wg protein was purified from S2 cell conditioned medium using Cibacron blue sepharose affinity chromatography and size exclusion chromatography.

Mass spectrometry data generated from purified wg protein indicated the presence of Swim peptides.

Bait expressed and purified from stably transfected cells.

Prey endogenous to cell extract.

Purified Swim protein was bound to plate, exposed to purified wg protein, and binding was detected using wg primary and alkaline phosphatase-conjugated secondary antibodies.

Binding affinity of wg-Swim interaction: 1.7 nM.

In the absence of detergent, purified Swim maintains the solubility of purified wg.

Palimitic acid disrupts the wg-Swim interaction, presumably by competing with wg lipid modifications for Swim binding. Consistent with this interpretation, Swim stimulates signaling by purified palmitoyl-modified wg, but does not stimulate the activity of an unmodified C93A mutant wg protein.

Bait expressed and purified from stably transfected cells.

Prey expressed and purified from stably transfected cells.

Swim-fz fusion expressed in cells and mixed with wg-containing conditioned medium; binding to cell surface detected by immunofluorescence using wg antibody.

Bait expressed in transiently transfected cells.

Prey expressed in stably transfected cells.