Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced and labeled by in vitro translation.

Caf1 directly interacts with E(z) and Rpd3, but not with esc, in this assay.

Source was embryonic nuclear extract wild-type fly line; proteins produced from endogenous genes.

The complex of Su(z)12, E(z), Rpd3, esc and Caf1 has an apparent size of 600-650 kDa (gel filtration).

A histone methyltransferase activity was purified from embryonic nuclear extracts.

Purified proteins visualized by Coomassie staining.

The E(z), Caf1, Su(z)12 and esc proteins interact as part of the tetrameric Polycomb repressive complex 2 (PRC2) complex.

Analysis of direct interactions suggests that E(z), Caf1, Su(z)12 proteins form a core trimeric complex (all paiwise interactions detected) to which esc protein binds via direct interaction with E(z).

Source was extract of Sf9 cell line; bait produced from transfected construct; prey produced from transfected construct.

Affinity purification of the esc-E(z) complex was carried out from a mixture of esc, E(z), Su(z)12, and Caf1-55 recombinant proteins.

Interaction in vitro; bait and prey produced recombinant fusion proteins in baculovirus and insect cell system.

Source was cell extract of Sf9 cell line; bait produced from baculovirus construct; prey produced from baculovirus construct.

PRC2 complexes were reconstituted from cells infected with baculoviruses for E(z), Su(z)12, Caf1-55 and esc or escl.

Su(z)12, E(z), Caf1 and esc were coexpressed in Sf9 cells and purified as a complex.

Su(z)12, E(z), Caf1 and esc form the PRC2 (Polycomb repressive complex 2).

Source was baculovirus-infected Sf9 cell extract; proteins produced from transfected constructs.