Figure 3. The GATOR2 complex regulates the recruitment of TSC to lysosomes via the Rag GTPase.

(A) Time point images from the Halo-TSC2 FRAP experiment in WT HeLa cells treated with Non-targeting, RAGA and RAGC siRNAs. Scale bar: 0.5 μm.

(B) Time point images from the Halo-TSC2 FRAP experiment in WDR24-KO HeLa cells treated with Non-targeting, RAGA and RAGC siRNAs.

(C) Fluorescence recovery versus time curves in A and B. A total of 30 lysosomes in different cells were used to plot the curve for each treatment. Error bars represent standard error.

(D) Plot showing the relationship between the recovery fraction versus half time (t_1/2) from the curves in C. Error bars represent standard error.

(E) Time point pictures from the GFP-TSC1 FRAP experiments in WT and ragA^RNAi Drosophila ovaries. Scale bar: 2 μm.

(F) Time point images from the GFP-TSC1 FRAP experiments in wdr24^RNAi and wdr24^RNAi ragA^RNAi Drosophila ovaries.

(G) Fluorescence recovery versus time curves in H and I. A total of 10 lysosomes from different ovaries were examined. Error bars represent standard error.

(H) Knockdowns of RAGA or RAGC increase the interaction between mTOR and Rheb in WDR24-KO cells. Cell lysates were immunoprecipitated by an anti-mTOR antibody and immunoblotted by an anti-Rheb antibody.

(I) Knockdowns of RAGA or RAGC decrease the interaction between TSC2 and Rheb in WDR24-KO HeLa cells. Cell lysates were immunoprecipitated by an anti-TSC2 antibody and immunoblotted by an anti-Rheb antibody.