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First published online 2 February 2005
doi: 10.1242/dev.01660

Development 132, 1117-1126 (2005)
Published by The Company of Biologists 2005
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RBPj{kappa}-dependent Notch function regulates Gata2 and is essential for the formation of intra-embryonic hematopoietic cells

Àlex Robert-Moreno1, Lluís Espinosa1, José Luis de la Pompa2 and Anna Bigas1,*

1 Centre Oncologia Molecular, IDIBELL-Institut de Recerca Oncologica, Hospitalet, Barcelona 08907, Spain
2 Department of Immunology and Oncology, Centro Nacional de Biotecnología, CSIC. Darwin, 3. Campus de Cantoblanco, Madrid 28049, Spain

* Author for correspondence (e-mail: abigas{at}iro.es)

Accepted 22 December 2004

Definitive hematopoiesis in the mouse embryo originates from the aortic floor in the P-Sp/AGM region in close association with endothelial cells. An important role for Notch1 in the control of hematopoietic ontogeny has been recently established, although its mechanism of action is poorly understood. Here, we show detailed analysis of Notch family gene expression in the aorta endothelium between embryonic day (E) 9.5 and E10.5. Since Notch requires binding to RBPj{kappa} transcription factor to activate transcription, we analyzed the aorta of the para-aortic splanchnopleura/AGM in RBPj{kappa} mutant embryos. We found specific patterns of expression of Notch receptors, ligands and Hes genes that were lost in RBPj{kappa} mutants. Analysis of these mutants revealed the absence of hematopoietic progenitors, accompanied by the lack of expression of the hematopoietic transcription factors Aml1/Runx1, Gata2 and Scl/Tal1. We show that in wild-type embryos, a few cells lining the aorta endothelium at E9.5 simultaneously expressed Notch1 and Gata2, and demonstrate by chromatin immunoprecipitation that Notch1 specifically associated with the Gata2 promoter in E9.5 wild-type embryos and 32D myeloid cells, an interaction lost in RBPj{kappa} mutants. Consistent with a role for Notch1 in regulating Gata2, we observe increased expression of this gene in 32D cells expressing activated Notch1. Taken together, these data strongly suggest that activation of Gata2 expression by Notch1/RBPj{kappa} is a crucial event for the onset of definitive hematopoiesis in the embryo.

Key words: Notch, Mouse, Rbpsuh




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