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Trudi Schüpbach

Publications and source records attributed to Trudi Schüpbach.

17 recordsLinked to original sources
PubMed2006

Differential effects of Cbl isoforms on Egfr signaling in Drosophila.

The Cbl family of proteins downregulate epidermal growth factor receptor (Egfr) signaling via receptor internalization and destruction. These proteins contain two functional domains, a RING finger domain with E3 ligase activity, and a proline rich domain mediating the formation of protein complexes. The Drosophila cbl gene encodes two isoforms, D-CblS and D-CblL. While both contain a RING finger domain, the proline rich domain is absent from D-CblS. We demonstrate that expression of either isoform is sufficient to rescue both the lethality of a D-cbl null mutant and the adult phenotypes characteristic of Egfr hyperactivation, suggesting that both isoforms downregulate Egfr signaling. Interestingly, targeted overexpression of D-CblL, but not D-CblS, results in phenotypes characteristic of reduced Egfr signaling and suppresses the effect of constitutive Egfr activation. The level of D-CblL was significantly correlated with the phenotypic severity of reduced Egfr signaling, suggesting that D-CblL controls the efficiency of downregulation of Egfr signaling. Furthermore, reduced dynamin function suppresses the effects of D-CblL overexpression in follicle cells, suggesting that D-CblL promotes internalization of activated receptors. D-CblL is detected in a punctate cytoplasmic pattern, whereas D-CblS is mainly localized at the follicle cell cortex. Therefore, D-CblS and D-CblL may downregulate Egfr through distinct mechanisms.

Alternative Splicing↗
PubMed2006

The role of brinker in eggshell patterning.

Drosophila oogenesis provides a useful system to study signal transduction pathways and their interactions. Through clonal analysis, we found that brinker (brk), a repressor of Dpp signaling, plays an important role in the Drosophila ovary, where its function is essential for dorsal appendage formation. In the absence of brk, operculum fates are specified at the expense of dorsal appendage fates. Brk is expressed by most of the oocyte associated follicle cells, starting from stage 8 of oogenesis. Transforming Growth Factor beta (TGFbeta) signaling represses brk expression in both the early stage egg chambers and in the anterior follicle cells. In brk mutant follicle cell clones at the dorsal anterior region, Broad Complex (BR-C) expression is down-regulated in a larger domain than in wild type. We show that BR-C is required for dorsal appendage development. In large anterior BR-C mutant clones, dorsal appendages are absent, and instead, the eggshell has an enlarged operculum like region at the anterior. In addition, we show that the Epidermal Growth Factor (EGF) receptor signaling represses the TGFbeta signaling in oogenesis by up-regulating brk expression. From our results and previously published data, it appears that anterior follicle cells integrate the levels of EGF receptor activation and TGFbeta receptor activation. Operculum fate results when the sum of the level of activation of both pathways reaches a threshold level, and reduction of activity of one pathway can be compensated to some extent by increase in the other pathway.

Animals↗
PubMed2006

Deadlock, a novel protein of Drosophila, is required for germline maintenance, fusome morphogenesis and axial patterning in oogenesis and associates with centrosomes in the early embryo.

The deadlock gene is required for a number of key developmental events in Drosophila oogenesis. Females homozygous for mutations in the deadlock gene lay few eggs and those exhibit severe patterning defects along both the anterior-posterior and dorsal-ventral axis. In this study, we analyzed eggs and ovaries from deadlock mutants and determined that deadlock is required for germline maintenance, stability of mitotic spindles, localization of patterning determinants, oocyte growth and fusome biogenesis in males and females. Deadlock encodes a novel protein which colocalizes with the oocyte nucleus at midstages of oogenesis and with the centrosomes of early embryos. Our genetic and immunohistological experiments point to a role for Deadlock in microtubule function during oogenesis.

Animals↗
PubMed2006

spn-F encodes a novel protein that affects oocyte patterning and bristle morphology in Drosophila.

The anteroposterior and dorsoventral axes of the Drosophila embryo are established during oogenesis through the activities of Gurken (Grk), a Tgfalpha-like protein, and the Epidermal growth factor receptor (Egfr). spn-F mutant females produce ventralized eggs similar to the phenotype produced by mutations in the grk-Egfr pathway. We found that the ventralization of the eggshell in spn-F mutants is due to defects in the localization and translation of grk mRNA during mid-oogenesis. Analysis of the microtubule network revealed defects in the organization of the microtubules around the oocyte nucleus. In addition, spn-F mutants have defective bristles. We cloned spn-F and found that it encodes a novel coiled-coil protein that localizes to the minus end of microtubules in the oocyte, and this localization requires the microtubule network and a Dynein heavy chain gene. We also show that Spn-F interacts directly with the Dynein light chain Ddlc-1. Our results show that we have identified a novel protein that affects oocyte axis determination and the organization of microtubules during Drosophila oogenesis.

Animals↗
PubMed2006

Quantitative analysis of the GAL4/UAS system in Drosophila oogenesis.

The GAL4/UAS system is extensively used for targeted gene expression in Drosophila, but the strength of the GAL4 drivers and their effects on target genes are rarely quantified. Quantitative information about the strength of the perturbations introduced by the GAL4/UAS system would further expand the usefulness of the GAL4/UAS system in studying gene functions and developmental processes. We have developed an assay to determine the relative level of expression for target genes tagged with green fluorescent protein (GFP). Our assay enables the relative quantitation of fluorescent proteins within specific cell types and developmental time windows in living eggs/embryos, and permits the analysis of samples from a broad expression range. We illustrate the assay using a panel of four GAL4 drivers and three UAS responder lines in Drosophila oogenesis, discuss the issues associated with the interpretation of the quantitative data, and correlate our results with the analysis of the GAL4/UAS system at the transcript level. The imaging-based strategy described here can be used to quantify other GAL4 drivers in Drosophila and other organisms.

Animals↗
PubMed2006

Quantifying the Gurken morphogen gradient in Drosophila oogenesis.

Quantitative information about the distribution of morphogens is crucial for understanding their effects on cell-fate determination, yet it is difficult to obtain through direct measurements. We have developed a parameter estimation approach for quantifying the spatial distribution of Gurken, a TGFalpha-like EGFR ligand that acts as a morphogen in Drosophila oogenesis. Modeling of Gurken/EGFR system shows that the shape of the Gurken gradient is controlled by a single dimensionless parameter, the Thiele modulus, which reflects the relative importance of ligand diffusion and degradation. By combining the model with genetic alterations of EGFR levels, we have estimated the value of the Thiele modulus in the wild-type egg chamber. This provides a direct characterization of the shape of the Gurken gradient and demonstrates how parameter estimation techniques can be used to quantify morphogen gradients in development.

Animals↗
PubMed2006

Quantitative models of developmental pattern formation.

Pattern formation in developing organisms can be regulated at a variety of levels, from gene sequence to anatomy. At this level of complexity, mechanistic models of development become essential for integrating data, guiding future experiments, and predicting the effects of genetic and physical perturbations. However, the formulation and analysis of quantitative models of development are limited by high levels of uncertainty in experimental measurements, a large number of both known and unknown system components, and the multiscale nature of development. At the same time, an expanding arsenal of experimental tools can constrain models and directly test their predictions, making the modeling efforts not only necessary, but feasible. Using a number of problems in fruit fly development, we discuss how models can be used to test the feasibility of proposed patterning mechanisms and characterize their systems-level properties.

Animals↗
PubMed2005

Drosophila female meiosis and embryonic syncytial mitosis use specialized Cks and CDC20 proteins for cyclin destruction.

Female meiosis and the rapid mitotic cycle of early embryos are two non-canonical cell cycles that occur sequentially in the same cell, the egg, and utilize the same pool of cell cycle proteins. Using a genetic approach to identify genes that are specifically required for these cell cycles in Drosophila, we found that a Drosophila Cks gene, Cks30A is required for spindle assembly and anaphase progression in both female meiosis and in the syncytial embryo. Cks30A interacts with Cdk1 to target cyclin A for destruction in the female germline, possibly through the activation of a novel germline specific CDC20 protein, Cortex. These results indicate that anaphase progression in female meiosis and the early embryo are under unique control in Drosophila.

Animals↗
PubMed2005

Drosophila Cks30A interacts with Cdk1 to target Cyclin A for destruction in the female germline.

Cks is a small highly conserved protein that plays an important role in cell cycle control in different eukaryotes. Cks proteins have been implicated in entry into and exit from mitosis, by promoting Cyclin-dependent kinase (Cdk) activity on mitotic substrates. In yeast, Cks can promote exit from mitosis by transcriptional regulation of cell cycle regulators. Cks proteins have also been found to promote S-phase via an interaction with the SCF(Skp2) Ubiquitination complex. We have characterized the Drosophila Cks gene, Cks30A and we find that it is required for progression through female meiosis and the mitotic divisions of the early embryo through an interaction with Cdk1. Cks30A mutants are compromised for Cyclin A destruction, resulting in an arrest or delay at the metaphase/anaphase transition, both in female meiosis and in the early syncytial embryo. Cks30A appears to regulate Cyclin A levels through the activity of a female germline-specific anaphase-promoting complex, CDC20-Cortex. We also find that a second closely related Cks gene, Cks85A, plays a distinct, non-overlapping role in Drosophila, and the two genes cannot functionally replace each other.

Amino Acid Sequence↗
PubMed2004

Hrb27C, Sqd and Otu cooperatively regulate gurken RNA localization and mediate nurse cell chromosome dispersion in Drosophila oogenesis.

Heterogeneous nuclear ribonucleoproteins, hnRNPs, are RNA-binding proteins that play crucial roles in controlling gene expression. In Drosophila oogenesis, the hnRNP Squid (Sqd) functions in the localization and translational regulation of gurken (grk) mRNA. We show that Sqd interacts with Hrb27C, an hnRNP previously implicated in splicing. Like sqd, hrb27C mutants lay eggs with dorsoventral defects and Hrb27C can directly bind to grk RNA. Our data demonstrate a novel role for Hrb27C in promoting grk localization. We also observe a direct physical interaction between Hrb27C and Ovarian tumor (Otu), a cytoplasmic protein implicated in RNA localization. We find that some otu alleles produce dorsalized eggs and it appears that Otu cooperates with Hrb27C and Sqd in the oocyte to mediate proper grk localization. All three mutants share another phenotype, persistent polytene nurse cell chromosomes. Our analyses support dual cooperative roles for Sqd, Hrb27C and Otu during Drosophila oogenesis.

Animals↗
PubMed2003

The Drosophila spn-D gene encodes a RAD51C-like protein that is required exclusively during meiosis.

In Drosophila, mutations in double-strand DNA break (DSB) repair enzymes, such as spn-B, activate a meiotic checkpoint leading to dorsal-ventral patterning defects in the egg and an abnormal appearance of the oocyte nucleus. Mutations in spn-D cause an array of ovarian phenotypes similar to spn-B. We have cloned the spn-D locus and found that it encodes a protein of 271 amino acids that shows significant homology to the human RAD51C protein. In mammals the spn-B and spn-D homologs, XRCC3 and RAD51C, play a role in genomic stability in somatic cells. To test for a similar role for spn-B and spn-D in double-strand DNA repair in mitotic cells, we analyzed the sensitivity of single and double mutants to DSBs induced by exposure to X rays and MMS. We found that neither singly mutant nor doubly mutant animals were significantly sensitized to MMS or X rays. These results suggest that spn-B and spn-D act in meiotic recombination but not in repair of DSBs in somatic cells. As there is no apparent ortholog of the meiosis-specific DMC1 gene in the Drosophila genome, and given their meiosis-specific requirement, we suggest that spn-B and spn-D may have a function comparable to DMC1.

Animals↗
PubMed2003

Cct1, a phosphatidylcholine biosynthesis enzyme, is required for Drosophila oogenesis and ovarian morphogenesis.

Patterning of the Drosophila egg requires cooperation between the germline cells and surrounding somatic follicle cells. In order to identify genes involved in follicle cell patterning, we analyzed enhancer trap lines expressed in specific subsets of follicle cells. Through this analysis, we have identified tandem Drosophila genes homologous to CTP: phosphocholine cytidylyltransferase (CCT), the second of three enzymes in the CDP-choline pathway, which is used to synthesize phosphatidylcholine. Drosophila Cct1 is expressed at high levels in three specific subsets of follicle cells, and this expression is regulated, at least in part, by the TGF-beta and Egfr signaling pathways. Mutations in Cct1 result in a number of defects, including a loss of germline stem cell maintenance, mispositioning of the oocyte, and a shortened operculum, suggesting that Cct1 plays multiple roles during oogenesis. In addition, Cct1 mutants display a novel branched ovariole phenotype, demonstrating a requirement for this gene during ovarian morphogenesis. These data provide the first evidence for a specific role for CCT, and thus for phosphatidylcholine, in patterning during development.

Amino Acid Sequence↗
PubMed2003

Encore facilitates SCF-Ubiquitin-proteasome-dependent proteolysis during Drosophila oogenesis.

Exit from the cell cycle requires the downregulation of Cyclin/Cdk activity. In the ovary of Drosophila, Encore activity is necessary in the germline to exit the division program after four mitotic divisions. We find that in encore mutant germaria, Cyclin A persists longer than in wild type. In addition, Cyclin E expression is not downregulated after the fourth mitosis and accumulates in a polyubiquitinated form. Mutations in genes coding for components of the SCF pathway such as cul1, UbcD2 and effete enhance the extra division phenotype of encore. We show that Encore physically interacts with the proteasome, Cul1 and Cyclin E. The association of Cul1, phosphorylated Cyclin E and the proteasome 19S-RP subunit S1 with the fusome is affected in encore mutant germaria. We propose that in encore mutant germaria the proteolysis machinery is less efficient and, in addition, reduced association of Cul1 and S1 with the fusome may compromise Cyclin E destruction and consequently promote an extra round of mitosis.

Animals↗
PubMed2002

Activation of a meiotic checkpoint during Drosophila oogenesis regulates the translation of Gurken through Chk2/Mnk.

BACKGROUND: During Drosophila oogenesis, unrepaired double-strand DNA breaks activate a mei-41-dependent meiotic checkpoint, which couples the progression through meiosis to specific developmental processes. This checkpoint affects the accumulation of Gurken protein, a transforming growth factor alpha-like signaling molecule, as well as the morphology of the oocyte nucleus. However, the components of this checkpoint in flies have not been completely elucidated. RESULTS: We show that a mutation in the Drosophila Chk2 homolog (DmChk2/Mnk) suppresses the defects in the translation of gurken mRNA and also the defects in oocyte nuclear morphology. We also found that DmChk2 is phosphorylated in a mei-41-dependent pathway. Analysis of the meiotic cell cycle progression shows that the Drosophila Chk2 homolog is not required during early meiotic prophase, as has been observed for Chk2 in C. elegans. We demonstrate that the activation of the meiotic checkpoint affects Dwee1 localization and is associated with DmChk2-dependent posttranslational modification of Dwee1. We suggest that Dwee1 has a role in the meiotic checkpoint that regulates the meiotic cell cycle, but not the translation of gurken mRNA. In addition, we found that p53 and mus304, the Drosophila ATR-IP homolog, are not required for the patterning defects caused by the meiotic DNA repair mutations. CONCLUSIONS: DmChk2 is a transducer of the meiotic checkpoint in flies that is activated by unrepaired double-strand DNA breaks. Activation of DmChk2 in this specific checkpoint affects a cell cycle regulator as well as mRNA translation.

Animals↗
PubMed2002

Half pint regulates alternative splice site selection in Drosophila.

Alternative splicing is used by metazoans to increase protein diversity and to alter gene expression during development. However, few factors that control splice site choice in vivo have been identified. Here we describe a factor, Half pint (Hfp), that regulates RNA splicing in Drosophila. Females harboring hypomorphic mutations in hfp lay short eggs and show defects in germline mitosis, nuclear morphology, and RNA localization during oogenesis. We find that hfp encodes the Drosophila ortholog of human PUF60 and functions in both constitutive and alternative splicing in vivo. In particular, hfp mutants display striking defects in the developmentally regulated splicing of ovarian tumor (otu). Furthermore, transgenic expression of the missing otu splice form can rescue the ovarian phenotypes of hfp.

Alternative Splicing↗