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. 2017 Mar 27;292(20):8207–8222. doi: 10.1074/jbc.M116.768101

Figure 8.

Figure 8.

HBP1 enhances Pim-1 promoter activity in a DNA-binding-dependent manner. A, schematic diagram of wild-type HBP1 and associated mutants. B, expression of exogenous HBP1 increases Pim-1 protein level. HEK293T cells were transfected with HA-HBP1, HA-pmHBP1, or delEx7. Western blotting was performed on protein lysates using anti-HA, Pim-1, or GAPDH antibodies. C, relative activities of HBP1 and associated mutants on the native Pim-1 promoter (Luc-Pim-1) or the mutant Pim-1 promoter (Luc-ΔPim-1). HEK293T cells were cotransfected with 0.1 μg of the indicated reporters and 0.8 μg of HBP1 or mutant expression plasmids. Error bars represent S.D. *, p < 0.05; **, p < 0.01. D, HBP1 enhances Pim-1 promoter activity in a dose-dependent manner. HEK293T cells were cotransfected with 0.1 μg of Luc-Pim-1 and different doses (0–2.0 μg) of HBP1. Error bars represent S.D. *, p < 0.05; **, p < 0.01. E, the phosphorylation of HBP1 at Ser-380 is indispensable for its induction of Pim-1 protein. The protein levels of HBP1 and its associated mutants or Pim-1 were measured by Western blotting in HeLa cells transfected with vector, HBP1, or its associated mutants (S372A/S380A, S372A, or S380A) individually. F, the phosphorylation of HBP1 at Ser-380 is indispensable for its enhancement of Pim-1 promoter. HEK293T cells were cotransfected with 0.1 μg of Pim-1 reporter and 0.8 μg of HBP1 or associated mutant expression plasmids. Error bars represent S.D. *, p < 0.05; **, p < 0.01.