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1.
Proc Natl Acad Sci U S A ; 110(42): 16892-7, 2013 Oct 15.
Article in English | MEDLINE | ID: mdl-24082091

ABSTRACT

The posterior lateral line primordium in zebrafish provides an amenable model to study mechanisms of collective cell migration. The directed migration of the cell cluster along the path of Sdf1a chemokine requires two receptors, Cxcr4b and Cxcr7b, which are expressed in the leading and trailing part of the primordium, respectively. The polarized expression of receptors is regulated by Wnt signaling, but downstream players mediating this control remain to be found. Here, we show that the Hox homeobox gene Hoxb8a is a critical component that acts downstream of the Wnt pathway to coordinate the expression of both chemokine receptors. We find that Hoxb8a is expressed in the leading part of the primordium and is required for the correct speed and extent of migration. Hoxb8a expression is dependent upon Wnt activity and needed both for cxcr4b expression and to repress and thus restrict cxcr7b expression to the trailing zone of the primordium. In the absence of Wnt activity, overexpressed Hoxb8a is able to repress cxcr7b but not up-regulate cxcr4b expression. Together with results from expressing dominant activator and repressor constructs, these findings suggest that Hoxb8a is induced by and cooperates with Wnt signaling to up-regulate cxcr4b, and acts through multiple mechanisms to repress cxcr7b expression.


Subject(s)
Gene Expression Regulation, Developmental/physiology , Homeodomain Proteins/metabolism , Receptors, CXCR4/biosynthesis , Receptors, CXCR/biosynthesis , Wnt Signaling Pathway/physiology , Zebrafish Proteins/biosynthesis , Zebrafish/embryology , Animals , Homeodomain Proteins/genetics , Receptors, CXCR/genetics , Receptors, CXCR4/genetics , Up-Regulation/physiology , Zebrafish/genetics , Zebrafish Proteins/genetics
2.
Development ; 140(10): 2235-43, 2013 May.
Article in English | MEDLINE | ID: mdl-23633515

ABSTRACT

We have generated an inducible system to control the timing of transgene expression in zebrafish and chick. An estrogen receptor variant (ERT2) fused to the GAL4 transcriptional activator rapidly and robustly activates transcription within 3 hours of treatment with the drug 4-hydroxy-tamoxifen (4-OHT) in tissue culture and transgenic zebrafish. We have generated a broadly expressed inducible ERT2-GAL4 zebrafish line using the ubiquitin (ubi) enhancer. In addition, use of ERT2-GAL4 in conjunction with tissue-specific enhancers enables the control of transgene expression in both space and time. This spatial restriction and the ability to sustain forced expression are important advantages over the currently used heat-shock promoters. Moreover, in contrast to currently available TET and LexA systems, which require separate constructs with their own unique recognition sequences, ERT2-GAL4 is compatible with the growing stock of UAS lines being generated in the community. We also applied the same inducible system to the chick embryo and find that it is fully functional, suggesting that this strategy is generally applicable.


Subject(s)
Developmental Biology/methods , Gene Expression Regulation, Developmental , Receptors, Estrogen/genetics , Transgenes , Animals , Animals, Genetically Modified , Chick Embryo , Crosses, Genetic , HEK293 Cells , Humans , Immunohistochemistry , In Situ Hybridization , Phenotype , Receptors, Estrogen/metabolism , Tamoxifen/analogs & derivatives , Tamoxifen/pharmacology , Zebrafish , Zebrafish Proteins/metabolism
3.
Development ; 139(12): 2246-53, 2012 Jun.
Article in English | MEDLINE | ID: mdl-22619392

ABSTRACT

The directional migration of many cell populations occurs as a coherent group. An amenable model is provided by the posterior lateral line in zebrafish, which is formed by a cohesive primordium that migrates from head to tail and deposits future neuromasts at intervals. We found that prior to the onset of migration, the compact state of the primordium is not fully established, as isolated cells with lateral line identity are present caudal to the main primordium. These isolated cells are retained in position such that they fuse with the migrating primordium as it advances, and later contribute to the leading zone and terminal neuromasts. We found that the isolated lateral line cells are positioned by two antagonistic cues: Fgf signalling attracts them towards the primordium, which counteracts Sdf1α/Cxcr4b-mediated caudal attraction. These findings reveal a novel chemotactic role for Fgf signalling in which it enables the coalescence of the lateral line primordium from an initial fuzzy pattern into a compact group of migrating cells.


Subject(s)
Chemokines/metabolism , Fibroblast Growth Factors/metabolism , Lateral Line System/embryology , Lateral Line System/metabolism , Signal Transduction , Zebrafish/embryology , Zebrafish/metabolism , Animals , Cell Fusion , Cell Movement , Cell Separation , Lateral Line System/cytology , Models, Biological , Receptors, CXCR/metabolism , Receptors, CXCR4/metabolism , Zebrafish Proteins/metabolism
4.
Development ; 136(16): 2791-801, 2009 Aug.
Article in English | MEDLINE | ID: mdl-19633172

ABSTRACT

Integrins are the major adhesive receptors for extracellular matrix and have various roles in development. To determine their role in cell migration, the gene encoding the beta1 integrin subunit (Itgb1) was conditionally deleted in mouse neural crest cells just after their emigration from the neural tube. We previously identified a major defect in gut colonisation by conditional Itgb1-null enteric neural crest cells (ENCCs) resulting from their impaired migratory abilities and enhanced aggregation properties. Here, we show that the migration defect occurs primarily during the invasion of the caecum, when Itgb1-null ENCCs stop their normal progression before invading the caecum and proximal hindgut by becoming abnormally aggregated. We found that the caecum and proximal hindgut express high levels of fibronectin and tenascin-C, two well-known ligands of integrins. In vitro, tenascin-C and fibronectin have opposite effects on ENCCs, with tenascin-C decreasing migration and adhesion and fibronectin strongly promoting them. Itgb1-null ENCCs exhibited an enhanced response to the inhibitory effect of tenascin-C, whereas they were insensitive to the stimulatory effect of fibronectin. These findings suggest that beta1 integrins are required to overcome the tenascin-C-mediated inhibition of migration within the caecum and proximal hindgut and to enhance fibronectin-dependent migration in these regions.


Subject(s)
Cecum , Cell Movement/physiology , Integrin beta1/metabolism , Intestinal Mucosa , Intestines , Neural Crest/cytology , Animals , Biomarkers/metabolism , Cecum/cytology , Cecum/embryology , Cecum/metabolism , Cell Adhesion/physiology , Cell Shape , Embryo, Mammalian/anatomy & histology , Embryo, Mammalian/physiology , Enteric Nervous System/cytology , Enteric Nervous System/physiology , Fibronectins/metabolism , Integrin beta1/genetics , Intestinal Mucosa/metabolism , Intestines/cytology , Intestines/embryology , Mice , Mice, Knockout , Tenascin/metabolism , Tissue Culture Techniques
5.
Development ; 133(9): 1725-34, 2006 May.
Article in English | MEDLINE | ID: mdl-16571628

ABSTRACT

The enteric nervous system arises mainly from vagal and sacral neural crest cells that colonise the gut between 9.5 and 14 days of development in mice. Using the Cre-LoxP system, we removed beta1 integrins in the neural crest cells when they emerge from the neural tube. beta1-null enteric neural crest cells fail to colonise the gut completely, leading to an aganglionosis of the descending colon, which resembles the human Hirschsprung's disease. Moreover, beta1-null enteric neural crest cells form abnormal aggregates in the gut wall, leading to a severe alteration of the ganglia network organisation. Organotypic cultures of gut explants reveal that beta1-null enteric neural crest cells show impaired adhesion on extracellular matrix and enhanced intercellular adhesion properties. They display migration defects in collagen gels and gut tissue environments. We also provide evidence that beta1 integrins are required for the villi innervation in the small intestine. Our findings highlight the crucial roles played by beta1 integrins at various steps of enteric nervous system development.


Subject(s)
Enteric Nervous System/embryology , Integrin beta1/genetics , Neural Crest/cytology , Neural Crest/embryology , Phenotype , Animals , Disease Models, Animal , Enteric Nervous System/cytology , Enteric Nervous System/physiology , Hirschsprung Disease/embryology , Immunohistochemistry , Integrases , Mice , Models, Genetic , Mutation , Organ Culture Techniques , Viral Proteins
6.
Blood ; 102(9): 3371-8, 2003 Nov 01.
Article in English | MEDLINE | ID: mdl-12869515

ABSTRACT

Retinoic acid induces clinical remission in acute promyelocytic leukemia (APL) by triggering differentiation of leukemia promyelocytes. Here, we have characterized a gene encoding a member of the immunoglobulin superfamily, among novel retinoic acid-induced genes identified in APL cells. This protein, which was named JAML (junctional adhesion molecule-like), contains 2 extracellular immunoglobulin-like domains, a transmembrane segment, and a cytoplasmic tail. JAML mRNA is expressed in hematopoietic tissues and is prominently expressed in granulocytes. The fact that JAML protein is localized at the cell plasma membrane in the areas of cell-cell contacts, whereas it is not detected at free cell borders, suggests that JAML is engaged in homophilic interactions. Furthermore, a conserved dimerization motif among JAM members was shown to be important for JAML localization at the cell membrane. Finally, exogenous expression of JAML in myeloid leukemia cells resulted in enhanced cell adhesion to endothelial cells. Altogether, our results point to JAML as a novel member of the JAM family expressed on leukocytes with a possible role in leukocyte transmigration.


Subject(s)
Cell Adhesion Molecules/biosynthesis , Gene Expression Regulation/drug effects , Leukemia, Myeloid/pathology , Base Sequence , Cell Adhesion/drug effects , Cell Adhesion Molecules/genetics , Cell Differentiation/genetics , Cell Line, Tumor , Endothelium, Vascular/cytology , Humans , Junctional Adhesion Molecules , Leukemia, Myeloid/metabolism , Leukocytes/cytology , Leukocytes/metabolism , Molecular Sequence Data , RNA, Messenger/biosynthesis , Sequence Analysis , Tretinoin/pharmacology , Tumor Cells, Cultured
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