Tyrosine 201 is required for efficient induction of MPNs by JAK2V617F. BM from 6-week female Balb/c mice was infected with retroviruses expressing vector (control), JAK2WT, JAK2V617F, or JAK2V617F/Y201F and then transplanted into lethally irradiated Balb/c recipient mice. Peripheral blood red blood cell (A), hemoglobin (B), white blood cell (C), and neutrophil (D) counts in mice receiving JAK2V617F/Y201F-transduced BM (n = 10) were significantly reduced compared with mice receiving JAK2V617F-transduced BM (n = 10) and were comparable with those receiving vector control- or JAK2WT-transduced BM (n = 5) 8 weeks after transplantation. (E) Spleen weight. Mice receiving JAK2V617F-transduced BM developed profound splenomegaly compared with the control (vector or JAK2WT) mice. The Y201F mutation significantly inhibited the JAK2V617F-evoked increase in spleen weight (n = 5 for mice receiving vector control- or JAK2WT-transduced BM; n = 6 for mice receiving JAK2V617F- or JAK2V617F/Y201F-transduced BM). (F) Liver weight. No significant difference was observed in the liver size among all 4 groups of mice (n = 5). (G) Flow cytometric analysis of GFP expression in the BM 8 weeks after transplantation. Histogram represents the percentage of GFP⁺ population in the BM of the transplanted mice (n = 5). *P < .05 (1-way ANOVA). **P < .005 (1-way ANOVA). Data are mean ± SEM. (H) Southern blot analysis with a radioactive GFP probe demonstrating oligoclonal integration of proviral clones in the BM of the transplanted animals expressing vector (control), JAK2WT, JAK2V617F, or JAK2V617F/Y201F. DNA from Ba/F3-EpoR-JAK2V617F cells is included at right. DNA size markers are shown in kilobases at left.