[I lead, follow me! How cells coordinate during collective migrations.] / Je mène donc tu suis - Comment les cellules se coordonnent lors de migrations collectives.

During development and wound healing, cells frequently move in a so-called "collective cell migration" process. The same type of migration is used by some cancer cells during metastasis formation. A powerful model to study collective cell migration is the border cell cluster in Drosophila as it allows the observation and manipulation of a collective cell migration in its normal environment. This review describes the molecular machinery used by the border cells to migrate directionally, focusing on the mechanisms used to detect and reacts to chemoattractants, and to organise the group in leader and follower cells.

Title: Je mène donc tu suis - Comment les cellules se coordonnent lors de migrations collectives. Abstract: Lors du développement et de la cicatrisation, les cellules se déplacent souvent par un processus de « migration cellulaire collective ¼. Un procédé identique est employé par les cellules de certaines tumeurs cancéreuses lors de la formation de métastases. Un remarquable modèle d'étude de la migration cellulaire collective est celui de l'étude du groupe (cluster) de cellules de bordure de la drosophile, qui permet d'observer et de manipuler une migration collective dans son environnement naturel. Cette revue décrit la machinerie moléculaire qui permet à ce groupe de cellules de migrer directionnellement, en se concentrant sur les mécanismes permettant aux cellules de détecter et réagir aux chimioattractants et d'organiser le groupe en cellules leaders et suiveuses.

ArfGAP1 regulates the endosomal sorting of guidance receptors to promote directed collective cell migration in vivo.

Chemotaxis drives diverse migrations important for development and involved in diseases, including cancer progression. Using border cells in the Drosophila egg chamber as a model for collective cell migration, we characterized the role of ArfGAP1 in regulating chemotaxis during this process. We found that ArfGAP1 is required for the maintenance of receptor tyrosine kinases, the guidance receptors, at the plasma membrane. In the absence of ArfGAP1, the level of active receptors is reduced at the plasma membrane and increased in late endosomes. Consequently, clusters with impaired ArfGAP1 activity lose directionality. Furthermore, we found that the number and size of late endosomes and lysosomes are increased in the absence of ArfGAP1. Finally, genetic interactions suggest that ArfGAP1 acts on the kinase and GTPase Lrrk to regulate receptor sorting. Overall, our data indicate that ArfGAP1 is required to maintain guidance receptors at the plasma membrane and promote chemotaxis.

Dual regulation of Misshapen by Tao and Rap2l promotes collective cell migration.

Collective cell migration occurs in various biological processes such as development, wound healing and metastasis. During Drosophila oogenesis, border cells (BC) form a cluster that migrates collectively inside the egg chamber. The Ste20-like kinase Misshapen (Msn) is a key regulator of BC migration coordinating the restriction of protrusion formation and contractile forces within the cluster. Here, we demonstrate that the kinase Tao acts as an upstream activator of Msn in BCs. Depletion of Tao significantly impedes BC migration and produces a phenotype similar to Msn loss-of-function. Furthermore, we show that the localization of Msn relies on its CNH domain, which interacts with the small GTPase Rap2l. Our findings indicate that Rap2l promotes the trafficking of Msn to the endolysosomal pathway. When Rap2l is depleted, the levels of Msn increase in the cytoplasm and at cell-cell junctions between BCs. Overall, our data suggest that Rap2l ensures that the levels of Msn are higher at the periphery of the cluster through the targeting of Msn to the degradative pathway. Together, we identified two distinct regulatory mechanisms that ensure the appropriate distribution and activation of Msn in BCs.

CNK2 promotes cancer cell motility by mediating ARF6 activation downstream of AXL signalling.

Cell motility is a critical feature of invasive tumour cells that is governed by complex signal transduction events. Particularly, the underlying mechanisms that bridge extracellular stimuli to the molecular machinery driving motility remain partially understood. Here, we show that the scaffold protein CNK2 promotes cancer cell migration by coupling the pro-metastatic receptor tyrosine kinase AXL to downstream activation of ARF6 GTPase. Mechanistically, AXL signalling induces PI3K-dependent recruitment of CNK2 to the plasma membrane. In turn, CNK2 stimulates ARF6 by associating with cytohesin ARF GEFs and with a novel adaptor protein called SAMD12. ARF6-GTP then controls motile forces by coordinating the respective activation and inhibition of RAC1 and RHOA GTPases. Significantly, genetic ablation of CNK2 or SAMD12 reduces metastasis in a mouse xenograft model. Together, this work identifies CNK2 and its partner SAMD12 as key components of a novel pro-motility pathway in cancer cells, which could be targeted in metastasis.

MAP4K4 regulates forces at cell-cell and cell-matrix adhesions to promote collective cell migration.

Collective cell migration is not only important for development and tissue homeostasis but can also promote cancer metastasis. To migrate collectively, cells need to coordinate cellular extensions and retractions, adhesion sites dynamics, and forces generation and transmission. Nevertheless, the regulatory mechanisms coordinating these processes remain elusive. Using A431 carcinoma cells, we identify the kinase MAP4K4 as a central regulator of collective migration. We show that MAP4K4 inactivation blocks the migration of clusters, whereas its overexpression decreases cluster cohesion. MAP4K4 regulates protrusion and retraction dynamics, remodels the actomyosin cytoskeleton, and controls the stability of both cell-cell and cell-substrate adhesion. MAP4K4 promotes focal adhesion disassembly through the phosphorylation of the actin and plasma membrane crosslinker moesin but disassembles adherens junctions through a moesin-independent mechanism. By analyzing traction and intercellular forces, we found that MAP4K4 loss of function leads to a tensional disequilibrium throughout the cell cluster, increasing the traction forces and the tension loading at the cell-cell adhesions. Together, our results indicate that MAP4K4 activity is a key regulator of biomechanical forces at adhesion sites, promoting collective migration.

SUMO Proteomics Analyses Identify Protein Inhibitor of Activated STAT-Mediated Regulatory Networks Involved in Cell Cycle and Cell Proliferation.

Protein inhibitor of activated STAT (PIAS) proteins are E3 SUMO ligases playing important roles in protein stability and signaling transduction pathways. PIAS proteins are overexpressed in the triple-negative breast cancer cell line MDA-MB-231, and PIAS knockout (KO) results in a reduction in cell proliferation and cell arrest in the S phase. However, the molecular mechanisms underlying PIAS functions in cell proliferation and cell cycle remain largely unknown. Here, we used quantitative SUMO proteomics to explore the regulatory role of PIAS SUMO E3 ligases upon CRISPR/Cas9 KO of individual PIAS. A total of 1422 sites were identified, and around 10% of SUMO sites were regulated following KO of one or more PIAS genes. We identified protein substrates that were either specific to individual PIAS ligase or regulated by several PIAS ligases. Ki-67 and TOP2A, which are involved in cell proliferation and epithelial-to-mesenchymal transition, are SUMOylated at several lysine residues by all PIAS ligases, suggesting a level of redundancy between these proteins. Confocal microscopy and biochemical experiments revealed that SUMOylation regulated TOP2A protein stability, while this modification is involved in the recruitment of Ki-67 nucleolar proteins containing the SUMO interacting motif. These results provide novel insights into both the redundant and specific regulatory mechanisms of cell proliferation and cell cycle mediated by PIAS SUMO E3 ligases.

Directing with restraint: Mechanisms of protrusion restriction in collective cell migrations.

Cell migration is necessary for morphogenesis, tissue homeostasis, wound healing and immune response. It is also involved in diseases. In particular, cell migration is inherent in metastasis. Cells can migrate individually or in groups. To migrate efficiently, cells need to be able to organize into a leading front that protrudes by forming membrane extensions and a trailing edge that contracts. This organization is scaled up at the group level during collective cell movements. If a cell or a group of cells is unable to limit its leading edge and hence to restrict the formation of protrusions to the front, directional movements are impaired or abrogated. Here we summarize our current understanding of the mechanisms restricting protrusion formation in collective cell migration. We focus on three in vivo examples: the neural crest cell migration, the rotatory migration of follicle cells around the Drosophila egg chamber and the border cell migration during oogenesis.

Rab11FIP1 maintains Rab35 at the intercellular bridge to promote actin removal and abscission.

Cytokinesis occurs at the end of mitosis/meiosis wherein the cytoplasms of daughter cells are separated. Before abscission, an intercellular bridge containing the remaining furrowing machinery, mitotic spindle and actin cytoskeleton connects the two daughter cells. To remove this actin and allow for the separation of daughter cells, Rab35 vesicles, loaded with the actin oxidizer MICAL1 and the inositol polyphosphate 5-phosphatase OCRL, are recruited to the midbody in a fine-tuned spatiotemporal manner. However, importantly, the means by which these vesicles are recruited is currently unclear. Here, we demonstrate that Rab11FIP1 is recruited to the midbody after Rab35 to scaffold it at the bridge and maintain Rab35 in this region. In the absence of Rab11FIP1, Rab35 dramatically drops from the midbody, inducing defects, such as cytokinetic delays and binucleation due to actin overaccumulation at the intercellular bridge, which can be rescued with Latrunculin A treatment. Importantly, we show that Rab11FIP1 is critical for Rab35 function in actin removal prior to cytokinesis. This article has an associated First Person interview with the first author of the paper.

Temporal Coordination of Collective Migration and Lumen Formation by Antagonism between Two Nuclear Receptors.

During development, cells undergo multiple, distinct morphogenetic processes to form a tissue or organ, but how their temporal order and time interval are determined remain poorly understood. Here we show that the nuclear receptors E75 and DHR3 regulate the temporal order and time interval between the collective migration and lumen formation of a coherent group of cells named border cells during Drosophila oogenesis. We show that E75, in response to ecdysone signaling, antagonizes the activity of DHR3 during border cell migration, and DHR3 is necessary and sufficient for the subsequent lumen formation that is critical for micropyle morphogenesis. DHR3's lumen-inducing function is mainly mediated through ßFtz-f1, another nuclear receptor and transcription factor. Furthermore, both DHR3 and ßFtz-f1 are required for chitin secretion into the lumen, whereas DHR3 is sufficient for chitin secretion. Lastly, DHR3 and ßFtz-f1 suppress JNK signaling in the border cells to downregulate cell adhesion during lumen formation.

Quantitative SUMO proteomics identifies PIAS1 substrates involved in cell migration and motility.

The protein inhibitor of activated STAT1 (PIAS1) is an E3 SUMO ligase that plays important roles in various cellular pathways. Increasing evidence shows that PIAS1 is overexpressed in various human malignancies, including prostate and lung cancers. Here we used quantitative SUMO proteomics to identify potential substrates of PIAS1 in a system-wide manner. We identified 983 SUMO sites on 544 proteins, of which 62 proteins were assigned as putative PIAS1 substrates. In particular, vimentin (VIM), a type III intermediate filament protein involved in cytoskeleton organization and cell motility, was SUMOylated by PIAS1 at Lys-439 and Lys-445 residues. VIM SUMOylation was necessary for its dynamic disassembly and cells expressing a non-SUMOylatable VIM mutant showed a reduced level of migration. Our approach not only enables the identification of E3 SUMO ligase substrates but also yields valuable biological insights into the unsuspected role of PIAS1 and VIM SUMOylation on cell motility.

The ArfGAP Drongo Promotes Actomyosin Contractility during Collective Cell Migration by Releasing Myosin Phosphatase from the Trailing Edge.

Collective cell migration is involved in various developmental and pathological processes, including the dissemination of various cancer cells. During Drosophila melanogaster oogenesis, a group of cells called border cells migrate collectively toward the oocyte. Herein, we show that members of the Arf family of small GTPases and some of their regulators are required for normal border cell migration. Notably, we found that the ArfGAP Drongo and its GTPase-activating function are essential for the initial detachment of the border cell cluster from the basal lamina. We demonstrate through protein localization and genetic interactions that Drongo controls the localization of the myosin phosphatase in order to regulate myosin II activity at the back of the cluster. Moreover, we show that toward the class III Arf, Drongo acts antagonistically to the guanine exchange factor Steppke. Overall, our work describes a mechanistic pathway that promotes the local actomyosin contractility necessary for border cell detachment.

Misshapen coordinates protrusion restriction and actomyosin contractility during collective cell migration.

Collective cell migration is involved in development, wound healing and metastasis. In the Drosophila ovary, border cells (BC) form a small cluster that migrates collectively through the egg chamber. To achieve directed motility, the BC cluster coordinates the formation of protrusions in its leader cell and contractility at the rear. Restricting protrusions to leader cells requires the actin and plasma membrane linker Moesin. Herein, we show that the Ste20-like kinase Misshapen phosphorylates Moesin in vitro and in BC. Depletion of Misshapen disrupts protrusion restriction, thereby allowing other cells within the cluster to protrude. In addition, we show that Misshapen is critical to generate contractile forces both at the rear of the cluster and at the base of protrusions. Together, our results indicate that Misshapen is a key regulator of BC migration as it coordinates two independent pathways that restrict protrusion formation to the leader cells and induces contractile forces.

Care pathway, towards the establishment of tailored funding: Reasons and criteria for success.

Care pathways are often at the forefront of political thinking about health care practices in France without ever finding a durable means for their extension. Closely linked to funding of healthcare system, they have, once again, been the object of so many economical discussions in 2017, as part of a more optimistic climate of governance which is therefore more open to change. Our changing system, the development and increasingly chronic nature of diseases, the scale of technological breakthroughs, these are all factors driving this topic forward. The object of this work, after a necessary study of the semantics of the term "pathway" and even "funding", was to identify all prerequisites and good practices for the stakeholders to develop a pilot pathway and then its relevant implementation in France. To do so, the members of the Round Table have relied on the presentation of examples of care pathways in order to identify triggers to a progressive, adapted extension to the whole territory. The group has identified key elements and priorities for the establishment of public funding beyond existing funding to incentive team work, particularly in the case of treatment rupture points and/or when they have diverging interests. Finally, creating a climate of confidence among patients, professionals, hospitals, the ARS, payers and manufacturers in handling change management will become the key challenges of the implementation of future pathways.

Non-autonomous role of Cdc42 in cell-cell communication during collective migration.

Collective cell migration is involved in numerous processes both physiological, such as embryonic development, and pathological such as metastasis. Compared to single cell migration, collective motion requires cell behaviour coordination through an as-yet poorly understood but critical cell-cell communication mechanism. Using Drosophila border cell migration, we show here that the small Rho GTPase Cdc42 regulates cell-cell communication. Indeed, we demonstrate that Cdc42 controls protrusion formation in a cell non-autonomous manner. Moreover, we found that the endocytic small GTPase Rab11, controls Cdc42 localisation to the periphery of migrating border cell clusters. Accordingly, over-expression of Cdc42 in border cells rescues the loss of Rab11 function. In addition, we showed that Cdc42 acts upstream of Moesin, a cytoskeletal regulator known to function downstream of rab11. Thus, our study positions Cdc42 as a new key player in cell-cell communication, acting downstream of Rab11.

Proteomics Screen Identifies Class I Rab11 Family Interacting Proteins as Key Regulators of Cytokinesis.

The 14-3-3 protein family orchestrates a complex network of molecular interactions that regulates various biological processes. Owing to their role in regulating the cell cycle and protein trafficking, 14-3-3 proteins are prevalent in human diseases such as cancer, diabetes, and neurodegeneration. 14-3-3 proteins are expressed in all eukaryotic cells, suggesting that they mediate their biological functions through evolutionarily conserved protein interactions. To identify these core 14-3-3 client proteins, we used an affinity-based proteomics approach to characterize and compare the human and Drosophila 14-3-3 interactomes. Using this approach, we identified a group of Rab11 effector proteins, termed class I Rab11 family interacting proteins (Rab11-FIPs), or Rip11 in Drosophila We found that 14-3-3 binds to Rip11 in a phospho-dependent manner to ensure its proper subcellular distribution during cell division. Our results indicate that Rip11 plays an essential role in the regulation of cytokinesis and that this function requires its association with 14-3-3 but not with Rab11. Together, our results suggest an evolutionarily conserved role for 14-3-3 in controlling Rip11-dependent protein transport during cytokinesis.

Spatial regulation of greatwall by Cdk1 and PP2A-Tws in the cell cycle.

Entry into mitosis requires the phosphorylation of multiple substrates by cyclin B-Cdk1, while exit from mitosis requires their dephosphorylation, which depends largely on the phosphatase PP2A in complex with its B55 regulatory subunit (Tws in Drosophila). At mitotic entry, cyclin B-Cdk1 activates the Greatwall kinase, which phosphorylates Endosulfine proteins, thereby activating their ability to inhibit PP2A-B55 competitively. The inhibition of PP2A-B55 at mitotic entry facilitates the accumulation of phosphorylated Cdk1 substrates. The coordination of these enzymes involves major changes in their localization. In interphase, Gwl is nuclear while PP2A-B55 is cytoplasmic. We recently showed that Gwl suddenly relocalizes from the nucleus to the cytoplasm in prophase, before nuclear envelope breakdown and that this controlled localization of Gwl is required for its function. We and others have shown that phosphorylation of Gwl by cyclin B-Cdk1 at multiple sites is required for its nuclear exclusion, but the precise mechanisms remained unclear. In addition, how Gwl returns to its nuclear localization was not explored. Here we show that cyclin B-Cdk1 directly inactivates a Nuclear Localization Signal in the central region of Gwl. This phosphorylation facilitates the cytoplasmic retention of Gwl, which is exported to the cytoplasm in a Crm1-dependent manner. In addition, we show that PP2A-Tws promotes the return of Gwl to its nuclear localization during cytokinesis. Our results indicate that the cyclic changes in Gwl localization at mitotic entry and exit are directly regulated by the antagonistic cyclin B-Cdk1 and PP2A-Tws enzymes.

In vitro and in vivo characterization of the Rab11-GAP activity of Drosophila Evi5.

Small GTPases of the Rab family are master regulators of vesicular trafficking. As such, they control the spatial distribution of various proteins, including proteins involved in cell signaling and the regulation of cell polarity. Misregulation of Rab proteins is associated with a large array of diseases. Surprisingly, the target of some key regulators of Rab proteins, including many GTPase-activating protein (GAP) is still unknown. Identifying the target of a specific GAP requires the combination of both in vitro and in vivo experiments to avoid any misinterpretation. Here is described the methodology we used to characterize the Rab11-GAP activity of Drosophila Evi5. We first focus on the in vitro Rab11 effector pull-down assay we developed and then we detail the in vivo characterization of Rab11 activity during Drosophila border cell migration.

[UNICANCER patient expectations observatory: a new role for patients in health care institutions]. / L'observatoire des attentes des patients UNICANCER: une nouvelle place des usagers dans les établissements de santé.

UNICANCER, recognizing the role played by patients in their own management, set up a unique initiative in France in November 2011: the patient expectations observatory, which is designed to reorient and improve the quality of care provided by comprehensive cancer centers of the UNICANCER group based on a better knowledge and understanding of patient perceptions and preferences. An innovative internet-based participative consultation recorded and prioritized patient expectations. Patient management improvement actions in cancer centres were then generalized to equitably satisfy the identified patient expectations. By using patient expectations concerning organization of health care, cancer cancers therefore provide an example of the new modalities of patient participation in health care institutions, in line with the changes proposed by public authorities.

Cell coordination of collective migration by Rab11 and Moesin.

Cell migration is an important process involved in developmental events and in pathologies such as cancer. Cell migration can be classified into two types: individual and collective cell movements. Compared with individual migration, collective cell migration is less understood and has drawn increasing attention lately because of its emerging role in cancer spreading. We have recently established that Rab11 is absolutely required for spatial control of Rac1 activity through the control of cell-cell communication during collective movements (Ramel, et al. 2013). Moreover, we demonstrated that Rab11 acts through the control of Moesin activity. Here, we discuss how Rab11 and Moesin could cooperate to transfer forces from cell to cell in order to insure coordinated collective cell migration.