Identification of Cdca7 as a novel Notch transcriptional target involved in hematopoietic stem cell emergence.
Hematopoietic stem cell (HSC) specification occurs in the embryonic aorta and requires Notch activation; however, most of the Notch-regulated elements controlling de novo HSC generation are still unknown. Here, we identify putative direct Notch targets in the aorta-gonad-mesonephros (AGM) embryonic tissue by chromatin precipitation using antibodies against the Notch partner RBPj. By ChIP-on-chip analysis of the precipitated DNA, we identified 701 promoter regions that were candidates to be regulated by Notch in the AGM. One of the most enriched regions corresponded to the Cdca7 gene, which was subsequently confirmed to recruit the RBPj factor but also Notch1 in AGM cells. We found that during embryonic hematopoietic development, expression of Cdca7 is restricted to the hematopoietic clusters of the aorta, and it is strongly up-regulated in the hemogenic population during human embryonic stem cell hematopoietic differentiation in a Notch-dependent manner. Down-regulation of Cdca7 mRNA in cultured AGM cells significantly induces hematopoietic differentiation and loss of the progenitor population. Finally, using loss-of-function experiments in zebrafish, we demonstrate that CDCA7 contributes to HSC emergence in vivo during embryonic development. Thus, our study identifies Cdca7 as an evolutionary conserved Notch target involved in HSC emergence.
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Jordi
J
Guiu
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Dylan J M
DJ
Bergen
Erasmus MC Stem Cell and Regenerative Medicine Institute, Erasmus Medical Center, 3000 CA Rotterdam, Netherlands.
Emma
E
De Pater
Research Unit on Biomedical Informatics, Department of Experimental and Health Sciences, Pompeu Fabra University, 08003 Barcelona, Spain Department of Genetic Engineering and Biotechnology, University of Dhaka, Dhaka 1000, Bangladesh.
Abul B M M K
AB
Islam
Centre for Genomics and Oncological Research (Genyo), Pfizer-University of Granada-Andalusian Government, 18016 Granada, Spain.
Verónica
V
Ayllón
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Leonor
L
Gama-Norton
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Cristina
C
Ruiz-Herguido
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Jessica
J
González
Research Unit on Biomedical Informatics, Department of Experimental and Health Sciences, Pompeu Fabra University, 08003 Barcelona, Spain Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain.
Nuria
N
López-Bigas
José Carreras Leukaemia Research Institute, Cell Therapy Program, School of Medicine, University of Barcelona, 08036 Barcelona, Spain Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain.
Pablo
P
Menendez
Erasmus MC Stem Cell and Regenerative Medicine Institute, Erasmus Medical Center, 3000 CA Rotterdam, Netherlands.
Elaine
E
Dzierzak
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain.
Lluis
L
Espinosa
Program de Recerca en Càncer, Institut Hospital del Mar d'Investigacions Mèdiques, Parc de Recerca Biomèdica de Barcelona, 08003 Barcelona, Spain abigas@imim.es.
Anna
A
Bigas
D000818:Animals; D001011:Aorta / Q000196:embryology / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000196:embryology / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D057809:HEK293 Cells; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein / Q000235:genetics / Q000378:metabolism; D017403:In Situ Hybridization; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D051881:Receptor, Notch1 / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000072099:Serrate-Jagged Proteins; D014158:Transcription, Genetic; D015027:Zebrafish / Q000196:embryology / Q000235:genetics
C445325:CDCA7 protein, human; D002135:Calcium-Binding Proteins; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; C497571:Rbpj protein, mouse; D051881:Receptor, Notch1; D000072099:Serrate-Jagged Proteins