RACK1 regulated ERK levels post-translationally.

(A) Shown is western blot analysis of this immunoprecipitation using antibodies against Myc or ubiquitin. (B) Quantification of relative ubiquitination levels of ERK–Myc from (A), data are the mean ± SEM, n = 3. (C) Knocking down rack1 under the control of en-GAL4 induced extra wing vein material (indicated by arrows) within the posterior compartment. The expression of erk Sem served as positive control. Scale bar: 250 μm. (D) Light microscope images of eyes of 1-day-old flies of GMR-63Q ( ey-flp GMR-63Q-HA ), GMR>erk ( GMR-GAL4/+;UAS-erk/+ ), GMR-63Q;GMR>erk ( ey-flp GMR-63Q-HA;GMR-GAL4/+;UAS-erk/+ ), GMR-63Q;GMR>erk RNAi ( ey-flp GMR-63Q-HA;GMR-GAL4/+;UAS-erk JF01366 /+ ), wild-type ( GMR-GAL4/+ ), GMR>erk RNAi ( GMR-GAL4/+;UAS-erk JF01366 /+ ), GMR-63Q;GMR > rack1 RNAi ( ey-flp GMR-63Q-HA;GMR-GAL4 UAS-rack1 KK109073 /+ ), GMR-63Q;GMR>rack1 RNAi erk RNAi ( ey-flp GMR-63Q-HA;GMR-GAL4 UAS-rack1 KK109073 /+;UAS-erk JF01366 /+ ) flies. Scale bar: 50 μm. (E) Quantification of average number of rhabdomeres per ommatidium of wild type, MJD-Q78 ( GMR-GAL4/+;UAS-MJD . tr-Q78/+ ), MJD-Q78 rack1 RNAi ( GMR-GAL4 UAS-rack1 KK109073 ;UAS-MJD . tr-Q78/+ ), MJD-Q78 erk RNAi ( GMR-GAL4/+;UAS-MJD . tr-Q78/UAS-erk JF01366 ), MJD-Q78 rack1 RNAi erk RNAi ( GMR-GAL4 UAS-rack1 KK109073 ;UAS-MJD . tr-Q78/UAS-erk JF01366 ), MJD-Q78 erk OE ( GMR-GAL4/+;UAS-MJD . tr-Q78/UAS-erk ). Transmission electron microscopy images from three one-day-old flies of each genotype were used for quantification. Data are represented as mean ± SEM, and significant differences compared with MJD-Q78 were determined using one-way ANOVA with Tukey’s post hoc test (n = 3).