American Society of Hematology

Figure 3

$Figure 3. The SH2 domain mutation does not impair membrane and IL-3 receptor association of JAK2. (A) Parental Ba/F3 (Ø) and Ba/F3 cells expressing WT-JAK2 (JAK2), JAK2V617F (V617F), JAK2V617F+R426K (V617 mSH2), and JAK2V617F+L40A/Y41A (V617F mFERM) were surface-biotinylated followed by streptavidin IP. The bound (membrane) and unbound (cytosolic) fractions were subjected to immunoblotting with antibodies to Flag and phospho-JAK2. IL-3Rβ and actin were used as loading controls. (B) IL-3Rβ chain was immunoprecipitated from parental Ba/F3 (Ø) and Ba/F3 cells expressing WT-JAK2 (JAK2), JAK2V617F (V617F), JAK2V617F+R426K (V617F mSH2), JAK2V617F+L40A/Y41A (V617F mFERM), JAK2R426K (mSH2), and JAK2L40A/Y41A (mFERM). Coprecipitated JAK2 and precipitated IL-3Rβ were determined by Western blot as indicated (top panels). Western blot analysis of lysates used is shown in the 2 bottom panels.$

The SH2 domain mutation does not impair membrane and IL-3 receptor association of JAK2. (A) Parental Ba/F3 (Ø) and Ba/F3 cells expressing WT-JAK2 (JAK2), JAK2V617F (V617F), JAK2V617F+R426K (V617 mSH2), and JAK2V617F+L40A/Y41A (V617F mFERM) were surface-biotinylated followed by streptavidin IP. The bound (membrane) and unbound (cytosolic) fractions were subjected to immunoblotting with antibodies to Flag and phospho-JAK2. IL-3Rβ and actin were used as loading controls. (B) IL-3Rβ chain was immunoprecipitated from parental Ba/F3 (Ø) and Ba/F3 cells expressing WT-JAK2 (JAK2), JAK2V617F (V617F), JAK2V617F+R426K (V617F mSH2), JAK2V617F+L40A/Y41A (V617F mFERM), JAK2R426K (mSH2), and JAK2L40A/Y41A (mFERM). Coprecipitated JAK2 and precipitated IL-3Rβ were determined by Western blot as indicated (top panels). Western blot analysis of lysates used is shown in the 2 bottom panels.

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