ABSTRACT
The ETS factor ETV2 (aka ER71) is essential for the generation of the blood and vascular system, as ETV2 deficiency leads to a complete block in blood and endothelial cell formation and embryonic lethality in the mouse. However, the ETV2-mediated gene regulatory network and signaling governing hematopoietic and endothelial cell development are poorly understood. Here, we map ETV2 global binding sites and carry out in vitro differentiation of embryonic stem cells, and germ line and conditional knockout mouse studies to uncover mechanisms involved in the hemangiogenic fate commitment from mesoderm. We show that ETV2 binds to enhancers that specify hematopoietic and endothelial cell lineages. We find that the hemangiogenic progenitor population in the developing embryo can be identified as FLK1(high)PDGFRα(-). Notably, these hemangiogenic progenitors are exclusively sensitive to ETV2-dependent FLK1 signaling. Importantly, ETV2 turns on other Ets genes, thereby establishing an ETS hierarchy. Consequently, the hematopoietic and endothelial cell program initiated by ETV2 is maintained partly by other ETS factors through an ETS switching mechanism. These findings highlight the critical role that transient ETV2 expression plays in the regulation of hematopoietic and endothelial cell lineage specification and stability.
Subject(s)
Blood Cells/cytology , Blood Cells/metabolism , Cell Differentiation , Endothelial Cells/cytology , Endothelial Cells/metabolism , Transcription Factors/metabolism , Animals , Binding Sites , Cell Differentiation/genetics , Cell Lineage/genetics , Chromatin Immunoprecipitation , Female , Gene Expression Regulation, Developmental , Gene Regulatory Networks , Hematopoietic Stem Cells/cytology , Hematopoietic Stem Cells/metabolism , High-Throughput Nucleotide Sequencing , Immunophenotyping , Male , Mice , Mice, Knockout , Nucleotide Motifs , Organ Specificity/genetics , Position-Specific Scoring Matrices , Protein Binding , Receptors, Vascular Endothelial Growth Factor/metabolism , Signal Transduction , Transcription Factors/genetics , Vascular Endothelial Growth Factor Receptor-2/metabolism , Vascular Endothelial Growth Factors/metabolismABSTRACT
MHC-related protein (MR)1 is an MHC class I-related molecule encoded on chromosome 1 that is highly conserved among mammals and is more closely related to classical class I molecules than are other nonclassical class I family members. In this report, we show for the first time that both mouse and human MR1 molecules can associate with the peptide-loading complex and can be detected at low levels at the surface of transfected cells. We also report the production of recombinant human MR1 molecules in insect cells using highly supplemented media and provide evidence that the MR1 H chain can assume a folded conformation and is stoichiometrically associated with beta(2)-microglobulin, similar to class I molecules. Cumulatively, these findings demonstrate that surface expression of MR1 is possible but may be limited by a specific ligand or associated molecule.
