Essential roles of G 12/13 signaling in distinct cell behaviors driving zebrafish convergence and extension gastrulation movements

Department of Pharmacology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
The Journal of Cell Biology (Impact Factor: 9.69). 07/2005; 169(5):777-87. DOI: 10.1083/jcb.200501104
Source: PubMed

ABSTRACT Galpha(12/13) have been implicated in numerous cellular processes, however, their roles in vertebrate gastrulation are largely unknown. Here, we show that during zebrafish gastrulation, suppression of both Galpha(12) and Galpha(13) signaling by overexpressing dominant negative proteins and application of antisense morpholino-modified oligonucleotide translation interference disrupted convergence and extension without changing embryonic patterning. Analyses of mesodermal cell behaviors revealed that Galpha(12/13) are required for cell elongation and efficient dorsalward migration during convergence independent of noncanonical Wnt signaling. Furthermore, Galpha(12/13) function cell-autonomously to mediate mediolateral cell elongation underlying intercalation during notochord extension, likely acting in parallel to noncanonical Wnt signaling. These findings provide the first evidence that Galpha(12) and Galpha(13) have overlapping and essential roles in distinct cell behaviors that drive vertebrate gastrulation.

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Available from: Fang Lin, Jul 25, 2015
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    • "S1pr2 overexpression induced defects in convergence and extension (C&E) movements (revealed by broadening of the axial mesoderm, Fig. 3A versus 3B, red lines), as well as in epiboly [evident from increase in the distance between the deep-cell margin (dcm) and the vegetal pole (VP), Fig. 3A versus 3B, blue lines with double arrows]. At 35 hpf, these embryos displayed a shortened body axis and frequently also cyclopia (Fig. 3D), phenotypes often associated with impaired C&E movements (Heisenberg et al., 2000; Jessen et al., 2002; Lin et al., 2005). Remarkably, the gastrulation defects induced by S1pr2 overexpression were largely suppressed by Ga 13 inhibition, as judged by morphological observation and analysis of ntl expression (Fig. 3C,F). "
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    ABSTRACT: A key process during vertebrate heart development is the migration of bilateral populations of myocardial precursors towards the midline to form the primitive heart tube. In zebrafish, signaling mediated by sphingosine-1-phosphate (S1P) and its cognate G protein-coupled receptor (S1pr2/Mil) is essential for myocardial migration, but the underlying mechanisms remain undefined. Here, we show that suppression of Gα(13) signaling disrupts myocardial migration, leading to the formation of two bilaterally located hearts (cardia bifida). Genetic studies indicate that Gα(13) acts downstream of S1pr2 to regulate myocardial migration through a RhoGEF-dependent pathway. Furthermore, disrupting any component of the S1pr2/Gα(13)/RhoGEF pathway impairs endoderm convergence during segmentation, and the endodermal defects correlate with the extent of cardia bifida. Moreover, endoderm transplantation reveals that the presence of wild-type anterior endodermal cells in Gα(13)-deficient embryos is sufficient to rescue the endoderm convergence defect and cardia bifida, and, conversely, that the presence of anterior endodermal cells defective for S1pr2 or Gα(13) in wild-type embryos causes such defects. Thus, S1pr2/Gα(13) signaling probably acts in the endoderm to regulate myocardial migration. In support of this notion, cardiac-specific expression of Gα(13) fails to rescue cardia bifida in the context of global Gα(13) inhibition. Our data demonstrate for the first time that the Gα(13)/RhoGEF-dependent pathway functions downstream of S1pr2 to regulate convergent movement of the endoderm, an event that is crucial for coordinating myocardial migration.
    Development 01/2013; 140(4). DOI:10.1242/dev.085340 · 6.27 Impact Factor
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    • "DIG-UTP-labeled sense and antisense RNA probes for each par gene were generated as described previously. Whole-mount in situ hybridization was performed as described (Lin et al., 2005). Frozen sections were generated from embryos labeled by whole-mount in situ hybridization using a cryostat. "
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    ABSTRACT: Protease-activated receptors (PARs) play critical roles in hemostasis in vertebrates including zebrafish. However, the zebrafish gene classification appears to be complex, and the expression patterns of par genes are not established. Based on analyses of genomic organization, phylogenetics, protein primary structure, and protein internalization, we report the identification of four zebrafish PARs: par1, par2a, par2b, and par3. This classification differs from one reported previously. We also show that these genes have distinct spatiotemporal expression profiles in embryos and larvae, with par1, par2a, and par2b expressed maternally and ubiquitously during gastrula stages and their expression patterns refined at later stages, and par3 expressed only in 3-day-old larvae. Notably, the expression patterns of zebrafish par1 and par2b resemble those of their mammalian counterparts, suggesting that receptor function is conserved among vertebrates. This conservation is supported by our findings that Par1 and Par2b are internalized following exposure to thrombin and trypsin, respectively.
    Developmental Dynamics 01/2011; 240(1):278-87. DOI:10.1002/dvdy.22517 · 2.67 Impact Factor
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    • "mutants manifest defects in epiboly, internalization, convergence and extension (Babb and Marrs, 2004; Kane et al., 2005; McFarland et al., 2005; Montero et al., 2005; Shimizu et al., 2005; von der Hardt et al., 2007). The requirement of direct and indirect inhibitors of E-cadherin, namely G 12/13 , Wnt11 and p38 (Mapk14a), for normal gastrulation movements underscores the significance of the precise and diverse regulation of cell adhesion in this process (Lin et al., 2005; Ulrich et al., 2005; Zohn et al., 2006; Lin et al., 2009). Because of the cell clumping observed in Ptges-deficient gastrulae (Fig. 1A), we hypothesized that increased cell adhesion contributed to the gastrulation movement phenotype. "
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    ABSTRACT: Gastrulation movements form the germ layers and shape them into the vertebrate body. Gastrulation entails a variety of cell behaviors, including directed cell migration and cell delamination, which are also involved in other physiological and pathological processes, such as cancer metastasis. Decreased Prostaglandin E(2) (PGE(2)) synthesis due to interference with the Cyclooxygenase (Cox) and Prostaglandin E synthase (Ptges) enzymes halts gastrulation and limits cancer cell invasiveness, but how PGE(2) regulates cell motility remains unclear. Here we show that PGE(2)-deficient zebrafish embryos, impaired in the epiboly, internalization, convergence and extension gastrulation movements, exhibit markedly increased cell-cell adhesion, which contributes to defective cell movements in the gastrula. Our analyses reveal that PGE(2) promotes cell protrusive activity and limits cell adhesion by modulating E-cadherin transcript and protein, in part through stabilization of the Snai1a (also known as Snail1) transcriptional repressor, an evolutionarily conserved regulator of cell delamination and directed migration. We delineate a pathway whereby PGE(2) potentiates interaction between the receptor-coupled G protein betagamma subunits and Gsk3beta to inhibit proteasomal degradation of Snai1a. However, overexpression of beta-catenin cannot stabilize Snai1a in PGE(2)-deficient gastrulae. Thus, the Gsk3beta-mediated and beta-catenin-independent inhibition of cell adhesion by Prostaglandins provides an additional mechanism for the functional interactions between the PGE(2) and Wnt signaling pathways during development and disease. We propose that ubiquitously expressed PGE(2) synthesizing enzymes, by promoting the stability of Snai1a, enable the precise and rapid regulation of cell adhesion that is required for the dynamic cell behaviors that drive various gastrulation movements.
    Development 04/2010; 137(8):1327-37. DOI:10.1242/dev.045971 · 6.27 Impact Factor
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