Fev acts upstream of ERK signaling to establish the HSC fate. (A) qPCR analysis of erk1 and erk2 expression at 24 and 36 hpf in the dissected trunk region of embryos injected with control MO or fevMO showed that erk2 but not erk1 expression in the trunk region is reduced in fev morphants (mean ± SD, n = 3,*P < .05). (B) Immunofluorescence shows pERK1/2 expression in the dorsal aorta is reduced in fev morphants (n = 5) compared with the controls (n = 5). Note that the somitic expression of pERK1/2 is not affected in fev morphants. Yellow squares indicate the dorsal aorta area. (C) Western blot showing that Runx1 protein is significantly downregulated at 36 hpf in embryos injected with fevMO or erk2MO. In addition, total ERK1/2 and pERK1/2 proteins are strongly down-regulated at 36 hpf in embryos injected with fevMO or erk2MO. Fev is downregulated in fev morphants but not altered in erk2 morphants. (D) runx1 and cmyb expression in the dorsal aorta is reduced at 36 hpf in embryos injected with erk2MO. Anterior to the left, lateral view, arrows mark the AGM region.