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1.
Sci Rep ; 9(1): 7464, 2019 05 16.
Article in English | MEDLINE | ID: mdl-31097722

ABSTRACT

Organisms use circulating diuretic hormones to control water balance (osmolarity), thereby avoiding dehydration and managing excretion of waste products. The hormones act through G-protein-coupled receptors to activate second messenger systems that in turn control the permeability of secretory epithelia to ions like chloride. In insects, the chloride channel mediating the effects of diuretic hormones was unknown. Surprisingly, we find a pentameric, cys-loop chloride channel, a type of channel normally associated with neurotransmission, mediating hormone-induced transepithelial chloride conductance. This discovery is important because: 1) it describes an unexpected role for pentameric receptors in the membrane permeability of secretory epithelial cells, and 2) it suggests that neurotransmitter-gated ion channels may have evolved from channels involved in secretion.


Subject(s)
Chloride Channels/metabolism , Chlorides/metabolism , Drosophila Proteins/metabolism , Insect Hormones/metabolism , Animals , Chloride Channels/chemistry , Chloride Channels/genetics , Drosophila melanogaster , Epithelium/metabolism , Ion Channel Gating , Ion Transport , Malpighian Tubules/metabolism , Osmoregulation , Protein Domains , Xenopus
2.
Curr Biol ; 26(19): 2572-2582, 2016 10 10.
Article in English | MEDLINE | ID: mdl-27593379

ABSTRACT

A relatively small number of signaling pathways drive a wide range of developmental decisions, but how this versatility in signaling outcome is generated is not clear. In the Drosophila follicular epithelium, localized epidermal growth factor receptor (EGFR) activation induces distinct cell fates depending on its location. Posterior follicle cells respond to EGFR activity by expressing the T-box transcription factors Midline and H15, while anterior cells respond by expressing the homeodomain transcription factor Mirror. We show that the choice between these alternative outputs of EGFR signaling is regulated by antiparallel gradients of JAK/STAT and BMP pathway activity and that mutual repression between Midline/H15 and Mirror generates a bistable switch that toggles between alternative EGFR signaling outcomes. JAK/STAT and BMP pathway input is integrated through their joint and opposing regulation of both sides of this switch. By converting this positional information into a binary decision between EGFR signaling outcomes, this regulatory network ultimately allows the same ligand-receptor pair to establish both the anterior-posterior (AP) and dorsal-ventral (DV) axes of the tissue.


Subject(s)
Drosophila Proteins/metabolism , Drosophila melanogaster/metabolism , ErbB Receptors/metabolism , Janus Kinases/metabolism , Receptors, Invertebrate Peptide/metabolism , STAT Transcription Factors/metabolism , Signal Transduction , Animals , Drosophila Proteins/genetics , Drosophila melanogaster/genetics , ErbB Receptors/genetics , Gene Expression Regulation , Janus Kinases/genetics , Receptors, Invertebrate Peptide/genetics , STAT Transcription Factors/genetics
3.
J Exp Biol ; 219(Pt 17): 2629-38, 2016 09 01.
Article in English | MEDLINE | ID: mdl-27358471

ABSTRACT

Pentameric ligand-gated ion channels (pLGICs) constitute a large protein superfamily in metazoa whose role as neurotransmitter receptors mediating rapid, ionotropic synaptic transmission has been extensively studied. Although the vast majority of pLGICs appear to be neurotransmitter receptors, the identification of pLGICs in non-neuronal tissues and homologous pLGIC-like proteins in prokaryotes points to biological functions, possibly ancestral, that are independent of neuronal signalling. Here, we report the molecular and physiological characterization of a highly divergent, orphan pLGIC subunit encoded by the pHCl-2 (CG11340) gene, in Drosophila melanogaster We show that pHCl-2 forms a channel that is insensitive to a wide array of neurotransmitters, but is instead gated by changes in extracellular pH. pHCl-2 is expressed in the Malpighian tubules, which are non-innervated renal-type secretory tissues. We demonstrate that pHCl-2 is localized to the apical membrane of the epithelial principal cells of the tubules and that loss of pHCl-2 reduces urine production during diuresis. Our data implicate pHCl-2 as an important source of chloride conductance required for proper urine production, highlighting a novel role for pLGICs in epithelial tissues regulating fluid secretion and osmotic homeostasis.


Subject(s)
Body Fluids/metabolism , Drosophila Proteins/metabolism , Drosophila melanogaster/metabolism , Ligand-Gated Ion Channels/metabolism , Malpighian Tubules/metabolism , Amino Acid Sequence , Animals , Chloride Channels/metabolism , Cyclic AMP/pharmacology , Diuresis/drug effects , Drosophila Proteins/chemistry , Drosophila Proteins/genetics , Drosophila melanogaster/drug effects , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Hydrogen-Ion Concentration/drug effects , Ligand-Gated Ion Channels/chemistry , Ligand-Gated Ion Channels/genetics , Malpighian Tubules/drug effects , Models, Biological , Mutation/genetics , Protein Subunits/chemistry , Protein Subunits/metabolism
4.
Cell Rep ; 4(4): 791-802, 2013 Aug 29.
Article in English | MEDLINE | ID: mdl-23972992

ABSTRACT

Spatially restricted epidermal growth factor receptor (EGFR) activity plays a central role in patterning the follicular epithelium of the Drosophila ovary. In midoogenesis, localized EGFR activation is achieved by the graded dorsal anterior localization of its ligand, Gurken. Graded EGFR activity determines multiple dorsal anterior fates along the dorsal-ventral axis but cannot explain the sharp posterior limit of this domain. Here, we show that posterior follicle cells express the T-box transcription factors Midline and H15, which render cells unable to adopt a dorsal anterior fate in response to EGFR activation. The posterior expression of Midline and H15 is itself induced in early oogenesis by posteriorly localized EGFR signaling, defining a feedback loop in which early induction of Mid and H15 confers a molecular memory that fundamentally alters the outcome of later EGFR signaling. Spatial regulation of the EGFR pathway thus occurs both through localization of the ligand and through localized regulation of the cellular response.


Subject(s)
Drosophila Proteins/metabolism , Drosophila/metabolism , ErbB Receptors/metabolism , Oogenesis , Receptors, Invertebrate Peptide/metabolism , Signal Transduction , Animals , Cell Lineage , Drosophila/physiology , Drosophila Proteins/genetics , Epithelium/metabolism , Epithelium/physiology , ErbB Receptors/genetics , Female , Mutation , Receptors, Invertebrate Peptide/genetics , T-Box Domain Proteins/genetics , T-Box Domain Proteins/metabolism , Transforming Growth Factor alpha/genetics , Transforming Growth Factor alpha/metabolism
5.
J Cell Biol ; 192(2): 335-48, 2011 Jan 24.
Article in English | MEDLINE | ID: mdl-21263031

ABSTRACT

During Drosophila melanogaster dorsal closure, lateral sheets of embryonic epidermis assemble an actomyosin cable at their leading edge and migrate dorsally over the amnioserosa, converging at the dorsal midline. We show that disappearance of the homophilic cell adhesion molecule Echinoid (Ed) from the amnioserosa just before dorsal closure eliminates homophilic interactions with the adjacent dorsal-most epidermal (DME) cells, which comprise the leading edge. The resulting planar polarized distribution of Ed in the DME cells is essential for the localized accumulation of actin regulators and for actomyosin cable formation at the leading edge and for the polarized localization of the scaffolding protein Bazooka/PAR-3. DME cells with uniform Ed fail to assemble a cable and protrude dorsally, suggesting that the cable restricts dorsal migration. The planar polarized distribution of Ed in the DME cells thus provides a spatial cue that polarizes the DME cell actin cytoskeleton, defining the epidermal leading edge and establishing its contractile properties.


Subject(s)
Cell Adhesion Molecules/metabolism , Drosophila Proteins/metabolism , Drosophila melanogaster/embryology , Drosophila melanogaster/metabolism , Epidermis/metabolism , Repressor Proteins/metabolism , Actins/metabolism , Actomyosin/metabolism , Amino Acid Motifs , Animals , Cell Adhesion , Cell Adhesion Molecules/chemistry , Cell Movement , Cell Polarity , Drosophila Proteins/chemistry , Epidermal Cells , Intracellular Signaling Peptides and Proteins/metabolism , Protein Structure, Tertiary , Repressor Proteins/chemistry
6.
Dev Dyn ; 239(9): 2509-19, 2010 Sep.
Article in English | MEDLINE | ID: mdl-20730906

ABSTRACT

Morphogenesis of the Drosophila embryonic trachea involves a stereotyped pattern of epithelial tube branching and fusion. Here, we report unexpected phenotypes resulting from maternal and zygotic (M/Z) loss of the homophilic cell adhesion molecule Echinoid (Ed), as well as the subcellular localization of Ed in the trachea. ed(M/Z) embryos have convoluted trachea reminiscent of septate junction (SJ) and luminal matrix mutants. However, Ed does not localize to SJs, and ed(M/Z) embryos have intact SJs and show normal luminal accumulation of the matrix-modifying protein Vermiform. Surprisingly, tracheal length is not increased in ed(M/Z) mutants, but a previously undescribed combination of reduced intersegmental spacing and deep epidermal grooves produces a convoluted tracheal phenotype. In addition, ed(M/Z) mutants have unique fusion defects involving supernumerary fusion cells, ectopic fusion events and atypical branch breaks. Tracheal-specific expression of Ed rescues these fusion defects, indicating that Ed acts in trachea to control fusion cell fate.


Subject(s)
Cell Adhesion Molecules/metabolism , Cell Fusion , Drosophila Proteins/metabolism , Drosophila melanogaster , Morphogenesis/physiology , Repressor Proteins/metabolism , Amidohydrolases/genetics , Amidohydrolases/metabolism , Animals , Cell Adhesion Molecules/genetics , Drosophila Proteins/genetics , Drosophila melanogaster/anatomy & histology , Drosophila melanogaster/embryology , Embryo, Nonmammalian/anatomy & histology , Embryo, Nonmammalian/physiology , Phenotype , Repressor Proteins/genetics , Trachea/anatomy & histology , Trachea/embryology , Wnt1 Protein/metabolism
7.
Development ; 136(17): 2893-902, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19641015

ABSTRACT

The pattern of the Drosophila eggshell is determined by the establishment of a complex and stereotyped pattern of cell fates in the follicular epithelium of the ovary. Localized activation of the Epidermal growth factor receptor (Egfr) is essential for this patterning. Modulation of Egfr pathway activity in time and space determines distinct fates at their appropriate locations, but the details of how Egfr signaling is regulated and how the profile of Egfr activity corresponds to cell fate remain unclear. Here we analyze the effect of loss of various Egfr regulators and targets on follicle cell patterning, using a marker for follicle cell fate, and on the mature eggshell phenotype, using a novel eggshell marker. We show, contrary to current patterning models, that feedback regulation of Egfr activity by the autocrine ligand Spitz and the inhibitor Argos is not necessary for patterning. Given the cell-autonomous nature of the mutant phenotypes we observed, we propose instead that the pattern of cell fates is generated by spatial information derived directly from the germline ligand Gurken, without a requirement for subsequent patterning by diffusible Egfr regulators in the follicular epithelium.


Subject(s)
Autocrine Communication , Body Patterning/physiology , Drosophila Proteins/metabolism , Drosophila melanogaster , ErbB Receptors/metabolism , Oocytes , Signal Transduction/physiology , Animals , Animals, Genetically Modified , Biomarkers/metabolism , Cell Lineage , Drosophila Proteins/genetics , Drosophila melanogaster/anatomy & histology , Drosophila melanogaster/physiology , Epidermal Growth Factor/genetics , Epidermal Growth Factor/metabolism , ErbB Receptors/genetics , Eye Proteins/genetics , Eye Proteins/metabolism , Female , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Male , Membrane Proteins/genetics , Membrane Proteins/metabolism , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Oocytes/cytology , Oocytes/physiology , Ovary/cytology , Ovary/metabolism , Phenotype , Transcription Factors/genetics , Transcription Factors/metabolism
8.
Dev Biol ; 326(2): 327-34, 2009 Feb 15.
Article in English | MEDLINE | ID: mdl-19100729

ABSTRACT

Establishment of the Drosophila dorsal-ventral axis depends upon the correct localization of gurken mRNA and protein within the oocyte. gurken mRNA becomes localized to the presumptive dorsal anterior region of the oocyte, but is synthesized in the adjoining nurse cells. Normal gurken localization requires the heterogeneous nuclear ribonucleoprotein Squid, which binds to the gurken 3' untranslated region. However, whether Squid functions in the nurse cells or the oocyte is unknown. To address this question, we generated genetic mosaics in which half of the nurse cells attached to a given oocyte are unable to produce Squid. In these mosaics, gurken mRNA is localized normally but ectopically translated during the dorsal anterior localization process, even though the oocyte contains abundant Squid produced by the wild type nurse cells. These data indicate that translational repression of gurken mRNA requires Squid function in the nurse cells. We propose that Squid interacts with gurken mRNA in the nurse cell nuclei and, together with other factors, maintains gurken in a translationally silent state during its transport to the dorsal anterior region of the oocyte. This translational repression is not required for gurken mRNA localization, indicating that the information repressing translation is separable from that regulating localization.


Subject(s)
Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Drosophila melanogaster , Gene Expression Regulation, Developmental , Heterogeneous-Nuclear Ribonucleoproteins/metabolism , Oocytes , Protein Biosynthesis , RNA-Binding Proteins/metabolism , Transforming Growth Factor alpha/genetics , Animals , Drosophila melanogaster/anatomy & histology , Drosophila melanogaster/physiology , Germ Cells/cytology , Germ Cells/physiology , Heterogeneous-Nuclear Ribonucleoproteins/genetics , Mosaicism , Oocytes/cytology , Oocytes/physiology , Protein Isoforms/genetics , Protein Isoforms/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA-Binding Proteins/genetics , Transforming Growth Factor alpha/metabolism
9.
Mech Dev ; 123(6): 450-62, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16844358

ABSTRACT

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.


Subject(s)
Drosophila Proteins/chemistry , Drosophila Proteins/physiology , ErbB Receptors/physiology , Gene Expression Regulation, Developmental , Proto-Oncogene Proteins c-cbl/chemistry , Proto-Oncogene Proteins c-cbl/physiology , Alternative Splicing , Animals , Body Patterning , Drosophila melanogaster , Endocytosis , ErbB Receptors/metabolism , Female , In Situ Hybridization , Ovary/metabolism , Phenotype , Protein Isoforms , Protein Structure, Tertiary , Signal Transduction
10.
Development ; 133(16): 3255-64, 2006 Aug.
Article in English | MEDLINE | ID: mdl-16854971

ABSTRACT

Epithelial morphogenesis requires cell movements and cell shape changes coordinated by modulation of the actin cytoskeleton. We identify a role for Echinoid (Ed), an immunoglobulin domain-containing cell-adhesion molecule, in the generation of a contractile actomyosin cable required for epithelial morphogenesis in both the Drosophila ovarian follicular epithelium and embryo. Analysis of ed mutant follicle cell clones indicates that the juxtaposition of wild-type and ed mutant cells is sufficient to trigger actomyosin cable formation. Moreover, in wild-type ovaries and embryos, specific epithelial domains lack detectable Ed, thus creating endogenous interfaces between cells with and without Ed; these interfaces display the same contractile characteristics as the ectopic Ed expression borders generated by ed mutant clones. In the ovary, such an interface lies between the two cell types of the dorsal appendage primordia. In the embryo, Ed is absent from the amnioserosa during dorsal closure, generating an Ed expression border with the lateral epidermis that coincides with the actomyosin cable present at this interface. In both cases, ed mutant epithelia exhibit loss of this contractile structure and subsequent defects in morphogenesis. We propose that local modulation of the cytoskeleton at Ed expression borders may represent a general mechanism for promoting epithelial morphogenesis.


Subject(s)
Cell Adhesion Molecules/metabolism , Drosophila Proteins/metabolism , Drosophila/embryology , Epithelium/embryology , Repressor Proteins/metabolism , Actomyosin/metabolism , Adherens Junctions/metabolism , Animals , Cell Adhesion Molecules/genetics , Cell Movement , Codon, Nonsense , Drosophila/genetics , Drosophila/metabolism , Drosophila Proteins/genetics , Embryo, Nonmammalian/metabolism , Epithelium/metabolism , Female , Morphogenesis/genetics , Mutation , Ovarian Follicle/embryology , Repressor Proteins/genetics
11.
Development ; 133(11): 2115-23, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16672346

ABSTRACT

The dorsoventral axis of the Drosophila egg is established by dorsally localized activation of the epidermal growth factor receptor (Egfr) in the ovarian follicular epithelium. Subsequent positive- and negative-feedback regulation generates two dorsolateral follicle cell primordia that will produce the eggshell appendages. A dorsal midline domain of low Egfr activity between the appendage primordia defines their dorsal boundary, but little is known about the mechanisms that establish their ventral limit. We demonstrate that the transcriptional repressor Capicua is required cell autonomously in ventral and lateral follicle cells to repress dorsal fates, and functions in this process through the repression of mirror. Interestingly, ectopic expression of mirror in the absence of capicua is observed only in the anterior half of the epithelium. We propose that Capicua regulates the pattern of follicle cell fates along the dorsoventral axis by blocking the induction of appendage determinants, such as mirror, by anterior positional cues.


Subject(s)
Cell Lineage , Drosophila Proteins/antagonists & inhibitors , Drosophila Proteins/metabolism , Drosophila melanogaster/cytology , Drosophila melanogaster/metabolism , Eye Proteins/antagonists & inhibitors , Homeodomain Proteins/antagonists & inhibitors , Ovarian Follicle/cytology , Ovarian Follicle/metabolism , Repressor Proteins/metabolism , Transcription Factors/antagonists & inhibitors , Animals , Down-Regulation , Drosophila Proteins/genetics , Drosophila melanogaster/genetics , Eye Proteins/genetics , Eye Proteins/metabolism , Female , Gene Expression Regulation, Developmental , HMGB Proteins , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Homozygote , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mutation/genetics , N-Acetylglucosaminyltransferases/genetics , N-Acetylglucosaminyltransferases/metabolism , Ovarian Follicle/growth & development , Ovum/metabolism , Phenotype , Repressor Proteins/genetics , Transcription Factors/genetics , Transcription Factors/metabolism , Transforming Growth Factor alpha/genetics , Transforming Growth Factor alpha/metabolism
12.
Development ; 132(10): 2345-53, 2005 May.
Article in English | MEDLINE | ID: mdl-15829517

ABSTRACT

The asymmetric localization of gurken mRNA and protein in the developing Drosophila oocyte defines both the anteroposterior and dorsoventral axes of the future embryo. Understanding the origin of these asymmetries requires knowledge of the source of gurken transcripts. During oogenesis most transcripts in the oocyte are produced by the associated nurse cells, but it has been proposed that gurken is an exceptional oocyte-derived transcript. Using a novel application of a standard mitotic recombination technique, we generated mosaic egg chambers in which the nurse cells, but not the oocyte, could produce gurken. Gurken was properly localized in these mosaics and oocyte axial polarity was established normally, indicating that the nurse cells synthesize gurken and that their contribution is sufficient for Gurken function. Our data demonstrate the existence of a mechanism for transport of gurken from the nurse cells and its subsequent localization within the oocyte.


Subject(s)
Body Patterning/physiology , Drosophila Proteins/metabolism , Drosophila/embryology , Oocytes/physiology , Ovarian Follicle/metabolism , Transforming Growth Factor alpha/metabolism , Animals , Female , Green Fluorescent Proteins , Immunohistochemistry , Oocytes/metabolism , Ovarian Follicle/cytology , Protein Transport/physiology , Transgenes/genetics
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