Discrete Notch signaling requirements in the specification of hematopoietic stem cells by Albert D Kim, Chase H Melick, Wilson K Clements, David L Stachura,

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Discrete Notch signaling requirements in the specification of hematopoietic stem cells by Albert D Kim, Chase H Melick, Wilson K Clements, David L Stachura, Martin Distel, Daniela Panáková, Calum MacRae, Lindsey A Mork, J Gage Crump, and David Traver EMBO J. Volume ():e September 17, 2014 ©2014 by European Molecular Biology Organization

Notch3 is required for HSPC specification AWISH of notch3 viewed dorsally at 13 hpf (left) and laterally in the trunk at 19 (middle) and 24 hpf (right). Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Notch3 is dispensable for DA, but required for sclerotome ABrightfield image of a 26 hpf zebrafish. Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Specific temporal activation of Notch signaling is sufficient to rescue HSPCs in notch3 morphants A, BWISH for runx1 in 26 hpf hsp70:gal4; UAS:NICD ‐ myc uninjected or notch3 morphant transgenic embryos with heat ‐ shock induction at 14 hpf (A) or 20 hpf (B)... Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Notch signaling is required during two distinct time windows for specification of sclerotome and dorsal aorta A–DWISH for runx1 (A), foxc1b (B), efnb2a (C), and dlc (D) in 26 hpf embryos treated with DMSO vehicle (left), 4 μM γ ‐ secretase Notch inhibitor... Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Specific spatial activation of Notch signaling is sufficient to rescue HSPCs in notch3 morphants A, BWISH for runx1 in 26 hpf kdrl:gal4 (A) or phldb1:gal4 ‐ mcherry (B) crossed to UAS:NICD ‐ myc transgenic embryos either uninjected or injected with notch3 mo... Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Notch3 cooperates synergistically with dlc and dld to specify HSPCs A–FWISH for runx1, twist1b, and foxc1b at 26 hpf in uninjected (A), low ‐ dose knockdown of notch3 (B), heterozygotes for dlc mutant bea (C), low ‐ dose knockdown of dld (D), bea heterozygot... Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization

Wnt16, dlc/dld, and notch3 function in a linear pathway to specify HSPCs A–EExpression of runx1 at 26 hpf in uninjected (A) or injected with wnt16MO (B), wnt16MO with 50 pg dlc/dld mRNA (C), notch3MO with 50 pg dlc/dld mRNA (D), and wnt16MO/notch3MO with... Albert D Kim et al. EMBO J. doi: /embj ©2014 by European Molecular Biology Organization