Extended data Figure 6. Nras^G12D/+ had a bimodal effect on HSC cycling but increased the rate at which MPPs divide.

a) Flow cytometric analysis of GFP expression in whole bone marrow cells from Nras^G12D/+ or littermate control mice after 12 weeks of chase without doxycycline. b) Median GFP fluorescence intensity of H2B-GFP⁻, H2B-GFP^lo and H2B-GFP^hi HSCs from wild type and Nras^G12D/+ mice (n=8 mice/genotype). GFP levels in control HSCs were set to one for comparison to relative levels in Nras^G12D/+ HSCs. c) Nras^G12D increased the rate of division by MPPs. Flow cytometric analysis of GFP expression in CD150⁻CD48⁻LSK MPPs from Mx1-cre; Nras^G12D/+; Col1A1-H2B-GFP; Rosa26-M2-rtTA mice (G12D/+) and littermate controls (con) after 12 weeks of chase(n=8 mice/genotype). Relative to control MPPs, Nras^G12D/+ MPPs included significantly more H2B-GFP⁻ frequently cycling cells and significantly fewer H2B-GFP^lo MPPs (p<0.05 by two-way ANOVA and posthoc pairwise t-tests). d) We continuously administered BrdU to Mx1-cre; Nras^G12D/+ versus control mice for 1 to 30 days and determined the frequency of BrdU+ HSCs (1 day BrdU data are from Figure 1a). Data represent mean±s.d.. Two-tailed student's t-tests were used to assess statistical significance unless stated otherwise. *P<0.05, **P<0.01, ***P<0.001.