ABSTRACT
The extracellular membrane-associated protein F-spondin has been implicated in cell-matrix and cell-cell adhesion and plays an important role in axonal pathfinding. We report here that F-spondin is expressed in non-neuronal cells in the embryonic chicken ciliary ganglion (CG) and robustly promotes survival of cultured CG neurons. Using deletion constructs of F-spondin we found that the amino-terminal Reelin/Spondin domain cooperates with thrombospondin type 1 repeat (TSR) 6, a functional TGFß-activation domain. In ovo treatment with blocking antibodies raised against the Reelin/Spondin domain or the TSR-domains caused increased apoptosis of CG neurons during the phase of programmed cell death and loss of about 30% of the neurons compared to controls. The Reelin/Spondin domain receptor - APP and its downstream signalling molecule disabled-1 are expressed in CG neurons. F-spondin induced rapid phosphorylation of disabled-1. Moreover, both blocking the central APP domain and interference with disabled-1 signalling disrupted the survival promoting effect of F-spondin. Taken together, our data suggest that F-spondin can promote neuron survival by a mechanism involving the Reelin/Spondin and the TSR domains.
Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Extracellular Matrix Proteins/metabolism , Ganglia, Parasympathetic/metabolism , Nerve Tissue Proteins/metabolism , Neurogenesis/physiology , Neurons/metabolism , Animals , Apoptosis/physiology , Cell Survival , Chick Embryo , Ganglia, Parasympathetic/embryology , Humans , Immunoblotting , Immunohistochemistry , In Situ Hybridization , In Situ Nick-End Labeling , Neurons/cytology , Reelin Protein , TransfectionABSTRACT
Two secreted signaling molecules, Xwnt-8 and BMP-4, play an essential role in the dorso-ventral patterning of the mesoderm in Xenopus. Here we investigate how the Wnt-8 and the BMP-4 pathways are connected and how they regulate target genes in the lateral and ventral marginal zone. BMP-4 regulates the transcription of Xwnt-8 in a threshold dependent manner. High levels of BMP-4 induce the expression of the Wnt antagonist sizzled in the ventral marginal zone, independent of Xwnt-8 signaling. Xwnt-8 induces the early muscle marker myf-5 in the lateral marginal zone in a BMP independent manner. The expression of the homeobox gene Xvent-1 can be modulated through both the BMP-4 and the Xwnt-8 pathways. The spatial distribution and the level of BMP-4 activity in the lateral and ventral marginal zone is reflected in the dynamic expression pattern of Xwnt-8. The data support the view that Xwnt-8 is involved in the specification of lateral (somitogenic) mesoderm and BMP-4 in the specification of ventral mesoderm.
Subject(s)
Bone Morphogenetic Proteins/metabolism , DNA-Binding Proteins , Gene Expression Regulation, Developmental , Mesoderm/physiology , Proto-Oncogene Proteins/metabolism , Trans-Activators , Xenopus Proteins , Zebrafish Proteins , Animals , Body Patterning , Bone Morphogenetic Protein 4 , Gastrula/metabolism , Gene Expression Regulation, Developmental/drug effects , In Situ Hybridization , Lithium Chloride/pharmacology , Mesoderm/drug effects , Muscle Proteins/metabolism , Myogenic Regulatory Factor 5 , Proteins/metabolism , RNA/metabolism , Signal Transduction , Time Factors , Transcription, Genetic , Wnt Proteins , XenopusABSTRACT
BMP-4 is believed to play a central role in the patterning of the mesoderm by providing a strong ventral signal. As part of this ventral patterning signal, BMP-4 has to activate a number of transcription factors to fulfill this role. Among the transcription factors regulated by BMP-4 are the Xvent and the GATA genes. A novel homeobox gene has been isolated termed Xvex-1 which represents a new class of homeobox genes. Transcription of Xvex-1 initiates soon after the midblastula transition. Xvex-1 transcripts undergo spatial restriction from the onset of gastrulation to the ventral marginal zone, and the transcripts will remain in this localization including at the tailbud stage in the proctodeum. Expression of Xvex-1 during gastrula stages requires normal BMP-4 activity as evidenced from the injection of BMP-4, Smad1, Smad5 and Smad6 mRNA and antisense BMP-4 RNA. Xvex-1 overexpression ventralizes the Xenopus embryo in a dose dependent manner. Partial loss of Xvex-1 activity induced by antisense RNA injection results in the dorsalization of embryos and the induction of secondary axis formation. Xvex-1 can rescue the effects of overexpressing the dominant negative BMP receptor. These results place Xvex-1 downstream of BMP-4 during gastrulation and suggest that it represents a novel homeobox family in Xenopus which is part of the ventral signaling pathway.
Subject(s)
Bone Morphogenetic Proteins/metabolism , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Receptors, Growth Factor , Repressor Proteins , Signal Transduction , Transcription Factors , Xenopus Proteins , Xenopus/embryology , Amino Acid Sequence , Animals , Base Sequence , Bone Morphogenetic Protein 4 , Bone Morphogenetic Protein Receptors , Bone Morphogenetic Proteins/genetics , Embryo, Nonmammalian/metabolism , Gastrula/metabolism , Gene Expression Regulation, Developmental , Goosecoid Protein , Mesoderm/physiology , Molecular Sequence Data , RNA, Antisense , Receptors, Cell Surface/genetics , Receptors, Cell Surface/metabolism , Sequence Homology, Amino AcidABSTRACT
The caudal genes in vertebrates as in invertebrates assume a posterior position along the anterior-posterior axis and they appear to regulate the expression of the Hox genes. The third chicken caudal gene, Cdx-C, was cloned. Extensive comparisons of the sequence of this protein to the other known members of this homeobox family has lead to the suggestion that vertebrate genomes contain three members of the caudal homeobox family. A comparative study of the chicken Cdx-A and Cdx-C genes during gastrulation and neurulation revealed the differences between the genes. The caudal genes exhibit sequential activation in the newly formed neural plate and sequential extinction in axial midline structures during the primitive streak regression along the anterior-posterior axis. This pattern of expression suggests that the number and identity of caudal genes expressed along the anterior-posterior axis changes dynamically.
Subject(s)
Gene Expression Regulation, Developmental , Genes, Homeobox , Homeodomain Proteins/genetics , Amino Acid Sequence , Animals , Base Sequence , Chick Embryo , Cloning, Molecular , DNA Primers/genetics , DNA, Complementary/genetics , Drosophila Proteins , Molecular Sequence Data , Multigene Family , Polymerase Chain Reaction , Sequence Homology, Amino Acid , Transcription FactorsABSTRACT
Specific signaling molecules play a pivotal role in the induction and specification of tissues during early vertebrate embryogenesis. BMP-4 specifies ventral mesoderm differentiation and inhibits neural induction in Xenopus, whereas three molecules secreted from the organizer, noggin, follistatin and chordin dorsalize mesoderm and promote neural induction. Here we report that follistatin antagonizes the activities of BMP-4 in frog embryos and mouse teratocarcinoma cells. In Xenopus embryos follistatin blocks the ventralizing effect of BMP-4. In mouse P19 cells follistatin promotes neural differentiation. BMP-4 antagonizes the action of follistatin and prevents neural differentiation. In addition we show that the follistatin and BMP-4 proteins can interact directly in vitro. These data provide evidence that follistatin might play a role in modulating BMP-4 activity in vivo.