Previous studies have suggested that embryonic vascular endothelial, endocardial and myocardial lineages originate from multipotential cardiovascular progenitors. However, their existence in vivo has been debated and molecular mechanisms that regulate specification of different cardiovascular lineages are poorly understood. An ETS domain transcription factor Etv2/Etsrp/ER71 has been recently established as a crucial regulator of vascular endothelial differentiation in zebrafish and mouse embryos. In this study, we show that etsrp-expressing vascular endothelial/endocardial progenitors differentiate as cardiomyocytes in the absence of Etsrp function during zebrafish embryonic development. Expression of multiple endocardial specific markers is absent or greatly reduced in Etsrp knockdown or mutant embryos. We show that Etsrp regulates endocardial differentiation by directly inducing endocardial nfatc1 expression. In addition, Etsrp function is required to inhibit myocardial differentiation. In the absence of Etsrp function, etsrp-expressing endothelial and endocardial progenitors initiate myocardial marker hand2 and cmlc2 expression. Furthermore, Foxc1a function and interaction between Foxc1a and Etsrp is required to initiate endocardial development, but is dispensable for the inhibition of myocardial differentiation. These results argue that Etsrp initiates endothelial and endocardial, and inhibits myocardial, differentiation by two distinct mechanisms. Our findings are important for the understanding of genetic pathways that control cardiovascular differentiation during normal vertebrate development and will also greatly contribute to the stem cell research aimed at regenerating heart tissues.

Vascular endothelial and myeloid cells have been proposed to originate from a common precursor cell, the hemangioblast. The mechanism of endothelial and myeloid cell specification and differentiation is poorly understood. We have previously described the endothelial-specific zebrafish Ets1-related protein (Etsrp), which was both necessary and sufficient to initiate vasculogenesis in the zebrafish embryos. Here we identify human Etv2/ER71 and mouse ER71 proteins as functional orthologs of Etsrp. Overexpression of mouse ER71 and Etsrp caused strong expansion of hemangioblast and vascular endothelial lineages in a zebrafish embryo. In addition, we show that etsrp is also required for the formation of myeloid but not erythroid cells. In the absence of etsrp function, the number of granulocytes and macrophages is greatly reduced. Etsrp overexpression causes expansion of both myeloid and vascular endothelial lineages. Analysis of mosaic embryos indicates that etsrp functions cell autonomously in inducing myeloid lineage. We further demonstrate that the choice of endothelial versus myeloid fate depends on a combinatorial effect of etsrp, scl, and alk8 genes.

Background: Signaling by the Wnt family of secreted glycoproteins through their receptors, the frizzled (Fz) family of seven-pass transmembrane proteins, is critical for numerous cell fate and tissue polarity decisions during development.Results: We report a novel role of Wnt signaling in organogenesis using the formation of the islet during pancreatic development as a model tissue. We used the advantages of the zebrafish to visualize and document this process in living embryos and demonstrated thatinsulin-positive cells actively migrate to form an islet. We used morpholinos (MOs), sequence-specific translational inhibitors, and time-lapse imaging analysis to show that the Wnt-5 ligand and the Fz-2 receptor are required for properinsulin-cell migration in zebrafish. Histological analyses of islets inWnt5a^-/-mouse embryos showed that Wnt5a signaling is also critical for murine pancreaticinsulin-cell migration.Conclusion: Our results implicate a conserved role of a Wnt5/Fz2 signaling pathway in islet formation during pancreatic development. This study opens the door for further investigation into a role of Wnt signaling in vertebrate organ development and disease.

Signaling pathways controlling vasculogenesis, angiogenesis, and myelopoiesis are still poorly understood, in part because not all genes important for vasculature or myeloid cell formation have been characterized. To identify novel potential regulators of vasculature and myeloid cell formation we performed microarray analysis of zebrafish embryos that overexpress Ets1‐related protein (Etsrp/Etv2/ER71), sufficient to induce vasculogenesis and myelopoiesis (Sumanas and Lin [2006] Development 121:3141–3150; Lee [2008] Cell Stem Cell 2:497–507; Sumanas et al. [2008] Blood 111:4500–4510). We performed sequence homology and expression analysis for up‐regulated genes that were novel or previously unassociated with the zebrafish vasculature formation. Angiotensin II type 2 receptor (agtr2), src homology 2 domain containing E (she), mannose receptor C1 (mrc1), endothelial cell‐specific adhesion molecule (esam), yes‐related kinase (yrk/fyn), zinc finger protein, multitype 2b (zfpm2b/fog2b), and stabilin 2 (stab2) were specifically expressed in vascular endothelial cells during early development while keratin18 expression was localized to the myeloid cells. Identification of vasculature and myeloid‐specific genes will be important for dissecting molecular mechanisms of vasculogenesis/angiogenesis and myelopoiesis. Developmental Dynamics 238:1836–1850, 2009.