The many roles of PTK7: a versatile regulator of cell-cell communication

a b s t r a c t PTK7 (protein tyrosine kinase 7) is an evolutionarily conserved transmembrane receptor with functions in various processes ranging from embryonic morphogenesis to epidermal wound repair. Here, we review recent findings indicating that PTK7 is a versatile co-receptor that functions as a molecular switch in Wnt, Semaphorin/Plexin and VEGF signaling pathways. We focus in particular on the role of PTK7 in Wnt signaling, as recent data indicate that PTK7 acts as a Wnt co-receptor, which activates the planar cell polarity pathway, but inhibits canonical Wnt signaling. Ó 2011 Elsevier Inc. All rights reserved. Introduction Cell-cell communication coordinates complex cell movements in embryogenesis as well as adult tissue homeostasis. During embryonic development cell divisions start the generation of a multicellular organism, and are followed by a series of complex and coordinated cell movements necessary for embryonic patterning and organ formation. Collectively, these morphogenetic movements change the shape and form of differentiating tissues through such processes as gastrulation, the closure of the neural tube, and the migration of neural crest cells. In adult organisms coordinated cell movements are relevant for wound healing and regeneration. In order to ensure precise regulation of these processes individual cells have to communicate with each other. To accomplish this, cells send out cues providing positional information that receiving cells translate into cellular asymmetries and directed locomotion. These molecular cues include members of the Wnt family of secreted glycoproteins, which are able to activate a broad range of downstream signaling events depending on cellular context One such molecular switch, PTK7 (protein tyrosine kinase 7) 1 is a transmembrane receptor that regulates morphogenetic processes. Identified in colon carcinoma cells and named colon carcinoma kinase-4 (CCK-4) PTK7 is a Wnt co-receptor involved in the choice of Wnt signaling outcome Recently, we described PTK7/Otk as a novel Wnt co-receptor that confers specificity in response to Wnt ligand

WNT signalling in prostate cancer

Genome sequencing and gene expression analyses of prostate tumours have highlighted the potential importance of genetic and epigenetic changes observed in WNT signalling pathway components in prostate tumours-particularly in the development of castration-resistant prostate cancer. WNT signalling is also important in the prostate tumour microenvironment, in which WNT proteins secreted by the tumour stroma promote resistance to therapy, and in prostate cancer stem or progenitor cells, in which WNT-β-catenin signals promote self-renewal or expansion. Preclinical studies have demonstrated the potential of inhibitors that target WNT receptor complexes at the cell membrane or that block the interaction of β-catenin with lymphoid enhancer-binding factor 1 and the androgen receptor, in preventing prostate cancer progression. Some WNT signalling inhibitors are in phase I trials, but they have yet to be tested in patients with prostate cancer

Live Imaging of Xwnt5A-ROR2 Complexes

Secreted molecules of the Wnt family regulate key decisions in embryogenesis and adult tissue homeostasis by activating a complex network of Wnt signaling pathways. Although the different branches of Wnt signaling have been studied for more than 25 years, fluorophore tagged constructs for live cell imaging of Wnt molecules activating the Wnt/β-catenin pathway have become available only recently. We have generated a fluorophore tagged Wnt construct of the Xenopus Wnt5a protein (Xwnt5A) with the enhanced green fluorescent protein (EGFP), Xwnt5A-EGFP. This construct activates non-canonical Wnt pathways in an endocytosis dependent manner and is capable of compensating for the loss of endogenous Xwnt5A in Xenopus embryos. Strikingly, non-canonical Wnt pathway activation was restricted to short-range signaling while an inhibitory effect was observed in transwell cell cultures taken as long-range signaling model sytem. We used our Xwnt5A-EGFP construct to analyze in vivo binding of Wnt5A to its co-receptor ROR2 on the microscopic and on the molecular level. On the microscopic level, Xwnt5A-EGFP clusters in the membrane and recruits ROR2-mCherry to these clusters. Applying dual-colour dual-focus line-scanning fluorescence correlation spectroscopy on dorsal marginal zone explants, we identified membrane tethered Xwnt5A-EGFP molecules binding to ROR2-mCherry molecules. Our data favour a model, in which membrane-tethered Wnt-5A recruits ROR2 to form large ligand/receptor clusters and signals in an endocytosis-dependent manner

Identifizierung und funktionelle Charakterisierung von PTK7-Liganden in Xenopus laevis

Wnt Signalwege sind evolutionär konserviert und regulieren zahlreiche Prozesse im sich entwickelnden sowie im ausgewachsenen Organismus. Der kanonische Wnt-Signalweg reguliert durch Veränderungen der Gentranskription die Zellproliferation und -differenzierung. Der nicht- kanonische oder planare Zellpolarität-Wnt-Signalweg (PCP) reguliert die Polarität von Zellen sowie die Zellmigration durch Veränderung der Zytoskelettorganisation. Neben signalwegspezifischen Molekülen werden einige Signalmoleküle wie Wnt, Frizzled (Fz), und Dishevelled (Dsh) von beiden Signalwegen gemeinsam genutzt. Bisher ist allerdings unklar wie eine Zelle zwischen den Signalwegen unterscheidet. PTK7 (Protein Tyrosin Kinase 7) reguliert den PCP Wnt Signalweg, welcher beim Schließen des Neuralrohres und der Polarität der Haarsinneszellen im Innenohr von Vertebraten benötigt wird. Bisher wurden einige intrazelluläre Interaktionspartner von PTK7 beschrieben. In Xenopus konnte zum Beispiel gezeigt werden, dass PTK7, Dsh an die innere Zellmembran rekrutiert und dadurch die Migration der Neuralleistenzellen reguliert. Extrazelluläre Interaktionspartner von PTK7 wurden bisher nicht identifiziert. In dieser Studie zeigen wir, dass PTK7 mit Wnt-Liganden interagiert, die im kanonischen Wnt-Signalweg eine Rolle spielen und dass diese Interaktion durch Fz vermittelt wird. Dieses Ergebnis weißt darauf hin, dass PTK7 auch den kanonischen Wnt-Signalweg beeinflussen kann. In der Tat inhibiert PTK7 den kanonischen Wnt-Signalweg sowohl in Xenopus Embryonen als auch in menschlichen Zelllinien. Des Weiteren haben Epistasis-Experimente ergeben, dass die Inhibierung des kanonischen Wnt-Signalweges durch PTK7 übergeordnet von Dsh auf der Ebene des Frizzled Rezeptors stattfindet. Die Verringerung der PTK7 Expression wiederum aktiviert den kanonischen Wnt-Signalweg. Die dadurch ausgelösten Defekte im Neuralrohrschluss können teilweise durch die Inhibierung des Kanonischen Wnt-Signalweges aufgehoben werden, was den inhibitorischen Effekt von PTK7 auf den Wnt-Signalweg bestätigt. Zusätzlich zur Unterdrückung des kanonischen Wnt-Signalweges aktiviert PTK7 die ATF2 vermittelte Gentranskription in Xenopus Embryonen, was auf eine Aktivierung des nicht-kanonischen Wnt-Signalweges hindeutet. Zusammenfassend konnte gezeigt werden das PTK7 in Xenopus den nicht-kanonischen Wnt-Signalweg aktiviert, indem über die Interaktion mit Wnt und Fz der kanonische Wnt-Signalweg inhibiert wird

Cell-state transitions and collective cell movement generate an endoderm-like region in gastruloids.

Shaping the animal body plan is a complex process that involves the spatial organization and patterning of the different germ layers. Recent advances in live imaging have started to unravel the cellular choreography underlying this process in mammals, however, the sequence of events transforming an unpatterned cell ensemble into structured territories is largely unknown. Here, using gastruloids -3D aggregates of mouse embryonic stem cells- we study the formation of one of the three germ layers, the endoderm. We show that the endoderm is generated from an epiblast-like homogeneous state by a three-step mechanism: (i) a loss of E-cadherin mediated contacts in parts of the aggregate leading to the appearance of islands of E-cadherin expressing cells surrounded by cells devoid of E-cadherin, (ii) a separation of these two populations with islands of E-cadherin expressing cells flowing toward the aggregate tip, and (iii) their differentiation into an endoderm population. During the flow, the islands of E-cadherin expressing cells are surrounded by cells expressing T-Brachyury, reminiscent of the process occurring at the primitive streak. Consistent with recent in vivo observations, the endoderm formation in the gastruloids does not require an epithelial-to-mesenchymal transition, but rather a maintenance of an epithelial state for a subset of cells coupled with fragmentation of E-cadherin contacts in the vicinity, and a sorting process. Our data emphasize the role of signaling and tissue flows in the establishment of the body plan

A conserved Oct4/POUV-dependent network links adhesion and migration to progenitor maintenance

SummaryBackgroundThe class V POU domain transcription factor Oct4 (Pou5f1) is a pivotal regulator of embryonic stem cell (ESC) self-renewal and reprogramming of somatic cells to induced pluripotent stem (iPS) cells. Oct4 is also an important evolutionarily conserved regulator of progenitor cell differentiation during embryonic development.ResultsHere we examine the function of Oct4 homologs in Xenopus embryos and compare this to the role of Oct4 in maintaining mammalian embryo-derived stem cells. Based on a combination of expression profiling of Oct4/POUV-depleted Xenopus embryos and in silico analysis of existing mammalian Oct4 target data sets, we defined a set of evolutionary-conserved Oct4/POUV targets. Most of these targets were regulators of cell adhesion. This is consistent with Oct4/POUV phenotypes observed in the adherens junctions in Xenopus ectoderm, mouse embryonic, and epiblast stem cells. A number of these targets could rescue both Oct4/POUV phenotypes in cellular adhesion and multipotent progenitor cell maintenance, whereas expression of cadherins on their own could only transiently support adhesion and block differentiation in both ESC and Xenopus embryos.ConclusionsCurrently, the list of Oct4 transcriptional targets contains thousands of genes. Using evolutionary conservation, we identified a core set of functionally relevant factors that linked the maintenance of adhesion to Oct4/POUV. We found that the regulation of adhesion by the Oct4/POUV network occurred at both transcriptional and posttranslational levels and was required for pluripotency