dachshund Potentiates Hedgehog Signaling during Drosophila Retinogenesis

Proper organ patterning depends on a tight coordination between cell proliferation and differentiation. The patterning of Drosophila retina occurs both very fast and with high precision. This process is driven by the dynamic changes in signaling activity of the conserved Hedgehog (Hh) pathway, which coordinates cell fate determination, cell cycle and tissue morphogenesis. Here we show that during Drosophila retinogenesis, the retinal determination gene dachshund (dac) is not only a target of the Hh signaling pathway, but is also a modulator of its activity. Using developmental genetics techniques, we demonstrate that dac enhances Hh signaling by promoting the accumulation of the Gli transcription factor Cubitus interruptus (Ci) parallel to or downstream of fused. In the absence of dac, all Hh-mediated events associated to the morphogenetic furrow are delayed. One of the consequences is that, posterior to the furrow, dac- cells cannot activate a Roadkill-Cullin3 negative feedback loop that attenuates Hh signaling and which is necessary for retinal cells to continue normal differentiation. Therefore, dac is part of an essential positive feedback loop in the Hh pathway, guaranteeing the speed and the accuracy of Drosophila retinogenesis.MINECO Spain grants: (BFU2012-34324, BFU2015- 66040); Research Foundation—Flanders FWO grants: (G.0640.13, G.0791.14, PhD fellowship); Fundação para a Ciência e Tecnologia grant: (IF/01031/2012)

Actin stress fiber organization promotes cell stiffening and proliferation of pre-invasive breast cancer cells

This deposit is composed by the main article and supplementary files of the publication.Studies of the role of actin in tumour progression have highlighted its key contribution in cell softening associated with cell invasion. Here, using a human breast cell line with conditional Src induction, we demonstrate that cells undergo a stiffening state prior to acquiring malignant features. This state is characterized by the transient accumulation of stress fibres and upregulation of Ena/VASP-like (EVL). EVL, in turn, organizes stress fibres leading to transient cell stiffening, ERK-dependent cell proliferation, as well as enhancement of Src activation and progression towards a fully transformed state. Accordingly, EVL accumulates predominantly in premalignant breast lesions and is required for Src-induced epithelial overgrowth in Drosophila. While cell softening allows for cancer cell invasion, our work reveals that stress fibre-mediated cell stiffening could drive tumour growth during premalignant stages. A careful consideration of the mechanical properties of tumour cells could therefore offer new avenues of exploration when designing cancer-targeting therapies.Bloomington Drosophila Stock Centre; Vienna Drosophila Research Center (VDRC); Developmental Studies Hybridoma Bank (DSHB); Fundação para a Ciência e Tecnologia (FCT) grant: (IF/01031/2012); Laço Grant in breast cancer 2015; Alexander von Humboldt Foundation grant: (Alexander von Humboldt Professorship); Liga Portuguesa contra o Cancro/Pfizer.info:eu-repo/semantics/publishedVersio

Enabled and Capping protein play important roles in shaping cell behavior during Drosophila oogenesis

During development, cells craft an impressive array of actin-based structures, mediating events as diverse as cytokinesis, apical constriction, and cell migration. One challenge is to determine how cells regulate actin assembly and disassembly to carry out these cell behaviors. During Drosophila oogenesis diverse cell behaviors are seen in the soma and germline. We used oogenesis to explore developmental roles of two important actin regulators: Enabled/VASP proteins and Capping protein. We found that Enabled plays an important role in cortical integrity of nurse cells, formation of robust bundled actin filaments in late nurse cells that facilitate nurse cell dumping, and migration of somatic border cells. During nurse cell dumping, Enabled localizes to barbed ends of the nurse cell actin filaments, suggesting its mechanism of action. We further pursued this mechanism using mutant Enabled proteins, each affecting one of its protein domains. These data suggest critical roles for the EVH2 domain and its tetramerization subdomain, while the EVH1 domain appears less critical. Enabled appears to be negatively regulated during oogenesis by Abelson kinase. We also explored the function of Capping protein. This revealed important roles in oocyte determination, nurse cell cortical integrity and nurse cell dumping, and support the idea that Capping protein and Enabled act antagonistically during dumping. Together these data reveal places these actin regulators shape oogenesis

In Silico Logical Modelling to Uncover Cooperative Interactions in Cancer

International audienceThe multistep development of cancer involves the cooperation between multiple molecular lesions, as well as complex interactions between cancer cells and the surrounding tumour microenvironment. The search for these synergistic interactions using experimental models made tremendous contributions to our understanding of oncogenesis. Yet, these approaches remain labour-intensive and challenging. To tackle such a hurdle, an integrative, multidisciplinary effort is required. In this article, we highlight the use of logical computational models, combined with experimental validations, as an effective approach to identify cooperative mechanisms and therapeutic strategies in the context of cancer biology. In silico models overcome limitations of reductionist approaches by capturing tumour complexity and by generating powerful testable hypotheses. We review representative examples of logical models reported in the literature and their validation. We then provide further analyses of our logical model of Epithelium to Mesenchymal Transition (EMT), searching for additional cooperative interactions involving inputs from the tumour microenvironment and gain of function mutations in NOTCH

In Silico Logical Modelling to Uncover Cooperative Interactions in Cancer

The multistep development of cancer involves the cooperation between multiple molecular lesions, as well as complex interactions between cancer cells and the surrounding tumour microenvironment. The search for these synergistic interactions using experimental models made tremendous contributions to our understanding of oncogenesis. Yet, these approaches remain labour-intensive and challenging. To tackle such a hurdle, an integrative, multidisciplinary effort is required. In this article, we highlight the use of logical computational models, combined with experimental validations, as an effective approach to identify cooperative mechanisms and therapeutic strategies in the context of cancer biology. In silico models overcome limitations of reductionist approaches by capturing tumour complexity and by generating powerful testable hypotheses. We review representative examples of logical models reported in the literature and their validation. We then provide further analyses of our logical model of Epithelium to Mesenchymal Transition (EMT), searching for additional cooperative interactions involving inputs from the tumour microenvironment and gain of function mutations in NOTCH

Cell Architecture-Dependent Constraints: Critical Safeguards to Carcinogenesis

Animal cells display great diversity in their shape. These morphological characteristics result from crosstalk between the plasma membrane and the force-generating capacities of the cytoskeleton macromolecules. Changes in cell shape are not merely byproducts of cell fate determinants, they also actively drive cell fate decisions, including proliferation and differentiation. Global and local changes in cell shape alter the transcriptional program by a multitude of mechanisms, including the regulation of physical links between the plasma membrane and the nuclear envelope and the mechanical modulation of cation channels and signalling molecules. It is therefore not surprising that anomalies in cell shape contribute to several diseases, including cancer. In this review, we discuss the possibility that the constraints imposed by cell shape determine the behaviour of normal and pro-tumour cells by organizing the whole interconnected regulatory network. In turn, cell behaviour might stabilize cells into discrete shapes. However, to progress towards a fully transformed phenotype and to acquire plasticity properties, pro-tumour cells might need to escape these cell shape restrictions. Thus, robust controls of the cell shape machinery may represent a critical safeguard against carcinogenesis

The retinal determination gene Dachshund controls the dynamics of cell shape changes during the differentiation of the Drosophila eye

Resumen del póster presentado al 52nd Annual Drosophila Research Conference, celebrado en San Diego, California (US) del 30 de marzo al 3 de abril de 2011.The Drosophila dachshund (dac) gene, the founder member of the DACH subfamily of nuclear proteins, forms part of the retinal determination pathway. Different expression and genetic studies suggest that this network is conserved in mammals. In Drosophila, previous work suggested that dac is required for the initiation of retinal differentiation. Accordingly, dac mutant flies are eyeless. However, in Dach2;Dach1 double mutant mice eyes are still present, suggesting that the functions of dac/DACH genes are not conserved. Due to this discrepancy, we have re-examined dac function during Drosophila eye development. We show that dac is not required neither for normal eye specification nor retinal differentiation, since dac mutant cells can still differentiate as photoreceptors. Furthermore, our data uncover a novel role for dac in the propagation of retinal differentiation: dac-mutant cells show abnormal cell shape dynamics that seems to cause a delay in exiting the morphogenetic furrow (MF) state, affecting MF formation and progression. Our results argue that the role of dac is not to promote the specification of the eye field, but to regulate MF dynamics.Peer reviewe

Homeostasis of the Drosophila adult retina by actin-capping protein and the Hippo pathway

The conserved Hippo signaling pathway regulates multiple cellular events, including tissue growth, cell fate decision and neuronal homeostasis. While the core Hippo kinase module appears to mediate all the effects of the pathway, various upstream inputs have been identified depending on tissue context. We have recently shown that, in the Drosophila wing imaginal disc, actin-Capping Protein and Hippo pathway activities inhibit F-actin accumulation. In turn, the reduction in F-actin sustains Hippo pathway activity, preventing Yorkie nuclear translocation and the upregulation of proliferation and survival genes. Here, we investigate the role of Capping Protein in growth-unrelated events controlled by the Hippo pathway. We provide evidence that loss of Capping Protein induces degeneration of the adult Drosophila retina through misregulation of the Hippo pathway. We propose a model by which F-actin dynamics might be involved in all processes that require the activity of the core Hippo kinase module

The novel SAM domain protein Aveugle is required for Raf activation in the Drosophila EGF receptor signaling pathway

Activation of the Raf kinase by GTP-bound Ras is a poorly understood step in receptor tyrosine kinase signaling pathways. One such pathway, the epidermal growth factor receptor (EGFR) pathway, is critical for cell differentiation, survival, and cell cycle regulation in many systems, including the Drosophila eye. We have identified a mutation in a novel gene, aveugle, based on its requirement for normal photoreceptor differentiation. The phenotypes of aveugle mutant cells in the eye and wing imaginal discs resemble those caused by reduction of EGFR pathway function. We show that aveugle is required between ras and raf for EGFR signaling in the eye and for mitogen-activated protein kinase phosphorylation in cell culture. aveugle encodes a small protein with a sterile α motif (SAM) domain that can physically interact with the scaffold protein connector enhancer of Ksr (Cnk). We propose that Aveugle acts together with Cnk to promote Raf activation, perhaps by recruiting an activating kinase

Subunits of the Drosophila actin-capping protein heterodimer regulate each other at multiple levels.

The actin-Capping Protein heterodimer, composed of the α and β subunits, is a master F-actin regulator. In addition to its role in many cellular processes, Capping Protein acts as a main tumor suppressor module in Drosophila and in humans, in part, by restricting the activity of Yorkie/YAP/TAZ oncogenes. We aimed in this report to understand how both subunits regulate each other in vivo. We show that the levels and capping activities of both subunits must be tightly regulated to control F-actin levels and consequently growth of the Drosophila wing. Overexpressing capping protein α and β decreases both F-actin levels and tissue growth, while expressing forms of Capping Protein that have dominant negative effects on F-actin promote tissue growth. Both subunits regulate each other's protein levels. In addition, overexpressing one of the subunit in tissues knocked-down for the other increases the mRNA and protein levels of the subunit knocked-down and compensates for its loss. We propose that the ability of the α and β subunits to control each other's levels assures that a pool of functional heterodimer is produced in sufficient quantities to restrict the development of tumor but not in excess to sustain normal tissue growth