Interaction in vitro; proteins produced as a recombinant fusion proteins in bacterial system.

Purified cact was assayed for its ability to block the binding of purified dl protein to a [32]P labeled DNA fragment of the zen promoter, as determined by gel shift assays. cact alone did not bind to zen DNA.

Interaction in vitro; proteins produced as a recombinant fusion proteins in bacterial system.

Purified cact was assayed for its ability to block the binding of purified dl protein to a [32]P labeled DNA fragment of the zen promoter, as determined by gel shift assays. cact alone did not bind to zen DNA, or inhibit other protein:DNA interactions.

Interaction in vitro; bait produced by in vitro translation; prey produced and labeled by in vitro translation.

No single cdc10/SWI6 repeat of cact is sufficient for binding to dl protein.

From deletion analysis, it is inferred that residues 168-349 of dl are sufficient for binding to cact.

Source was embryos of wild-type fly line; bait produced from endogenous gene; prey produced from endogenous gene.

Coimmunoprecipitation of cact with dl is decreased in an activated Tl background (Tl[10b]).

Interaction in vitro; proteins derived from wild-type embryo extract.

cact and dl exist together in a 270 kDa complex.

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

The "Region I" and the NLS ("Region II") motifs that mediate binding to cact are similar between Drosophila dl and Dif proteins, but diverge from mammalian Rel proteins.

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent.

Yeast cells carried GAL1 and URA3 reporters under the control of the dl-regulated VRE regulatory element of the zen gene. These cells were transformed with dl and cact. Interaction inferred from ability of cact to inhibit dl-dependent activation of the reporters.

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent).

Two-hybrid system: yeast GAL4-BD

Endogenous dl activation domain used.

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent).

Two-hybrid system: yeast LexA-BD/B42-AD

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent).

Two-hybrid system: yeast LexA-BD/B42-AD

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey from ovary cDNA expression library).

10 out of 20 dl-interacting hits in this screen represented cact cDNAs.

Two-hybrid system: yeast LexA-BD/B42-AD

Interaction in vitro; proteins produced as a recombinant fusion proteins in bacterial system.

Purified cact was assayed for its ability to block the binding of purified dl protein to a [32]P labeled DNA fragment of the zen promoter, as determined by gel shift assays. cact alone did not bind to zen DNA.

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey was previously cloned reagent).

Two-hybrid system: yeast LexA-BD/B42-AD

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey from embryo cDNA expression library).

Two-hybrid system: yeast LexA-BD/GAL4-AD

Two-hybrid system: yeast LexA-BD/B42-AD

Source was yeast cell line; bait produced as transgenic fusion protein; prey produced as transgenic fusion protein (prey from ovarian cDNA expression library).

Positive control.

Source was embryos of wild-type fly line; bait produced from endogenous gene; prey produced from endogenous gene.

Proteins were mixed with [32]P labeled zen DNA probe. Binding to dl by cact inferred from inhibition of the zen-dl DNA-protein complex.

Positive control.

Interaction in vitro; proteins produced as a recombinant fusion protein in bacterial system.

Bait expressed and purified from bacteria.

dl homodimers tested in this assay.

Prey expressed and purified from bacteria.

cact binds the Rel homology domains of Dif and dl more strongly than that of Rel.

Purified proteins were mixed and precipitated with anti-thioredoxin (Trx tag) antibodies.

Bait expressed and purified from bacteria.

Prey expressed and purified from bacteria.

The cact ankyrin repeat domain binds the Rel homology domains of Dif and dl more strongly than the Rel Rel homology domain.

The interaction of the cact ankyrin repeat domain and the Dif Rel homology domain is not affected by the presence of 200-fold molar excess of the Rel ankyrin repeat domain.

dl coimmunoprecipitates cact, but not the cact-interacting protein CalpA.

Source was cell extract of S2 cell line; bait produced from endogenous gene; prey produced from endogenous gene.

Source was cell extract of S2 cell line; bait produced from endogenous gene; prey produced from transfected construct.

C-terminal cact fragments generated upon co-transfection with CalpA can coimmunoprecipitate with dl.

Source was cell extract of S2 cell line; bait produced from transfected construct; prey produced from transfected construct.

Positive control.