Fig. 6. Mechanism of lateral signaling inhibition by persistent LIN-12 in the 1° lineage. All pictures are ventral views. In left panels, GFP (green), AJM-1(blue) and nuclear ß-galactosidase expressed from egl-17p::lacZ (red) are visualized in Pn.px stage hermaphrodites. In right panels, AJM-1 (teal) and egl-17p::LacZ (red) are visualized in Pn.pxx stage hermaphrodites. (A) LIN-12(EDTS)::GFP is an activated form of LIN-12 that is not downregulated. Most of the GFP is nuclear localized. A' shows magnification of GFP staining, and A" shows overlapping nuclear ß-galactosidase and GFP. (B) LIN-12(EDTS)::GFP does not affect VPC fates. The 1° fate is assessed by the presence of P6.pxx cells marked with egl-17p::LacZ and AJM-1. The 2° fate is scored as described in Fig. 5B. (C) LIN-12(extra)::TM::GFP is not downregulated, and accumulates at the apical plasma membrane. C' shows magnification of GFP staining, while C" shows that most of the GFP is enclosed inside the AJM-1 boundary (pseudocolored red). (D) LIN-12(extra)::TM::GFP does not affect the 1° fate, but inhibits lateral signaling as evidenced by loss of AJM-1-marked granddaughters of P5.p/P7.p. (E,F) Adding region 1 promotes downregulation of LIN-12(extra)::TM::GFP and allows for normal lateral signaling. (G) Adding the DTS to LIN-12(extra)::TM::GFP does not promote downregulation. G' shows magnification of GFP staining, while G" shows that most of the GFP is outside the AJM-1 boundary (pseudocolored red), consistent with basolateral accumulation. (H) LIN-12(extra)::TM::DTS::GFP does not inhibit lateral signaling. (I-L) Structure of GFP-tagged proteins expressed above, and quantification of downregulation (gray bars) and 2° fate defects (black bars).