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Zhang Y, Kim TH, Niswander L. Phactr4 regulates directional migration of enteric neural crest through PP1, integrin signaling, and cofilin activity. Genes Dev 26: 69-81

Howard Hughes Medical Institute, Department of Pediatrics, Graduate Program in Cell Biology, Stem Cells, and Development, Children's Hospital Colorado, University of Colorado, Aurora, Colorado 80045, USA.
Genes & development (Impact Factor: 12.64). 01/2012; 26(1):69-81. DOI: 10.1101/gad.179283.111
Source: PubMed

ABSTRACT Hirschsprung disease (HSCR) is caused by a reduction of enteric neural crest cells (ENCCs) in the gut and gastrointestinal blockage. Knowledge of the genetics underlying HSCR is incomplete, particularly genes that control cellular behaviors of ENCC migration. Here we report a novel regulator of ENCC migration in mice. Disruption of the Phactr4 gene causes an embryonic gastrointestinal defect due to colon hypoganglionosis, which resembles human HSCR. Time-lapse imaging of ENCCs within the embryonic gut demonstrates a collective cell migration defect. Mutant ENCCs show undirected cellular protrusions and disrupted directional and chain migration. Phactr4 acts cell-autonomously in ENCCs and colocalizes with integrin and cofilin at cell protrusions. Mechanistically, we show that Phactr4 negatively regulates integrin signaling through the RHO/ROCK pathway and coordinates protein phosphatase 1 (PP1) with cofilin activity to regulate cytoskeletal dynamics. Strikingly, lamellipodia formation and in vivo ENCC chain migration defects are rescued by inhibition of ROCK or integrin function. Our results demonstrate a previously unknown pathway in ENCC collective migration in vivo and provide new candidate genes for human genetic studies of HSCR.

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    • "NC cells crawl through a variety of extracellular matrix (ECM) including regions rich in fibronectin and laminin (Newgreen and Thiery, 1980; Strachan and Condic, 2003; Brauer and Markwald, 1987). Evidence from chick and mouse time-lapse imaging has revealed that NC cell chain migration occurs in the cranial, trunk and intestinal subregions of the embryo (Kulesa and Fraser 1998; Young, Anderson et al. 2004; Kasemeier-Kulesa, Bradley et al. 2006; Druckenbrod and Epstein 2007; Rupp and Kulesa 2007; Nishiyama, Uesaka et al. 2012; Zhang, Kim et al. 2012). NC cells in nearly every vertebrate model system (chick, mouse, zebrafish, axolotl, turtle, snake) have also been observed to travel in multicellular streams (Reyes et al., 2010; Kulesa and Fraser, 1998; Schilling and Kimmel, 1994; Golding et al., 2000; Epperlein et al., 2007; Gilbert et al., 2007). "
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    ABSTRACT: Directed cell migration often involves at least two types of cell motility that include multicellular streaming and chain migration. However, what is unclear is how cell contact dynamics and the distinct microenvironments through which cells travel influence the selection of one migratory mode or the other. The embryonic and highly invasive neural crest (NC) are an excellent model system to study this question since NC cells have been observed in vivo to display both of these types of cell motility. Here, we present data from tissue transplantation experiments in chick and in silico modeling that test our hypothesis that cell contact dynamics with each other and the microenvironment promote and sustain either multicellular stream or chain migration. We show that when premigratory cranial NC cells (at the pre-otic level) are transplanted into a more caudal region in the head (at the post-otic level), cells alter their characteristic stream behavior and migrate in chains. Similarly, post-otic NC cells migrate in streams after transplantation into the pre-otic hindbrain, suggesting that local microenvironmental signals dictate the mode of NC cell migration. Simulations of an agent-based model (ABM) that integrates the NC cell behavioral data predict that chain migration critically depends on the interplay of biased cell-cell contact and local microenvironment signals. Together, this integrated modeling and experimental approach suggests new experiments and offers a powerful tool to examine mechanisms that underlie complex cell migration patterns.
    Physical Biology 06/2013; 10(3):035003. DOI:10.1088/1478-3975/10/3/035003 · 3.14 Impact Factor
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    • "A complete sequence search for PP1 BMs, RVxF, SILK, MyphoNE, and apoptotic signature for all the proteins from the YTHs was performed (Tables 1, 2, and 3). From the proteins in YTH1, nine do not have any PP1 BM, including PHACTR4, a known PIP (Kim et al. 2007; Zhang et al. 2012). The SILK motif was described as being present in seven different PIPs (Hendrickx et al. 2009) and always N-terminal to a RVxF motif, being 7 to 55 aa distant. "
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    ABSTRACT: Posttranslational protein modifications, in particular reversible protein phosphorylation, are important regulatory mechanisms involved in cellular signaling transduction pathways. Thousands of human proteins are phosphorylatable and the tight regulation of phosphorylation states is crucial for cell maintenance and development. Protein phosphorylation occurs primarily on serine, threonine, and tyrosine residues, through the antagonistic actions of protein kinases and phosphatases. The catalytic subunit of protein phosphatase 1 (PP1), a major Ser/Thr-phosphatase, associates with a large variety of regulatory subunits that define substrate specificity and determine specific cellular pathway responses. PP1 has been shown to bind to different proteins in the brain in order to execute key and differential functions. This work reports the identification of proteins expressed in the human brain that interact with PP1γ1 and PP1γ2 isoforms by the yeast two-hybrid method. An extensive search of PP1-binding motifs was performed for the proteins identified, revealing already known PP1 regulators but also novel interactors. Moreover, our results were integrated with the data of PP1γ interacting proteins from several public web databases, permitting the development of physical maps of the novel interactions. The PP1γ interactome thus obtained allowed for the identification of novel PP1 interacting proteins, supporting novel functions of PP1γ isoforms in the human brain.
    Journal of Molecular Neuroscience 10/2012; 50(1). DOI:10.1007/s12031-012-9902-6 · 2.76 Impact Factor
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    • "To address these issues we studied a second family of RPEL proteins, the Phactr family of PP1 cofactors (Allen et al., 2004; Sagara et al., 2003). In humans, the four Phactr proteins have been implicated in a variety of human diseases, including Parkinson's, myocardial infarction, and cancer (Bankovic et al., 2010; Kathiresan et al., 2009; Smith et al., 2005; Trufant, 2010; Wider et al., 2009), while in the mouse the Phactr4-PP1 interaction is required for neural tube closure and enteric neural cell migration (Kim et al., 2007; Zhang et al., 2012). Each Phactr protein contains an N-terminal RPEL motif together with a C-terminal triple RPEL repeat (Allen et al., 2004; Sagara et al., 2003). "
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    ABSTRACT: The Phactr family of PP1-binding proteins and the myocardin-related transcription factor family of transcriptional coactivators contain regulatory domains comprising three copies of the RPEL motif, a G-actin binding element. We report the structure of a Phactr1 G-actin⋅RPEL domain complex. Three G-actins surround the crank-shaped RPEL domain forming a closed helical assembly. Their spatial relationship is identical to the RPEL-actins within the pentavalent MRTF G-actin⋅RPEL domain complex, suggesting that conserved cooperative interactions between actin⋅RPEL units organize the assembly. In the trivalent Phactr1 complex, each G-actin⋅RPEL unit makes secondary contacts with its downstream actin involving distinct RPEL residues. Similar secondary contacts are seen in G-actin⋅RPEL peptide crystals. Loss-of-secondary-contact mutations destabilize the Phactr1 G-actin⋅RPEL assembly. Furthermore, actin-mediated inhibition of Phactr1 nuclear import requires secondary contact residues in the Phactr1 N-terminal RPEL-N motif, suggesting that it involves interaction of RPEL-N with the C-terminal assembly. Secondary actin contacts by actin-bound RPEL motifs thus govern formation of multivalent actin⋅RPEL assemblies.
    Structure 10/2012; 20(11). DOI:10.1016/j.str.2012.08.031 · 6.79 Impact Factor
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