Fig 1. CycG protein interacts with Hairless protein in vitro and in vivo.

(A) Yeast two-hybrid assay for pairwise interactions between CycG and Hairless (HFL): interaction is observed irrespective of the orientation of the assay (blue coloured colonies). pEG vector expressed the DNA-binding protein fusion; pJG and VP16 vectors the DNA-activation domain fusions. Empty vectors served as negative control (mock). Of the two CycG deletion constructs 1–215 and 215–566, only the latter containing the Cyclin domains (blue) binds to HFL. (B) Left panel shows the constructs: a sketch of the full length Hairless protein with the Su(H) binding domain (SBD, orange), the Gro binding domain (GBD, purple) and the CtBP binding domain (CBD, yellow). Numbers correspond to the codons contained in the construct according to 35, 37]. Right panel: Quantification of the interaction between CycG, full length Hairless (HFL) and Hairless deletion constructs as indicated. Values represent Miller units (M.u.), determined as outlined in the Methods section. Note that the C-terminal deletion H-CX shows no interaction, whereas H-C6 shows a strongly reduced binding to CycG. (C) Co-immunoprecipitations (IP) were performed on embryonic extracts using polyclonal antisera as indicated, directed either against Hairless or CycG. Co-precipitates were detected by Western blot (WB) in both experiments indicating in vivo interaction of the two proteins. Mock control contained no primary antiserum.