American Society of Hematology

Figure 6.

$Figure 6. Flt3-ITD enhances SMRT-p65 interaction through the SMRT-RID. (A) Subcellular localization of p65 in the absence (vec) or presence of Flt3-ITD. HA-p65 was transfected into 293T cells, which were subjected to biochemical fractionation. Equal proportions of the resulting lysates were immunoblotted with anti-HA antibody to detect p65. (B) Importance of RID for p65-SMRT interaction and enhancement by Flt3-ITD. Cells were transfected with HA-p65 and GFP-SMRT (1-1495) or GFPSMRT (1-1073) in the presence (F) or absence (vec; V) of Flt3-ITD. Cytosolic extracts from the transfected cells were coimmunoprecipitated with anti-SMRT antibody and immunoblotted with anti-HA antibody to detect p65. The immunoblots were quantified by densitometry and the relative density was calculated. (C) Overexpression of ssIκBα inhibits the effect of Flt3-ITD. Cells transfected with the full-length GFP-SMRT (1-1495) constructs in the presence or absence of coexpressed Flt3-ITD and ssIκBα were visualized by immunofluorescence microscopy. The percentage of cells exhibiting nuclear (N) or cytoplasmic (C) SMRT expression was scored in 100 cells. Data are representative of 3 experiments.$

Flt3-ITD enhances SMRT-p65 interaction through the SMRT-RID. (A) Subcellular localization of p65 in the absence (vec) or presence of Flt3-ITD. HA-p65 was transfected into 293T cells, which were subjected to biochemical fractionation. Equal proportions of the resulting lysates were immunoblotted with anti-HA antibody to detect p65. (B) Importance of RID for p65-SMRT interaction and enhancement by Flt3-ITD. Cells were transfected with HA-p65 and GFP-SMRT (1-1495) or GFPSMRT (1-1073) in the presence (F) or absence (vec; V) of Flt3-ITD. Cytosolic extracts from the transfected cells were coimmunoprecipitated with anti-SMRT antibody and immunoblotted with anti-HA antibody to detect p65. The immunoblots were quantified by densitometry and the relative density was calculated. (C) Overexpression of ssIκBα inhibits the effect of Flt3-ITD. Cells transfected with the full-length GFP-SMRT (1-1495) constructs in the presence or absence of coexpressed Flt3-ITD and ssIκBα were visualized by immunofluorescence microscopy. The percentage of cells exhibiting nuclear (N) or cytoplasmic (C) SMRT expression was scored in 100 cells. Data are representative of 3 experiments.

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