Dataset Information

In vivo analysis of formation and endocytosis of the Wnt/?-catenin signaling complex in zebrafish embryos.

ABSTRACT: After activation by Wnt/?-Catenin ligands, a multi-protein complex assembles at the plasma membrane as membrane-bound receptors and intracellular signal transducers are clustered into the so-called Lrp6-signalosome [Corrected]. However, the mechanism of signalosome formation and dissolution is yet not clear. Our imaging studies of live zebrafish embryos show that the signalosome is a highly dynamic structure. It is continuously assembled by Dvl2-mediated recruitment of the transducer complex to the activated receptors and partially disassembled by endocytosis. We find that, after internalization, the ligand-receptor complex and the transducer complex take separate routes. The Wnt-Fz-Lrp6 complex follows a Rab-positive endocytic path. However, when still bound to the transducer complex, Dvl2 forms intracellular aggregates. We show that this endocytic process is not only essential for ligand-receptor internalization but also for signaling. The ?2-subunit of the endocytic Clathrin adaptor Ap2 interacts with Dvl2 to maintain its stability during endocytosis. Blockage of Ap2?2 function leads to Dvl2 degradation, inhibiton of signalosome formation at the plasma membrane and, consequently, reduction of signaling. We conclude that Ap2?2-mediated endocytosis is important to maintain Wnt/?-catenin signaling in vertebrates.

SUBMITTER: Hagemann AI

PROVIDER: S-EPMC4163645 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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In vivo analysis of formation and endocytosis of the Wnt/β-catenin signaling complex in zebrafish embryos.

Hagemann Anja I H AI Kurz Jennifer J Kauffeld Silke S Chen Qing Q Reeves Patrick M PM Weber Sabrina S Schindler Simone S Davidson Gary G Kirchhausen Tomas T Scholpp Steffen S

Journal of cell science 20140729 Pt 18

After activation by Wnt/β-Catenin ligands, a multi-protein complex assembles at the plasma membrane as membrane-bound receptors and intracellular signal transducers are clustered into the so-called Lrp6-signalosome [Corrected]. However, the mechanism of signalosome formation and dissolution is yet not clear. Our imaging studies of live zebrafish embryos show that the signalosome is a highly dynamic structure. It is continuously assembled by Dvl2-mediated recruitment of the transducer complex to ...[more]

PMID: 25074807

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Project description:Embryonic axis formation in vertebrates is initiated by the establishment of the dorsal Nieuwkoop blastula organizer, marked by the nuclear accumulation of maternal ?-catenin, a transcriptional effector of canonical Wnt signaling. Known regulators of axis specification include the canonical Wnt pathway components that positively or negatively affect ?-catenin. An involvement of G-protein coupled receptors (GPCRs) was hypothesized from experiments implicating G proteins and intracellular calcium in axis formation, but such GPCRs have not been identified. Mobilization of intracellular Ca(2+) stores generates Ca(2+) transients in the superficial blastomeres of zebrafish blastulae when the nuclear accumulation of maternal ?-catenin marks the formation of the Nieuwkoop organizer. Moreover, intracellular Ca(2+) downstream of non-canonical Wnt ligands was proposed to inhibit ?-catenin and axis formation, but mechanisms remain unclear. Here we report a novel function of Ccr7 GPCR and its chemokine ligand Ccl19.1, previously implicated in chemotaxis and other responses of dendritic cells in mammals, as negative regulators of ?-catenin and axis formation in zebrafish. We show that interference with the maternally and ubiquitously expressed zebrafish Ccr7 or Ccl19.1 expands the blastula organizer and the dorsoanterior tissues at the expense of the ventroposterior ones. Conversely, Ccr7 or Ccl19.1 overexpression limits axis formation. Epistatic analyses demonstrate that Ccr7 acts downstream of Ccl19.1 ligand and upstream of ?-catenin transcriptional targets. Moreover, Ccl19/Ccr7 signaling reduces the level and nuclear accumulation of maternal ?-catenin and its axis-inducing activity and can also inhibit the Gsk3? -insensitive form of ?-catenin. Mutational and pharmacologic experiments reveal that Ccr7 functions during axis formation as a GPCR to inhibit ?-catenin, likely by promoting Ca(2+) transients throughout the blastula. Our study delineates a novel negative, Gsk3?-independent control mechanism of ?-catenin and implicates Ccr7 as a long-hypothesized GPCR regulating vertebrate axis formation.

Dataset Information

In vivo analysis of formation and endocytosis of the Wnt/?-catenin signaling complex in zebrafish embryos.

Publications

In vivo analysis of formation and endocytosis of the Wnt/β-catenin signaling complex in zebrafish embryos.

Similar Datasets

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