophila oogenesis. In the search for genes downstream of the Grk/EGFR signal transduction pathway (STP), we isolated a number of genes that are components of other STPs. One of them is a known gene, called fringe (fng). Drosophila fng encodes a putative secreted protein that is required at other development stages for mediating interactions between dorsal and ventral cells via Notch signalling. Here we show that fng has a dynamic expression pattern in oogenesis and that its expression in specific groups of follicle cells along the anterior-posterior and dorsal-ventral axes is defined by the repression of fng by Grk. Interfering with fng expression using antisense RNA experiments resulted in a typical fng mutant phenotype in the wing, and malformed egg chambers and abnormal organisation of the follicle cells in the ovaries, revealing that fng is essential in oogenesis for the proper formation of the egg chamber and for epithelial morphogenesis. This has been confirmed by re-examination of fng mutants and analysis of fng mutant clones in oogenesis.

Animals↗

PubMed2000

The Dstpk61 locus of Drosophila produces multiple transcripts and protein isoforms, suggesting it is involved in multiple signalling pathways.

The Drosophila gene Dstpk61 encodes a serine threonine protein kinase homologous to human phosphoinositide-dependent protein kinase (PDK1), and also has homologues in S. cerevisiae, S. pombe, C. elegans, A. thaliana, mouse, and sheep. Where its function has been investigated, this kinase is thought to be involved in regulating cell growth and survival in response to extracellular signals such as insulin and growth factors. In Drosophila it produces multiple transcripts, some of which appear to be sex-specific. In addition to the five Dstpk61 cDNAs we have described previously we report the existence of a further 18 expressed sequence tag (EST) cDNAs, three of which we have fully sequenced. We conclude that Dstpk61 is a complex locus that utilises a combination of alternative promoters, alternative splice sites and alternative polyadenylation sites to produce a vast array of different transcripts. These cDNAs encode at least four different DSTPK61 protein isoforms with variant N-termini. In this paper, we discuss the possible functions of the distinct Dstpk61 transcripts and how they might be differentially regulated. We also discuss the roles that DSTPK61 protein isoforms might play in relation to the protein domains they contain and their potential targets in the cell. Finally, we report the putative structure of the human PDK1 gene based on computer comparisons of available mRNA and genomic sequences. The value of using sequence data from other species for experimental design in mammalian systems is discussed.

3' Untranslated Regions↗

PubMed1999

Control of oocyte maturation in sexually mature Drosophila females.

In many sexually mature insects egg production and oviposition are tightly coupled to copulation. Sex-Peptide is a 36-amino-acid peptide synthesized in the accessory glands of Drosophila melanogaster males and transferred to the female during copulation. Sex-Peptide stimulates vitellogenic oocyte progression through a putative control point at about stage 9 of oogenesis. Here we show that application of the juvenile hormone analogue methoprene mimics the Sex-Peptide-mediated stimulation of vitellogenic oocyte progression in sexually mature virgin females. Apoptosis is induced by 20-hydroxyecdysone in nurse cells of stage 9 egg chambers at physiological concentrations (10(-7) M). 20-Hydroxyecdysone thus acts as an antagonist of early vitellogenic oocyte development. Simultaneous application of juvenile hormone analogue, however, protects early vitellogenic oocytes from 20-hydroxyecdysone-induced resorption. These results suggest that the balance of these hormones in the hemolymph regulates whether oocytes will progress through the control point at stage 9 or undergo apoptosis. These data are further supported by a molecular analysis of the regulation of yolk protein synthesis and uptake into the ovary by the two hormones. We conclude that juvenile hormone is a downstream component in the Sex-Peptide response cascade and acts by stimulating vitellogenic oocyte progression and inhibiting apoptosis. Since juvenile hormone analogue does not elicit increased oviposition and reduced receptivity, Sex-Peptide must have an additional, separate effect on these two postmating responses.

Animals↗

PubMed1999

Misexpression of argos, an inhibitor of EGFR signaling in oogenesis, leads to the production of bicephalic, ventralized, and lateralized Drosophila melanogaster eggs.

Epidermal growth factor receptor (EGFR) signaling pathways are frequently involved in generating cell fate diversity in a number of organisms. During anterior-posterior and dorso-ventral polarity in the Drosophila egg chamber and eggshell, EGFR signaling leads to a number of determinative events in the follicle cell layer. A high level of Gurken signal leads to the expression of argos in dorsal midline cells. Lateral follicle cells, receiving a lower level of Gurken signal, can continue to express the Broad-Complex (BR-C) and differentiate into cells which produce chorionic appendages. Misexpression of argos in mid-oogenesis causes the midline cells to retain expression of BR-C, resulting in a single fused large appendage. Evidence that argos can directly repress Gurken-induced EGFR signaling is seen when premature expression of argos is induced earlier in oogenesis. It represses the Gurken signal at stage 5-6 of oogenesis which determines posterior follicle cells and occasionally leads to eggs with anteriors at both ends. We propose that the Gurken signal at stage 9 of oogenesis induces follicle cells to take on two fates, dorsal midline and lateral, each producing different parts of the eggshell and that argos is one of the key downstream genes required to select between these two fates.

Animals↗

PubMed1999

Sex-specific transcripts of the Dstpk61 serine/threonine kinase gene in Drosophila melanogaster.

We describe the characterization of several transcripts of the Drosophila serine/threonine protein kinase 61 (Dstpk61) gene. Dstpk61 produces at least four transcripts, including a 3.0-kb testis-specific transcript, a 4.5-kb female-specific carcass transcript, a 3.5-kb ovary-specific transcript, and a 4.7-kb non-sex-specific transcript. Two cDNAs, a 4.5-kb cDNA (cDNAB) and a 3.0-kb cDNA (cDNAA), likely to correspond to either the non-specific or the female-specific carcass and the testis-specific transcript, respectively, were fully sequenced and found to encode a novel OPA-repeat-containing serine/threonine-specific protein kinase. cDNAA and cDNAB both contain the entire ORF that encodes this predicted protein, but differ in the untranslated regions. The cDNAs contain translational control elements which are found in transcripts under male germline-specific translational control, and doublesex-like 13-nucleotide repeat elements, which are required for transformer/transformer-2-mediated splicing of the female doublesex transcript. The complex tissue and sex-specific transcripts, differing in the untranslated regions which are likely to be crucial in translational control, suggest that this kinase may have both general and sex-specific functions. The protein is homologous to human 3-phosphoinositide dependent protein kinase, which is involved in transduction of insulin signalling.

3-Phosphoinositide-Dependent Protein Kinases↗

PubMed1999

A yolk protein mutant leads to defects in the secretion machinery of Drosophila melanogaster.

The three yolk proteins of Drosophila melanogaster are synthesized in the fat body and ovarian follicle cells. A mutation in yolk protein 3, YP3S1, has been described in which the leader sequence is not cleaved from the protein. We describe here ultrastructural and molecular studies on the YP3S1 mutant and show that the mutant protein enters the secretory pathway and forms precipitates, often as electron dense material in excessive elaborations of the plasma membrane. Females homozygous for YP3S1 lay fewer eggs than wild type flies and these embryos are less viable. The abnormal ultrastructure of the yolk spheres observed suggests that whilst YP3 is not completely essential for viability, it is required for normal yolk sphere morphogenesis.

Animals↗