Figure 6. The rapamycin-induced turnover of Hxt7 signals through Tor1, but is Npr1-independent.

(A) BY4742 and tor1-1 expressing CUP1pro-GFP-HXT7 were cultured in raffinose media plus CuSO₄ as described in the methods (time 0). After harvesting and washing, the cells were resuspended in raffinose media devoid of CuSO₄ and treated with rapamycin. Samples were collected at the indicated times and analyzed by (A) fluorescence microscopy and (B) Western analysis with anti-GFP antibodies. Identical blots were also probed with anti-ADH antibody as a loading control. BY4742 and npr1Δ strains expressing CUP1pro-GFP-HXT7 were cultured and treated as outlined above. Samples were collected at the indicated times and analyzed by (C) fluorescence microscopy, and (D) Western analysis with anti-GFP antibodies. Identical blots were also probed with anti-ADH antibody as a loading control.