ABSTRACT
E-twenty six variant 2 (Etv2) transcription factor participates in cardiac, vascular-endothelial and blood cell lineage specification decisions during embryonic development. Previous studies have identified genomic elements in the etv2 locus responsible for vascular endothelial cell specification. Using transgenic analysis in zebrafish, we report here an etv2 proximal promoter fragment that prevents transgene misexpression in myocardial progenitor cells. This inhibition of etv2 expression in the cardiac progenitor population is partly mediated by Scl and Nkx2.5, likely through direct binding to the etv2 promoter, and cis-regulatory elements located in the first and second introns. The results identify an etv2 cis-regulatory mechanism controlling cardiovascular fate choice implying that etv2 participates in a transcriptional network mediating developmental plasticity of endothelial progenitor cells during embryonic development.
Subject(s)
Endothelium, Vascular/embryology , Heart/embryology , Transcription, Genetic/genetics , Zebrafish Proteins/genetics , Zebrafish/embryology , Animals , Animals, Genetically Modified , Basic Helix-Loop-Helix Transcription Factors/genetics , Cell Differentiation/genetics , Cell Line , Cell Lineage , Embryonic Stem Cells , Endothelial Cells/cytology , Endothelium, Vascular/cytology , Erythrocytes/cytology , Gene Expression Regulation, Developmental , Gene Knockdown Techniques , Gene Silencing , Homeobox Protein Nkx-2.5 , Morpholinos/genetics , Promoter Regions, Genetic , Proto-Oncogene Proteins/genetics , T-Cell Acute Lymphocytic Leukemia Protein 1 , Transcription Factors/genetics , Transgenes , Zebrafish/genetics , Zebrafish Proteins/biosynthesisABSTRACT
The extracellular matrix plays a critical role in neural crest (NC) cell migration. In this study, we characterize the contribution of the novel GPI-linked matrix metalloproteinase (MMP) zebrafish mmp17b. Mmp17b is expressed post-gastrulation in the developing NC. Morpholino inactivation of mmp17b function, or chemical inhibition of MMP activity results in aberrant NC cell migration with minimal change in NC proliferation or apoptosis. Intriguingly, a GPI anchored protein with metalloproteinase inhibitor properties, Reversion-inducing-Cysteine-rich protein with Kazal motifs (RECK), which has previously been implicated in NC development, is expressed in close apposition to NC cells expressing mmp17b, raising the possibility that these two gene products interact. Consistent with this possibility, embryos silenced for mmp17b show defective development of the dorsal root ganglia (DRG), a crest-derived structure affected in RECK mutant fish sensory deprived (sdp). Taken together, this study has identified the first pair of MMP, and their putative MMP inhibitor RECK that functions together in NC cell migration.
Subject(s)
Cell Movement/genetics , Matrix Metalloproteinase 17/genetics , Matrix Metalloproteinase 17/metabolism , Neural Crest/cytology , Neural Crest/metabolism , Amino Acid Sequence , Animals , Body Patterning/genetics , Embryonic Development/genetics , Enzyme Activation , GPI-Linked Proteins/genetics , GPI-Linked Proteins/metabolism , Gene Expression Profiling , Matrix Metalloproteinase 17/chemistry , Molecular Sequence Data , Organ Specificity/genetics , Sequence Alignment , ZebrafishABSTRACT
BACKGROUND: Vasculogenesis, the de novo formation of blood vessels from precursor cells is critical for a developing embryo. However, the signals and events that dictate the formation of primary axial vessels remain poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we use ets-related protein-1 (etsrp), which is essential for vascular development, to analyze the early stages of vasculogenesis in zebrafish. We found etsrp(+) cells of the head, trunk and tail follow distinct developmental sequences. Using a combination of genetic, molecular and chemical approaches, we demonstrate that fli(+)etsrp(+) hemato-vascular progenitors (FEVPs) are proliferating at the lateral plate mesoderm (LPM). The Shh-VEGF-Notch-Hey2 signaling pathway controls the proliferation process, and experimental modulation of single components of this pathway alters etsrp(+) cell numbers at the LPM. CONCLUSIONS/SIGNIFICANCE: This study for the first time defines factors controlling proliferation, and cell numbers of pre-migratory FEVPs in zebrafish.
Subject(s)
Blood Vessels/embryology , Cell Proliferation , Hematopoietic Stem Cells/physiology , Mesoderm/cytology , Proto-Oncogene Protein c-fli-1/metabolism , Zebrafish Proteins/metabolism , Zebrafish/embryology , Animals , Blood Vessels/metabolism , Cell Movement/physiology , Embryo, Nonmammalian , Endothelium, Vascular/embryology , Endothelium, Vascular/metabolism , Endothelium, Vascular/physiology , Gene Expression Regulation, Developmental , Head/embryology , Hematopoietic Stem Cells/metabolism , Mesoderm/metabolism , Mesoderm/physiology , Models, Biological , Neovascularization, Physiologic/genetics , Neovascularization, Physiologic/physiology , Proto-Oncogene Protein c-fli-1/genetics , Proto-Oncogene Protein c-fli-1/physiology , Tail/embryology , Zebrafish/genetics , Zebrafish/metabolism , Zebrafish/physiology , Zebrafish Proteins/genetics , Zebrafish Proteins/physiologyABSTRACT
BACKGROUND: Astacins are a large family of zinc metalloproteases found in bacteria and animals. They have diverse roles ranging from digestion of food to processing of extracellular matrix components. The C. elegans genome contains an unusually large number of astacins, of which the majority have not been functionally characterized yet. RESULTS: We analyzed the expression pattern of previously uncharacterized members of the astacin family to try and obtain clues to potential functions. Prominent sites of expression for many members of this family are the hypodermis, the alimentary system and several specialized cells including sensory sheath and sockets cells, which are located at openings in the body wall. We isolated mutants affecting representative members of the various subfamilies. Mutants in nas-5, nas-21 and nas-39 (the BMP-1/Tolloid homologue) are viable and have no apparent phenotypic defects. Mutants in nas-6 and nas-6; nas-7 double mutants are slow growing and have defects in the grinder of the pharynx, a cuticular structure important for food processing. CONCLUSIONS: Expression data and phenotypic characterization of selected family members suggest a diversity of functions for members of the astacin family in nematodes. In part this might be due to extracellular structures unique to nematodes.
