Fig. 6. Structure-function analysis of Drm-Lin interaction in vitro and in vivo. (A) Drm constructs. The full-length, wild-type Drm construct contains two zinc fingers (ZF1 and ZF2). The N-Drm construct consists of the 25-residue N-terminal portion of Drm, and the C-Drm construct contains the C-terminal 63-residue domain, including the two zinc fingers. R46C is the strong drm⁶ missense mutation within ZF1, and C57G is a missense mutation in one of the conserved zinc-binding Cys residues of ZF2. The ability of each derivative to bind Lin in vitro was determined by coimmunoprecipitation of epitope-tagged Drm and Lin from Schneider S2 cells (B). Lin binds with high affinity to full-length Drm and C-Drm, with lower affinity to C57G and C-Drm(C57G), and does not bind to N-Drm, R46C, or C-Drm(R46C). Control samples of cell lysates show that Lin was expressed in all transfection assays. To assess the effect of these constructs in vivo, byn-GAL4 was used to drive expression of each throughout the hindgut, and the phenotype characterized by anti-Crb staining (C). Expression of N-Drm results in a wild-type appearing hindgut, while expression of C-Drm produces a lin-like phenotype similar to the gain-of-function phenotype produced by ectopic expression of full-length Drm (compare Fig. 5A). Expression of R46C results in a wild-type hindgut while C57G produces a lin-like phenotype. +++ or +, strong or weak interaction, respectively; –, no interaction.