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. Author manuscript; available in PMC: 2014 Apr 10.
Published in final edited form as: Mol Cell. 2012 Jan 27;45(2):233–243. doi: 10.1016/j.molcel.2011.11.031

Figure 4. The KLF4 amino-terminus is important for degradation and interaction with pVHL.

Figure 4

(A) Schematic of human KLF4 with previously identified domains as well as deletion mutants used in the present study. AD, activation domain. RD, repression domain. NLS, nuclear localization signal. DBD, DNA-binding domain. (B) Deletion of KLF4 amino acids 1–108 stabilizes KLF4. Vectors for FLAG-HA-tagged wild type (wt) or deletion mutants of KLF4 were transfected into U2OS. 2 days after transfection cells were mock or cycloheximide (CHX) treated for 6 hours. Equal protein amounts of whole cell lysates were probed for KLF4 mutant levels by immunoblot with an antibody against the hemagglutinin (HA) tag. (C) Fine mapping of the amino-terminal KLF4 region required for degradation. The indicated deletion mutants were tested like in B. (D, E) An amino-terminal region of KLF4 is required for its interaction with KLF4. Beads with GST alone or GST-fused pVHL were incubated with various in vitro translated deletion mutants of KLF4 and bound proteins were analyzed by SDS-PAGE plus autoradiography. Deletion of amino acids 41–50 that stabilize KLF4 (C) does not affect pVHL binding (E).