csf1rb deficiency does not interfere with the formation of HSPCs but leads to HSPC reduction and multiple lineage defects at early developmental stages. (A) Budding behavior of the converted cell in the VDA in siblings (sib) and csf1rb mutants (mut). Orange line in the diagram indicates the VDA region. Upper panels show flk1-NLS-Eos+ green endothelial cells in the VDA; nine cells were converted into red after UV light conversion. Lower panels show the budding behavior of the cell in the white dashed line, enlarged at different time points in the bottom panels and indicated by white arrowheads. The time scale is shown in the left corner. (B) Quantification of the number of cells budding among converted endothelial cells over 26 h in siblings (n=11) and csf1rb mutants (n=8). (C) WISH of cmyb in siblings (n=16) and csf1rb mutants (n=16) at 30 hpf; red arrows show naïve HSPC signal in the VDA. (D) WISH of cmyb in siblings and csf1rb mutants at 2 dpf. (E) Quantification of cmyb+ HSPCs in siblings (n=12) and csf1rb mutants (n=12) in the VDA and the CHT at 2 dpf. (F) WISH of cmyb in csf1rb mutants and siblings at 3 dpf. (G) Quantification of cmyb+ HSPCs in siblings (n=40) and csf1rb mutants (n=16) in the CHT at 3 dpf. (H) WISH of runx1 in siblings and csf1rb mutants at 3 dpf. (I) Quantification of runx1+ HSPCs in siblings (n=20) and csf1rb mutants (n=12) in the CHT at 3 dpf. (J) WISH of αe1-globin in siblings and csf1rb mutants at 5 dpf. (K) Quantification of αe1-globin+ erythrocytes in siblings (n=52) and csf1rb mutants (n=22) in the CHT at 5 dpf. (L) Immunostaining of Lcp1 in the CHT at 5 dpf in siblings and csf1rb mutants. (M) Quantification of the Lcp1+ myeloid cells in siblings (n=11) and csf1rb mutant (n=10) embryos. (N) WISH of rag1 at 5 dpf in siblings and csf1rb mutants; red dashed circle indicates the position of the thymus. (O) Quantification of the rag1+ thymus area in siblings (n=59) and csf1rb mutant (n=19). Scale bars: 50 μm. Data are presented as mean±s.e.m. *P<0.05, **P<0.01, ****P<0.0001; ns, not significant.
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