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	Fig. 2. Effects of Wnt5a and Wnt5b on IL-3 and GM-CSF induced myeloid colony formation.A) Total numbers of colonies formed by IL-3 (10 ng/mL) in the absence and presence of Wnt5a (50 and 200 ng/mL) and Wnt5b (50 and 200 ng/mL). Panels B-D show differential data for B) GM-CFU; C) M-CFU and D) G-CFU. E) Total numbers of colonies formed by GM-CSF (10 ng/mL) in the absence and presence of Wnt5a and Wnt5b. Panels F- H show differential data for F) GM-CFU; G) M-CFU and H) G-CFU. Data are presented as mean ± SEM. *p < .05, n =6-8.
	Fig. 3. Similar effects of IL-3 and GM-CSF on myeloid differentiation.A) CFU assay after treatment with IL-3 (10 ng/mL) and GM-CSF (10 ng/mL). Colorsrepresent different types of myeloid colonies - GM-CFU = Granulocyte-monocyte colony formation unit (white); M-CFU = Monocyte colony formation unit (grey); G- CFU = Granulocyte colony formation unit (black). B) Log10 transformed colony-areas in μm2. C) CFU replating assay after treatment with IL-3. Colors represent the same types of colonies as described for panel A. Half-life represents calculation of the number of rounds needed to reach a 50% decay in colony formation. D) CFU replating assay after treatment with GM-CSF. Colors represent the same types of colonies as described for panel A. Half-life represents calculation of the number of rounds needed to reach a 50% decay in colony formation. E) Cells/colony after IL-3 and GM-CSF treatment. F) Cell number after 1 week of LC with IL-3 and GM-CSF treatment. G) Percentage of Lin+ cells after 1 week of liquid culture (LC) with IL-3 and GM-CSF treatment. H) Percentage of Lin+ cells after 1 week of liquid culture (LC) with IL-3 and GM-CSF treatment. I) Percentage of Lin-c-Kit+ cells after 1 week of liquid culture (LC) with IL-3 and GM-CSF treatment. J-K) Analysis of phosphorylation of GSK3β after 15, 20 and 30 min of stimulation with IL-3 or GM-CSF (respectively) in HSPC (Lin-Sca-1 +c-Kit+ — black bars) and more committed progenitors (Lin- Sca-1- c-Kit+ - stripped bars). Comparison was done between HPSC and progenitors in each treatment. Data are presented as mean ± SEM, except for area data, which is plotted as individual colonies and their median. *p < .05.
	Fig. 4. Wnt5b maintains primitive progenitors in combination with IL-3.A) CFU Replating assay in the presence of IL-3 (10 ng/mL). B) CFU Replating assay in the presence of Wnt5b (200 ng/mL) and IL-3. Bars show total colony number; pie charts represent types of colonies in each round of CFU reading, being white = GM- CFU; grey = M-CFU and black = G-CFU. C) Colony-areas. D) Cell number/colony after IL-3 and IL-3 + Wnt5b. E) CFU replating of subtypes of colonies after IL-3 and IL-3 + Wnt5b. Colors represent GM-CFU = white, M-CFU = grey; G-CFU = black. F-I) Cdk1 (F,H) and Cyclin D1 (G,I) gene expression in relation to housekeeping genes after 1 week (F,G) and 2 weeks (H,I) of CFU assay. J-M) GATA-2 (J,L) and Ifitm-1 (K,M) gene expression in relation to housekeeping genes after 1 week (J,K) and 2 weeks (L,M) of CFU assay. N-P) Axin2 (N), Dkk1 (O) and LRP5 (P) gene expression in relation to housekeeping genes after 2 weeks of CFU assay. P) LRP5 gene expression in relation to housekeeping genes after 2 weeks of CFU assay. Q-S) Gene expression of proteins involved in non-canonical Wnt signaling (c-Fos, c-Jun and Cdc42) in relation to housekeeping genes after 2 weeks of CFU assay. For gene expression, housekeeping genes used were GAPDH and β-actin. Data are presented as mean ± SEM. *p < .05.
	Fig. 5. Wnt5b induces committed progenitor activation and progenitor exhaustion in the presence of GM-CSF.A) CFU Replating assay in the presence of GM-CSF (10 ng/mL). B) CFU Replating assay in the presence of Wnt5b (200 ng/mL) and GM-CSF. Pie charts represent types of colonies in each rounds of CFU reading, being white = GM-CFU; grey = M-CFU and black = G-CFU. C) Colony-areas. D) Cell number/colony after GM-CSF and GM-CSF + Wnt5b treatment. E) CFU replating of subtypes of colonies after GM-CSF and GM-CSF + Wnt5b treatment. F) Ratio between Lin + and Lin- populations after 1 and 2 weeks of CFU assay. G-L) Cdk1 (G), Cyclin D1 (H), GATA-2 (I), Ifitm-1 (J), Axin2 (K), Dkk1 (L), LRP5 (M), c-Fos (N), c-Jun (O) and Cdc42 (P) gene expression in relation to housekeeping genes after 2 weeks of CFU assay. For gene expression, housekeeping genes used were GAPDH and β-actin. Data are presented as mean ± SEM. *p < .05.

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