Par-1 Regulates Tissue Growth by Influencing Hippo Phosphorylation Status and Hippo-Salvador Association
Figure 4
Par-1 functionally interacts with the components of the Hpo pathway.
(A–B′) Loss of Par-1 induced a phenotype that was suppressed by Yki overexpression. The protein levels of DIAP1 in the wing discs expressing UAS-Par-1 RNAi (A–A′) or coexpressing UAS-Yki and UAS-Par-1 RNAi (B–B′) with hh-Gal4 were detected. Note that Yki overexpression overcame the inhibitory effect of Par-1 RNAi on diap1 expression. The arrows indicate the P-compartment. (C) Par-1 inhibits Yki phosphorylation. S2 cells were transfected with the indicated constructs. Phosphorylated Yki was detected using the p-Yki antibody, which recognizes the phosphorylated site of Yki at Ser168. (D–E′) ex functions upstream of par-1 in the Hpo pathway. Wing discs expressing UAS-ex RNAi (D–D′) or coexpressing UAS-ex RNAi and UAS-Par-1 RNAi (E–E′) with hh-Gal4 were subjected to immunostaining. The transgene expression regions were marked by the lack of Ci (red) staining and are indicated by arrows. Note that ex RNAi expression resulted in an enlarged P-compartment and increased expression of diap1-GFP 3.5, whereas coexpression with Par-1 RNAi fully suppressed these phenotypes. (F–F′″) wts functions downstream of par-1 in the Hpo pathway. Clones were generated using the MARCM system. The genotypes were the following: ey-flp, Ubi-Gal4, UAS-GFP; FRT82B/FRT82B Gal80 (F), ey-flp, Ubi-Gal4, UAS-GFP; Par-1-RNAi; FRT82B/FRT82B Gal80 (F′), ey-flp, Ubi-Gal4, UAS-GFP; Par-1-RNAi; FRT82B wtslatsX1/FRT82B Gal80 (F″), and ey-flp, Ubi-Gal4, UAS-GFP; FRT82B wtslatsX1/FRT82B Gal80 (F′″). Note that Par-1 RNAi reduced the clone size, whereas wtslatsX1 rescued this phenotype. (G) Quantification of the relative clone size. The relative clone size was calculated as the GFP area divided by the entire disc area. All of these data were expressed as the mean ± SEM. **p<0.01. **p<0.001. n>5, for each group. (H) Par-1 modulates Wts phosphorylation status. S2 cells were transfected with the indicated plasmids. The cell lysates were harvested and followed by Western blot analysis. Note that the phosphorylation shift of Wts mediated by Hpo/Sav/Merlin/Tao-1 was partially blocked by Par-1 expression. The shifted Wts bands are indicated by the small circles. (I–I″) par-1 functions upstream of hpo in the Hpo pathway. Clones were generated using the MARCM system. The genotypes were the following: eyflp, ubi-Gal4, UAS-GFP; FRT42D/FRT42D Gal80 (I), eyflp, ubi-Gal4, UAS-GFP; FRT42D hpoBF33/FRT42DGal80 (I′), eyflp, ubi-Gal4, UAS-GFP; FRT42D hpoBF33/FRT42DGal80; Par-1-RNAi (I″). Note that the Hpo null clones caused tumorous growth, whereas Par-1 RNAi could not rescue this phenotype.