Delphine Cerezo

The Drosophila cytokine receptor Domeless controls border cell migration and epithelial polarization during oogenesis

In mammals, the JAK/STAT (Janus Kinase/Signal Transducer and Activator of Transcription) signaling pathway is activated in response to cytokines and growth factors to control blood cell development, proliferation and cell determination. In Drosophila, a conserved JAK/STAT signaling pathway controls segmentation in embryos, as well as blood cell development and other processes in larvae and adults. During embryogenesis, transduction of the Unpaired [Upd; also known as Outstretched (Os)] ligand through the JAK/STAT pathway requires Domeless, a putative membrane protein with distant homology to vertebrate type I cytokine receptors. We have isolated domeless (dome) in a screen to identify genes essential in epithelial morphogenesis during oogenesis. The level of dome activity is critical for proper border cell migration and is controlled in part through a negative feedback loop. In addition to its essential role in border cells, we show that dome is required in the germarium for the polarization of follicle cells during encapsulation of germline cells. In this process, dome controls the expression of the apical determinant Crumbs. In contrast to the ligand Upd, whose expression is limited to a pair of polar cells at both ends of the egg chamber, dome is expressed in all germline and follicle cells. However, the Dome protein is specifically localized at apicolateral membranes and undergoes ligand-dependent internalization in the follicle cells. dome mutations interact genetically with JAK/STAT pathway genes in border cell migration and abolish the nuclear translocation of Stat92E in vivo. We also show that dome functions downstream of upd and that both the extracellular and intracellular domains of Dome are required for JAK/STAT signaling. Altogether, our data indicate that Dome is an essential receptor molecule for Upd and JAK/STAT signaling during oogenesis.

Drosophila RalA is essential for the maintenance of Jak/Stat signalling in ovarian follicles.

Small GTPases of the Ras‐like (Ral) family are crucial for signalling functions in both normal and cancer cells; however, their role in a developing organism is poorly understood. Here, we identify the Drosophila Ral homologue RalA as a new key regulator of polar‐cell differentiation during oogenesis. Polar cells have a crucial role in patterning the egg chamber and in recruiting border cells, which undergo collective and guided migration. We show that RalA function is essential for the maintenance of anterior and posterior polar‐cell fate and survival. RalA is required cell autonomously to control the expression of polar‐cell‐specific markers, including the Jak/Stat ligand Unpaired. The loss of RalA also causes a cell non‐autonomous phenotype owing to reduced Jak/Stat signalling in neighbouring follicle cells. As a result, border‐cell assembly and migration as well as the polarization of the oocyte are defective. Thus, RalA is required in organizing centres to control proper patterning and migration in vivo.

The Atypical Cadherin Dachsous Controls Left-Right Asymmetry in Drosophila

Left-right (LR) asymmetry is essential for organ development and function in metazoans, but how initial LR cue is relayed to tissues still remains unclear. Here, we propose a mechanism by which the Drosophila LR determinant Myosin ID (MyoID) transfers LR information to neighboring cells through the planar cell polarity (PCP) atypical cadherin Dachsous (Ds). Molecular interaction between MyoID and Ds in a specific LR organizer controls dextral cell polarity of adjoining hindgut progenitors and is required for organ looping in adults. Loss of Ds blocks hindgut tissue polarization and looping, indicating that Ds is a crucial factor for both LR cue transmission and asymmetric morphogenesis. We further show that the Ds/Fat and Frizzled PCP pathways are required for the spreading of LR asymmetry throughout the hindgut progenitor tissue. These results identify a direct functional coupling between the LR determinant MyoID and PCP, essential for non-autonomous propagation of early LR asymmetry.

Signalling crosstalk at the leading edge controls tissue closure dynamics in the Drosophila embryo

Tissue morphogenesis relies on proper differentiation of morphogenetic domains, adopting specific cell behaviours. Yet, how signalling pathways interact to determine and coordinate these domains remains poorly understood. Dorsal closure (DC) of the Drosophila embryo represents a powerful model to study epithelial cell sheet sealing. In this process, JNK (JUN N-terminal Kinase) signalling controls leading edge (LE) differentiation generating local forces and cell shape changes essential for DC. The LE represents a key morphogenetic domain in which, in addition to JNK, a number of signalling pathways converges and interacts (anterior/posterior -AP- determination; segmentation genes, such as Wnt/Wingless; TGFβ/Decapentaplegic). To better characterize properties of the LE morphogenetic domain, we sought out new JNK target genes through a genomic approach: 25 were identified of which 8 are specifically expressed in the LE, similarly to decapentaplegic or puckered. Quantitative in situ gene profiling of this new set of LE genes reveals complex patterning of the LE along the AP axis, involving a three-way interplay between the JNK pathway, segmentation and HOX genes. Patterning of the LE into discrete domains appears essential for coordination of tissue sealing dynamics. Loss of anterior or posterior HOX gene function leads to strongly delayed and asymmetric DC, due to incorrect zipping in their respective functional domain. Therefore, in addition to significantly increasing the number of JNK target genes identified so far, our results reveal that the LE is a highly heterogeneous morphogenetic organizer, sculpted through crosstalk between JNK, segmental and AP signalling. This fine-tuning regulatory mechanism is essential to coordinate morphogenesis and dynamics of tissue sealing.

Drosophila apc regulates delamination of invasive epithelial clusters

Border Cells in the Drosophila ovaries are a useful genetic model for understanding the molecular events underlying epithelial cell motility. During stage 9 of egg chamber development they detach from neighboring stretched cells and migrate between the nurse cells to reach the oocyte. RNAi screening allowed us to identify the dapc1 gene as being critical in this process. Clonal and live analysis showed a requirement of dapc1 in both outer border cells and contacting stretched cells for delamination. This mutant phenotype was rescued by dapc1 or dapc2 expression. Loss of dapc1 function was associated with an abnormal lasting accumulation of β-catenin/Armadillo and E-cadherin at the boundary between migrating border and stretched cells. Moreover, β-catenin/armadillo or E-cadherin downregulation rescued the dapc1 loss of function phenotype. Altogether these results indicate that Drosophila Apc1 is required for dynamic remodeling of β-catenin/Armadillo and E-cadherin adhesive complexes between outer border cells and stretched cells regulating proper delamination and invasion of migrating epithelial clusters.

In Vivo Characterization of Dynein-Driven nanovectors Using Drosophila Oocytes

Molecular motors transport various cargoes including vesicles, proteins and mRNAs, to distinct intracellular compartments. A significant challenge in the field of nanotechnology is to improve drug nuclear delivery by engineering nanocarriers transported by cytoskeletal motors. However, suitable in vivo models to assay transport and delivery efficiency remain very limited. Here, we develop a fast and genetically tractable assay to test the efficiency and dynamics of fluospheres (FS) using microinjection into Drosophila oocytes coupled with time-lapse microscopy. We designed dynein motor driven FS using a collection of dynein light chain 8 (LC8) peptide binding motifs as molecular linkers and characterized in real time the efficiency of the FS movement according to its linker’s sequence. Results show that the conserved LC8 binding motif allows fast perinuclear nanoparticles accumulation in a microtubule and dynein dependent mechanism. These data reveal the Drosophila oocyte as a new valuable tool for the design of motor driven nanovectors.

The Drosophila insulin pathway controls Profilin expression and dynamic actin-rich protrusions during collective cell migration

ABSTRACT Understanding how different cell types acquire their motile behaviour is central to many normal and pathological processes. Drosophila border cells represent a powerful model for addressing this issue and to specifically decipher the mechanisms controlling collective cell migration. Here, we identify the Drosophila Insulin/Insulin-like growth factor signalling (IIS) pathway as a key regulator in controlling actin dynamics in border cells, independently of its function in growth control. Loss of IIS activity blocks the formation of actin-rich long cellular extensions that are important for the delamination and the migration of the invasive cluster. We show that IIS specifically activates the expression of the actin regulator chickadee, the Drosophila homolog of Profilin, which is essential for promoting the formation of actin extensions and migration through the egg chamber. In this process, the transcription factor FoxO acts as a repressor of chickadee expression. Altogether, these results show that local activation of IIS controls collective cell migration through regulation of actin homeostasis and protrusion dynamics. Highlighted Article: The Insulin pathway and FoxO are key regulators of Profilin expression, which is essential for the actin dynamics that support collective cell migration in Drosophila.

Myosin1D is an evolutionarily conserved determinant of animal Left/Right asymmetry

The establishment of Left/Right (LR) asymmetry is fundamental to animal development. While the pathways governing antero-posterior and dorso-ventral patterning are well conserved among different phyla, divergent mechanisms have been implicated in the specification of LR asymmetry in vertebrates and invertebrates. A cilia-driven, directional fluid flow is important for symmetry breaking in numerous vertebrates, including zebrafish1–10. Alternatively, LR asymmetry can be established independently of motile cilia, notably through the intrinsic chirality of the acto-myosin cytoskeleton11–18. Here we show that MyosiniD (Myo1D), which has been previously identified as a key determinant of LR asymmetry in Drosophila12,13, is essential for the formation and the function of the zebrafish LR Organizer (LRO). We show that Myo1D controls the polarity of LRO cilia and interacts functionally with the Planar Cell Polarity (PCP) gene VanGogh-like2 (Vangl2)19, to promote the establishment of a functional LRO flow. Our findings identify Myo1D as the first evolutionarily conserved determinant of LR asymmetry, and show that functional interactions between Myo1D and PCP are central to the establishment of animal LR asymmetry.