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Identification of novel regulators of developmental hematopoiesis using Endoglin regulatory elements as molecular probes.

Accepted version
Peer-reviewed

Type

Article

Change log

Authors

Nasrallah, Rabab 
Fast, Eva M 
Solaimani, Parham 
Knezevic, Kathy 
Eliades, Alexia 

Abstract

Enhancers are the primary determinants of cell identity, and specific promoter/enhancer combinations of Endoglin (ENG) have been shown to target blood and endothelium in the embryo. Here, we generated a series of embryonic stem cell lines, each targeted with reporter constructs driven by specific promoter/enhancer combinations of ENG, to evaluate their discriminative potential and value as molecular probes of the corresponding transcriptome. The Eng promoter (P) in combination with the -8/+7/+9-kb enhancers, targeted cells in FLK1 mesoderm that were enriched for blast colony forming potential, whereas the P/-8-kb enhancer targeted TIE2+/c-KIT+/CD41- endothelial cells that were enriched for hematopoietic potential. These fractions were isolated using reporter expression and their transcriptomes profiled by RNA-seq. There was high concordance between our signatures and those from embryos with defects at corresponding stages of hematopoiesis. Of the 6 genes that were upregulated in both hemogenic mesoderm and hemogenic endothelial fractions targeted by the reporters, LRP2, a multiligand receptor, was the only gene that had not previously been associated with hematopoiesis. We show that LRP2 is indeed involved in definitive hematopoiesis and by doing so validate the use of reporter gene-coupled enhancers as probes to gain insights into transcriptional changes that facilitate cell fate transitions.

Description

Keywords

Animals, Cell Line, Embryo, Mammalian, Endoglin, Endothelial Cells, Enhancer Elements, Genetic, Hematopoiesis, Low Density Lipoprotein Receptor-Related Protein-2, Mesoderm, Mice, Molecular Probes, Mouse Embryonic Stem Cells

Journal Title

Blood

Conference Name

Journal ISSN

0006-4971
1528-0020

Volume Title

Publisher

American Society of Hematology
Sponsorship
Wellcome Trust (097922/Z/11/Z)
Medical Research Council (MC_PC_12009)
Leukemia & Lymphoma Society (7001-12)
Cancer Research Uk (None)
Biotechnology and Biological Sciences Research Council (BB/I00050X/1)
National Health and Medical Research Council of Australia, Australian Research Council, Dr Tom Bee Stem Cell Research Fund, Cancer Research UK, Biotechnology and Biological Sciences Research Council, Leukaemia and Lymphoma Research, The Leukaemia and Lymphoma Society, core support grants by the Wellcome Trust to the Cambridge Institute for Medical Research and Wellcome Trust - MRC Cambridge Stem Cell Institute (Grant IDs: R01 HL04880, P015PO1HL32262-32, 5P30 DK49216, 5R01 DK53298, 5U01 HL10001-05, R24 DK092760)