The Stubble-stubbloid locus encodes a transmembrane serine protease (Stubble) necessary for the proper formation of sensory bristles, and the morphogenesis of leg and wing epithelia. Genetic and cell biological analysis indicate a role for Stubble in actin cytoskeletal dynamics and cell shape changes in developing epithelia and bristles. Previously reported genetic interactions between Stubble and the Rho1 signaling pathway suggest Stubble influences actin cytoskeleton dynamics in developing imaginal discs through interactions with the Rho1 pathway. This work will discuss a genetic screen conducted to further investigate the role of Stubble in bristle and imaginal disc morphogenesis. From 50,000 EMS-mutagenized chromosomes 12 enhancers of the recessive sbd201 allele were identified, including 6 new sbd alleles. Consistent with the current understanding of genetic interactions regulating imaginal disc morphogenesis, mutations in two Rho1 pathway genes, zipper (2 alleles) and Rho1, were isolated. Additionally, three new mutant enhancers of sbd201 were isolated, one of which has been identified as an allele of the cadherin gene Dacshous, another as an allele of the muscle myosin heavy chain gene, and the last as an allele of Notopleural (Np). Dominant and recessive mutations in the Stubble locus interact with the Np allele identified in this screen, in regards to both limb and bristle development, respectively. Mutations in the Np locus were first identified in 1936, but this locus remains poorly characterized and has never been cloned The genetic and phenotypic characterization of Np will be discussed along with experiments that have mapped the position of the Np locus to a 50kb region at the border of the 44F12, 45A1 cytological regions.

Το φυλοσύνδετο γονίδιο wiser (CG32711) είναι απαραίτητο για την ανάπτυξη της Drosophila melanogaster. Η μελέτη μιας θερμοευαίσθητης, θανατογόνου μετάλλαξης που ονομάζεται wisertsl αποκάλυψε ότι το γονίδιο wiser εμπλέκεται μεταξύ άλλων στην ανάπτυξη των φτερών. Η μετάλλαξη wisertsl οφείλεται σε ένα P στοιχείο (7E P) που βρίσκεται 490 bp ανοδικά του σημείου έναρξης της μεταγραφής του γονιδίου wiser. 95 bp καθοδικά του 7E P στοιχείου υπάρχει μια P{lacW} ένθεση υπεύθυνη για τη θανατογόνο μετάλλαξη PL26. Οι μεταλλάξεις wisertsl και PL26 είναι αλληλόμορφα του ίδιου γονιδίου ενώ 12000 περίπου βάσεις ανοδικά του γονιδίου wiser και 490 bp ανοδικά του γονιδίου trf2 υπάρχει μια άλλη P{lacW} ένθεση που είναι υπεύθυνη για τη θανατογόνο μετάλλαξη PL28. Οι PL26 και PL28 δεν δείχνουν συμπληρωματικότητα με τη μετάλλαξη wisertsl όσον αφορά το θανατογόνο φαινότυπο στους 29ºC. Όμως το διαγονίδιο UAS-wiser δε διασώζει το θανατογόνο φαινότυπο του PL28. Τα αποτελέσματα της παρούσας εργασίας αποκάλυψαν ότι: 1) Το γονίδιο wiser αλληλεπιδρά με το γονίδιο dpp. Εκτοπική έκφραση του διαγονιδίου (UAS wiser) υπό τον έλεγχο του οδηγού στελέχους apGAL4, μειώνει την έκφραση του dpp στην περιοχή του εμβρυικού δίσκου που θα δώσει τμήμα του θώρακα (notum). 2) Σε ομόζυγα wisertsl άτομα η έκφραση των γονιδίων dpp, dad, omb και salm (όπως αποκαλύπτεται από την έκφραση των αντίστοιχων –lacZ διαγονιδίων) μειώνεται στον εμβρυικό δίσκο του φτερού. Τα παραπάνω γονίδια είναι απαραίτητα για την ανάπτυξη του προσθοπίσθιου άξονα του εμβρυικού δίσκου του φτερού που σημαίνει ότι και το γονίδιο wiser εμπλέκεται στο σχηματισμό του. 3) Το γονίδιο wiser αλληλεπιδρά με το γονίδιο Notch (N) καθώς N wisertsl /wisertsl θηλυκά έχουν εντονότερα φαγωμένα φτερά. 4) Οι μεταλλάξεις wisertsl και PL28 είτε αφορούν και οι δύο το γονίδιο wiser ή η PL28 αφορά το γονίδιο trf2 που σημαίνει ότι και αυτό εμπλέκεται στο σχηματισμό του φτερού. / The X- linked wiser (CG32711) gene is a vital gene for the development of Drosophila melanogaster. The study of a temperature sensitive lethal mutation, named wisertsl, revealed that the wiser gene is implicated among others in the development of wings. The wisertsl mutation is due to a wild P element (7E P) located 490 bp upstream of the presumed transcription start site of the gene wiser at the region 7Ε. 95 bp downstream of the 7E P element is located a P{lacW} responsible for the lethal mutation PL26 and ~ 12000 bp upstream of the gene wiser and 490 bp upstream of the gene trf2 exists another P{lacW} insertion which is responsible of the lethal mutation PL28. The mutations PL26 and PL28 do not show complementation with the wisertsl mutation as regards the lethal phenotype at 29°C. However, while the transgene UAS-wiser saves the lethal phenotypes of wisertsl and PL26 it does not save the lethal phenotype of the mutation PL28. The present data study revealed that: 1) The wiser gene interacts with the dpp gene. Ectoping expression of the UAS wiserCDS construct under the control of apGAL4 driver, reduced the dpp expression (revealed by dpp-lacZ) in the notum territory of the wing imaginal disc. 2) In the homozygous wisertsl individuals the expression of dpp, dad, salm and omb genes (revealed by the corresponding -lacZ strains) is reduced in the wing imaginal disc. The above genes are implicated in the development of the anterior-posterior (A/P) axis of the wing imaginal disc. 3) The wiser gene interacts with the Notch (N) gene. N wisertsl/wisertsl females have stronger notching phenotype. 4) The induction of mitotic clones revealed that the mutation PL28 either concerns an enhancer of the wiser gene or the gene trf2. At the late case the gene trf2 must affect the development of the wings as well.

Two groups of growth regulators were described in Drosophila imaginal disc cell culture Cl.8+. Imaginal disc growth factors (IDGFs) belonging to chitinase-like protein family of carbohydrate binding proteins and Adenosine deaminase-related growth factors (ADGFs), which are active adenosine deaminases influencing homeostasis of key cellular metabolite adenosine. The functions of two of the IDGFs, as well as the effects of extracellular adenosine and its receptor were studied primarily in in vitro cell culture. Our results supported their roles in the regulation of cell survival and energy homeostasis especially in imaginal disc cells. Both the IDGFs and adenosine also play important roles in organismal responses to stress and infection and may interact in vivo.

Epithelial morphogenesis is contingent upon cell shape changes. Cell shape changes are the driving force for the metamorphosis of the adult Drosophila leg from the leg imaginal disc precursor. Genetic analysis has identified several Drosophila genes involved in regulating cell shape changes during leg disc morphogenesis. These include members of the RhoA signaling pathway and the product of the Stubble-stubbloid (Sb-sbd) locus, a transmembrane serine protease. Mutations in the Sb-sbd gene interact genetically with the members of the RhoA signaling pathway, however the nature of the relationship between Sb-sbd serine protease activity and RhoA signaling is not understood. To identify additional components of the RhoA signaling pathway that may help us to understand the role of the Sb-sbd protease in RhoA signaling the Drosophila genome was systematically scanned for genes that interact with Sb-sbd and RhoA mutations using deletions/deficiencies of specified regions of each chromosome. A total of 201 deficiencies uncovering approximately 84.9-91% of the euchromatic genome and spanning the X, second, and third chromosoms were tested. Of the 201 deficiencies tested, five putative interacting genetic regions and one gene within these deficiencies were identified. The candidate gene Eip78C encodes a nuclear steroid hormone receptor previously identified as having an important role in metamorphosis.

Le réticulum endoplasmique (RE) est un organite assurant de nombreuses fonctionscellulaires telles que la conformation et des modifications post-traductionnelles des protéines ou lemaintien de l’homéostasie calcique. Cet organite est donc un site crucial pour réguler le maintien del’homéostasie cellulaire et tissulaire des organismes multicellulaires. Des altérations de ses fonctionsconduisent à l’accumulation de protéines mal-conformées qui sont observées dans de nombreusespathologies humaines telles que des cancers ou des maladies inflammatoires chroniques. Ce stressdéclenche une réponse adaptative connue sous le nom de réponse aux protéines mal-conformées(UPR) qui permet à la cellule de supprimer ses sources et conséquences. Néanmoins, l’intensité et lachronicité du stress peuvent entrainer une modification de l’UPR qui conduit alors à l’élimination dela cellule par apoptose. A ce jour, les processus moléculaires qui permettent à l’UPR d’induirel’apoptose restent flous. De plus, l’implication de l'UPR dans la régulation de processuscompensatoires n'a jamais été étudiée. Mes travaux de thèse apportent une meilleurecompréhension de ces mécanismes à travers l’étude comparative de différents modèles de stresschronique du RE, qui dépendent d’une dérégulation de l’homéostasie protéique et/ou calcique. Ilssoulignent également le rôle essentiel de la branche PERK/ATF4 de l’UPR dans l’induction de deuxvoies parallèles et indépendantes. D’une part, PERK promeut une apoptose dépendante des caspasesvia une répression de l'expression de diap1, et d‘autre part, elle induit un retard de développement àtravers une induction de l’expression de dilp8 dépendante de la voie JNK. Mes données suggèrentégalement une spécificité tissulaire des signalisations déclenchées en réponse à un stress chroniquedu RE. / The endoplasmic reticulum (ER) is an organelle which ensures various cellular functionssuch as protein maturation and folding or calcium homeostasis maintenance. That is why ER is acrucial site of cell and tissue homeostasis regulation in multicellular organisms. Disruption of ERfunctions leads to misfolded-protein accumulation and is observed in a great number of devastatinghuman diseases. This ER stress triggers an adaptive response named Unfolded Protein Response(UPR) in order to attempt to resolve its sources and consequences. Nevertheless, the intensity andchronicity of ER stress can change this response and lead to the apoptosis of stressed cells. To thisdate, the molecular processes that regulate UPR-induced apoptosis remain unclear. Furthermore, theUPR contribution in the modulation of compensatory mechanisms in response to ER stress has neverbeen studied. This work contributes to a better understanding of these processes through acomparative study of various chronic ER stresses, which depend on the disruption of proteostasis orcalcium homeostasis. During my thesis, I have established the essential role of the PERK/ATF4 branchof the UPR in the induction of two parallel and independent pathways. One promotes apoptosisthrough the down-regulation of the diap1 gene while the other interferes with the induction of adevelopmental delay though a JNK signaling-dependent dilp8 expression. My results also suggest thatchronic ER stress response is tissue specific.

No description available.

The development of a multicellular organism, such as a human or an animal, begins with the fertilization of an egg cell. Thereupon the organism grows by repeated cell divisions until the adult size is reached and growth stops. Although it is known that intrinsic mechanisms determine the final size of developing organs and organisms, the basic principles of growth control are still poorly understood. However, there is strong evidence that certain morphogens, which are a special class of signaling molecules, act as growth factors and play a key role in growth control. In this work, growth control is studied from a mainly theoretical viewpoint. A discrete vertex model describing the organization of cells by a network of polygons is used, including a description of the cell cycle and a description of dynamical morphogen distributions. Self-organized growth is studied by introducing growth rules that govern cell divisions based on the local morphogen level. This discrete description is complemented by a continuum theory to gain further insight into the dynamics of self-organized growth processes. The theoretical description is applied to the developing wing of the fruit fly Drosophila melanogaster. In the developing wing, which is an epithelium consisting of single-layered cell sheets, the morphogen Decapentaplegic (Dpp) acts as a key growth factor. Experimental data shows that the Dpp distribution is dynamic and adapts to the size of the developing wing. Two mechanisms that rely on a regulatory molecule species and lead to such a dynamic behaviour of the Dpp distribution are studied. Several growth rules are tested and the resulting growth behaviour is quantitatively compared to experimental data of the developing wing. A particular growth rule, that triggers a cell division when the local morphogen level has increased by a certain relative amount, is found to be consistent with experimental observations under normal and several perturbed conditions. It is shown that mechanical stresses that arise due to spatial growth inhomogeneities can have a stabilizing effect on the growth process.

Animal morphogenesis is largely driven by concerted changes in the shape of individual cells. However, how cell shape changes are regulated and coordinated in developing animals is not well understood. Here we show that the two perpendicular signaling gradients of the morphogens Dpp, a TGF-β homologue, and Wingless, a Wnt family member, maintain tissue homoeostasis and control cell shape changes in the developing Drosophila wing. Clones of cells lacking Dpp or Wingless signaling invaginate apically, shorten apico-basally and subsequently extrude basally without disruption of the epithelium. During early larval development, the onset of Dpp and Wingless signaling correlates with the cuboidal-to-columnar cell shape transition of wing disc cells. Gradients in apical-basal length of columnar cells correlate during late larval development with the gradients of Dpp and Wingless signaling activities. Cells receiving high levels of Dpp and Wingless signaling are most elongated and apically constricted. Low levels of Dpp and Wingless signaling correlate with a shorter and apically wider cell morphology. Dpp and Wingless signaling is cell-autonomously required for maintaining the elongated columnar cell shape of late larval wing disc cells. Overactivation of these pathways results in precocious cell elongation during early larval development. These morphogenetic responses to Dpp and Wingless require the transcription factor complexes Mad and Tcf/β-catenin, respectively, indicating that they are mediated by changes in gene expression. The morphogenetic function of Wingless is in part mediated by one of its target genes, the transcription factor Vestigial. Wingless signaling promotes an enrichment of E-cadherin at the adherens junctions, and we show that E-cadherin is required to maintain apical-basal cell length. Dpp signaling controls the subcellular distribution of the activities of the small GTPase Rho1 and the regulatory light chain of non-muscle myosin II (MRLC). Alteration of Rho1 or MRLC activity has a profound effect on apical-basal cell length. Finally, we demonstrate that a decrease in Rho1 or MRLC activity rescues the shortening of cells with compromised Dpp signaling. Our results identify cell-autonomous roles for Dpp and Wingless signaling in promoting and maintaining the elongated columnar shape of wing disc cells. Furthermore, they suggest that Dpp and Wingless signaling control cell shape by regulating the actin-MyosinII/E-cadherin network. / Morphogenese in Tieren wird in hohem Maße von konzertierten Zellformveränderungen einzelner Zellen bewirkt. Es ist jedoch noch nicht hinreichend verstanden, wie Zellformveränderungen in sich entwickelnden Tieren reguliert und koordiniert werden. Hier zeigen wir, dass die zwei zueinander senkrecht stehenden Signalgradienten der Morphogene Dpp, eines TGF-β Homologs, und Wingless, eines Mitglieds der Wnt Familie, im sich entwickelnden Drosophila-Flügel Gewebe-Homöostase aufrechterhalten und Zellformveränderungen kontrollieren. Klone von Zellen, denen Dpp oder Wingless Signalaktivität fehlt, invaginieren von ihrer apikalen Seite her, verkürzen sich in apiko-basaler Richtung und extruieren im Folgenden auf der basalen Seite des Epithels, ohne es zu zerstören. Während der frühen Larvalentwicklung korreliert das Anschalten der Dpp und Wingless Signale mit der Zellformveränderung der Flügelscheibenzellen von kuboidal zu kolumnar. Gradienten in der apiko-basalen Länge von kolumnaren Zellen korrelieren während der späten Larvalentwicklung mit den Gradienten der Dpp und Wingless Signalaktivitäten. Zellen, die hohe Werte an Dpp und Wingless Signalen empfangen, sind am meisten elongiert und apikal konstringiert. Niedrige Werte von Dpp und Wingless Signalen korrelieren mit kürzerer und apikal weiterer Zellmorphologie. Dpp und Wingless Signale werden zellautonom gebraucht für die Aufrechterhaltung der elongierten Zellform von späten larvalen Flügelscheibenzellen. Die Überaktivierung dieser Signalwege führt zu vorzeitiger Zellverlängerung während der frühen Larvalentwicklung. Diese morphogenetischen Antworten auf Dpp und Wingless benötigen die Transkriptionsfaktor-Komplexe Mad beziehungsweise Tcf/β-catenin, was darauf hindeutet, dass sie durch Änderungen in der Genexpression vermittelt werden. Die morphogenetische Funktion von Wingless wird teilweise durch eines seiner Zielgene, Vestigial, vermittelt. Wingless Signale fördern die Anreicherung von E-cadherin an den Adherensverbindungen. Wir zeigen hier, dass E-cadherin gebraucht wird, um apiko-basale Zelllänge aufrechtzuerhalten. Dpp Signale kontrollieren die subzelluläre Verteilung der Aktivitäten der kleinen GTPase Rho1 und der regulatorischen leichten Kette von nicht-muskulärem Myosin II (MRLC). Eine Änderung in der Rho1 oder MRLC Aktivität hat weitreichende Auswirkungen auf die apiko-basale Zelllänge. Schließlich zeigen wir noch, dass eine Verringerung der Rho1 oder MRLC Aktivitäten die Zellverkürzung von Dpp-Signal kompromittierten Zellen rettet. Unsere Resultate identifizieren zellautonome Rollen für Dpp und Wingless Signale in der Förderung und Aufrechterhaltung der elongierten kolumnaren Zellform von Flügelimaginalscheibenzellen. Darüber hinaus suggerieren sie, dass Dpp und Wingless Signale die Zellform durch die Regulierung des Aktin-MyosinII/E-cadherin-Netzwerks kontrollieren.

Animal morphogenesis is largely driven by concerted changes in the shape of individual cells. However, how cell shape changes are regulated and coordinated in developing animals is not well understood. Here we show that the two perpendicular signaling gradients of the morphogens Dpp, a TGF-β homologue, and Wingless, a Wnt family member, maintain tissue homoeostasis and control cell shape changes in the developing Drosophila wing. Clones of cells lacking Dpp or Wingless signaling invaginate apically, shorten apico-basally and subsequently extrude basally without disruption of the epithelium. During early larval development, the onset of Dpp and Wingless signaling correlates with the cuboidal-to-columnar cell shape transition of wing disc cells. Gradients in apical-basal length of columnar cells correlate during late larval development with the gradients of Dpp and Wingless signaling activities. Cells receiving high levels of Dpp and Wingless signaling are most elongated and apically constricted. Low levels of Dpp and Wingless signaling correlate with a shorter and apically wider cell morphology. Dpp and Wingless signaling is cell-autonomously required for maintaining the elongated columnar cell shape of late larval wing disc cells. Overactivation of these pathways results in precocious cell elongation during early larval development. These morphogenetic responses to Dpp and Wingless require the transcription factor complexes Mad and Tcf/β-catenin, respectively, indicating that they are mediated by changes in gene expression. The morphogenetic function of Wingless is in part mediated by one of its target genes, the transcription factor Vestigial. Wingless signaling promotes an enrichment of E-cadherin at the adherens junctions, and we show that E-cadherin is required to maintain apical-basal cell length. Dpp signaling controls the subcellular distribution of the activities of the small GTPase Rho1 and the regulatory light chain of non-muscle myosin II (MRLC). Alteration of Rho1 or MRLC activity has a profound effect on apical-basal cell length. Finally, we demonstrate that a decrease in Rho1 or MRLC activity rescues the shortening of cells with compromised Dpp signaling. Our results identify cell-autonomous roles for Dpp and Wingless signaling in promoting and maintaining the elongated columnar shape of wing disc cells. Furthermore, they suggest that Dpp and Wingless signaling control cell shape by regulating the actin-MyosinII/E-cadherin network. / Morphogenese in Tieren wird in hohem Maße von konzertierten Zellformveränderungen einzelner Zellen bewirkt. Es ist jedoch noch nicht hinreichend verstanden, wie Zellformveränderungen in sich entwickelnden Tieren reguliert und koordiniert werden. Hier zeigen wir, dass die zwei zueinander senkrecht stehenden Signalgradienten der Morphogene Dpp, eines TGF-β Homologs, und Wingless, eines Mitglieds der Wnt Familie, im sich entwickelnden Drosophila-Flügel Gewebe-Homöostase aufrechterhalten und Zellformveränderungen kontrollieren. Klone von Zellen, denen Dpp oder Wingless Signalaktivität fehlt, invaginieren von ihrer apikalen Seite her, verkürzen sich in apiko-basaler Richtung und extruieren im Folgenden auf der basalen Seite des Epithels, ohne es zu zerstören. Während der frühen Larvalentwicklung korreliert das Anschalten der Dpp und Wingless Signale mit der Zellformveränderung der Flügelscheibenzellen von kuboidal zu kolumnar. Gradienten in der apiko-basalen Länge von kolumnaren Zellen korrelieren während der späten Larvalentwicklung mit den Gradienten der Dpp und Wingless Signalaktivitäten. Zellen, die hohe Werte an Dpp und Wingless Signalen empfangen, sind am meisten elongiert und apikal konstringiert. Niedrige Werte von Dpp und Wingless Signalen korrelieren mit kürzerer und apikal weiterer Zellmorphologie. Dpp und Wingless Signale werden zellautonom gebraucht für die Aufrechterhaltung der elongierten Zellform von späten larvalen Flügelscheibenzellen. Die Überaktivierung dieser Signalwege führt zu vorzeitiger Zellverlängerung während der frühen Larvalentwicklung. Diese morphogenetischen Antworten auf Dpp und Wingless benötigen die Transkriptionsfaktor-Komplexe Mad beziehungsweise Tcf/β-catenin, was darauf hindeutet, dass sie durch Änderungen in der Genexpression vermittelt werden. Die morphogenetische Funktion von Wingless wird teilweise durch eines seiner Zielgene, Vestigial, vermittelt. Wingless Signale fördern die Anreicherung von E-cadherin an den Adherensverbindungen. Wir zeigen hier, dass E-cadherin gebraucht wird, um apiko-basale Zelllänge aufrechtzuerhalten. Dpp Signale kontrollieren die subzelluläre Verteilung der Aktivitäten der kleinen GTPase Rho1 und der regulatorischen leichten Kette von nicht-muskulärem Myosin II (MRLC). Eine Änderung in der Rho1 oder MRLC Aktivität hat weitreichende Auswirkungen auf die apiko-basale Zelllänge. Schließlich zeigen wir noch, dass eine Verringerung der Rho1 oder MRLC Aktivitäten die Zellverkürzung von Dpp-Signal kompromittierten Zellen rettet. Unsere Resultate identifizieren zellautonome Rollen für Dpp und Wingless Signale in der Förderung und Aufrechterhaltung der elongierten kolumnaren Zellform von Flügelimaginalscheibenzellen. Darüber hinaus suggerieren sie, dass Dpp und Wingless Signale die Zellform durch die Regulierung des Aktin-MyosinII/E-cadherin-Netzwerks kontrollieren.

The development of a multicellular organism, such as a human or an animal, begins with the fertilization of an egg cell. Thereupon the organism grows by repeated cell divisions until the adult size is reached and growth stops. Although it is known that intrinsic mechanisms determine the final size of developing organs and organisms, the basic principles of growth control are still poorly understood. However, there is strong evidence that certain morphogens, which are a special class of signaling molecules, act as growth factors and play a key role in growth control. In this work, growth control is studied from a mainly theoretical viewpoint. A discrete vertex model describing the organization of cells by a network of polygons is used, including a description of the cell cycle and a description of dynamical morphogen distributions. Self-organized growth is studied by introducing growth rules that govern cell divisions based on the local morphogen level. This discrete description is complemented by a continuum theory to gain further insight into the dynamics of self-organized growth processes. The theoretical description is applied to the developing wing of the fruit fly Drosophila melanogaster. In the developing wing, which is an epithelium consisting of single-layered cell sheets, the morphogen Decapentaplegic (Dpp) acts as a key growth factor. Experimental data shows that the Dpp distribution is dynamic and adapts to the size of the developing wing. Two mechanisms that rely on a regulatory molecule species and lead to such a dynamic behaviour of the Dpp distribution are studied. Several growth rules are tested and the resulting growth behaviour is quantitatively compared to experimental data of the developing wing. A particular growth rule, that triggers a cell division when the local morphogen level has increased by a certain relative amount, is found to be consistent with experimental observations under normal and several perturbed conditions. It is shown that mechanical stresses that arise due to spatial growth inhomogeneities can have a stabilizing effect on the growth process.