IP:

1. Supplementary Figure 1. POP-1 interacts with CBP/p300 proteins in vitro and in vivo. A - - - - - - (1-438) + (282-438) - - - - - + GST-POP-1 - - - + - - - (188-281) - - - - - - + (48-187) - - - - - - + (1-47) GST - - + - - - - Input p300 (50%) - - - - - - + 35 S B - - + - GST-CBP-1 - + - - GST-p300 GST - + - - Input POP-1 (50%) - - - + 35 S C a-POP-1 a-CBP-1 IP: a-HA WB a-CBP-1 WB a-POP-1 (A) Human p300 interacts with POP-1 in vitro. 35S-methionine-labeled, in vitro translated, human p300 protein was incubated with equivalent amounts of bacterially expressed GST fused to either the full-length POP-1 (amino acids 1-438) or various truncated POP-1 proteins (spanning amino acids 1-47, 48-187, 188-281, or 282-438 of POP-1). Input lane corresponds to 50% of the amount of 35S-methionine-labeled p300 used for the GST pull-down assays. While p300 did not bind the GST moiety, it interacted efficiently with full-length POP-1. With the exeption of the N-terminal region (amino acids 1-47), all other fragments of POP-1 retained a significant amount of 35S-methionine-labeled p300 protein. (B) The converse GST pull-down experiment demonstrated that the 35S-methionine-labeled, in vitro translated, full-length POP-1 protein interacts with the human p300 and the C. elegans CBP-1 proteins in vitro. (C) Endogenous CBP-1 and POP-1 also interact in vivo. Wild-type worm embryo extracts were incubated with the a-POP-1 (Lin et al. 1995), the a-CBP-1 (Shi and Mello 1998) or the unrelated a-HA (Santa Cruz) rabbit polyclonal antibodies. The immunoprecipitates were then analyzed by Western blotting (WB) with the same a-POP-1 and a-CBP-1 antibodies used for the immunoprecipitation (IP).