Figure 2. Interaction of SIRT1 with the pocket region of Rb in vivo.
(A) Schematic representation of large-pocket Rb and mutations in the pocket region of Rb. Δ indicates mutations in the LXCXE Rb-binding site (residues 709, 713 and 757). (B) Whole-cell lysates from transfected C33A cells were immunoprecipitated (IP) with the anti-GAL4 antibody. Complexes were resolved by SDS/PAGE, transferred on to nitrocellulose and immunoblotted using the anti-SIRT1 antibody (top panel). The blot was stripped and reprobed with the anti-Rb antibody for the presence of immunoprecipitated Rb (middle panel). Input protein representing 10% of the cell lysate is shown in the bottom panel. (C) C33A cells were transfected with the indicated expression vectors. The experiment was performed as described in (B).