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. 2009 May 6;284(27):18184–18193. doi: 10.1074/jbc.M109.007096

FIGURE 1.

FIGURE 1.

KLF5 interacts with other transcriptional regulators of p15. A and B, interaction of KLF5 with Smads2–4 as detected by co-IP and immunoblotting (IB). Lysates from HaCaT cells treated with TGFβ (5 ng/ml for 1 h) were subjected to IP with antibodies against Smad4 or Smad2/3 followed by IB with antibodies against KLF5 and Smads. IgG or serum was used as a negative control. Inputs are from cell lysates before IP. C, HaCaT cells were transfected with FLAG-tagged Smad3 (FLAG-Smad3), and IP and IB were performed with FLAG and KLF5 antibodies. D, pulldown of in vitro translated full length KLF5 (F) but not C terminally truncated KLF5 (ΔKLF5 or Δ) by GST-fused Smad4. GST alone served as a negative control. Inputs are translated proteins not subjected to GST pulldown. E, interaction of KLF5 (HA-tagged) with Miz-1 (FLAG-tagged) in the presence or absence of TGFβ signaling (TβRI) also detected by IP combined with IB in the HEK-293 cell line transfected with the respective plasmids. F, subcellular localization of KLF5, Smad2, and Smad4 in HaCaT cells. Nuclear and cytoplasmic fractions of HaCaT cells treated with or without TGFβ were subjected to IB with the indicated antibodies. G, TGFβ-dependent interaction between KLF5 and Myc as detected by IP with anti-Myc antibody and IB with anti-KLF5 antibody in HaCaT cells treated with or without TGFβ (5 ng/ml for 1 h).