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. Author manuscript; available in PMC: 2013 Aug 10.
Published in final edited form as: Mol Cell. 2012 Jun 14;47(3):410–421. doi: 10.1016/j.molcel.2012.05.025

Figure 3. AH2 interacts with PCNA via a conserved PIP box.

Figure 3

(A) AH2 contains a PIP box that is highly conserved among some other PCNA interacting proteins. The consensus sequence of PIP box is indicated. (B) The PIP box of AH2 is required for AH2 to bind to PCNA. 293T cells were transfected with plasmids encoding SFB-tagged wild-type AH2 or three PIP box mutants (dPIPB, Q519A and F525A) of AH2. Co-precipitation was carried out using S-protein beads and immunoblotting was performed using antibodies as indicated. (C) AH2-depleted (AH2 shRNA#55) HeLa derivative cell lines stably expressing shRNA-resistant wild-type AH2 or three PIP box mutants (dPIPB, Q519A and F525A) of AH2 were generated. The empty vector was included as control. The endogenous and exogenous AH2 expression was confirmed by immunoblotting using indicated antibodies. (D) PIP box mutants of AH2 could not rescue HU hypersensitivity of AH2-depleted cells. Survival curves are shown for indicated cell lines in response to increasing doses of HU. Data are presented as mean ± s.d. from three different experiments. See also Figure S2.