Constitutive activation of SHP2 in mice cooperates with ICSBP deficiency to accelerate progression to acute myeloid leukemia
J. Clin. Invest. Iwona Konieczna, et al. 118:853 doi:10.1172/JCI33742 [Go to this article.]

Figure 1
ICSBP tyrosine phosphorylation influenced target gene transcription. (A) Representation of tyrosine residues in ICSBP. The ICSBP amino acid sequence contains 12 tyrosine residues, including 2 highly conserved residues in the IRF domain (indicated by *). (B) The switch from PRDI cis element repression to activation during myelopoiesis is mediated by non–IRF domain ICSBP tyrosines. U937 cells were transfected with an artificial promoter construct containing 4 copies of the PRDI consensus (PRDITATACAT) or control (TATACAT) and a vector to express ICSBP; ICSBP with mutation of conserved IRF domain tyrosines (Y92/95F ICSBP); or all 12 tyrosines (Y-mut ICSBP). Reporter assays were performed with or without IFN-γ. The PRDI cis element was equivalently repressed by all forms of ICSBP in undifferentiated transfectants (statistically significant results are indicated by *P ≤ 0.04). Differentiation increased PRDI cis element activity in transfectants with ICSBP, Y92/95F ICSBP, or vector control. This increase was not significantly different with WT versus Y92/95F ICSBP (**P ≤ 0.04). Repression activity of Y-mut ICSBP was not significantly altered by IFN-γ. (C) WT and Y-mut ICSBP were equivalently expressed in U937 transfectants. U937 cells were stably transfected with a vector to express ICSBP or Y-mut ICSBP, or vector control. Total lysate proteins were analyzed by Western blotting for ICSBP and tubulin (as a loading control). (D) WT and Y-mut ICSBP were similarly expressed in the nucleus. U937 cells were stably transfected with a vector to express epitope-tagged WT or Y-mut ICSBP, or vector control. Nuclear proteins from these transfectants were immunoprecipitated with an anti-ICSBP antibody and Western blotting performed with an anti–epitope tag antibody.