Figure 1.

Autophagy is induced upon zinc starvation. A, growth in SD or SD(−zinc). Wild-type or atg2Δ cells were pre-grown in SD to mid-log phase (A₆₀₀ =1) and then inoculated into fresh SD or SD(−zinc) at A₆₀₀ = 0.05. Growth was monitored by measuring absorbance at 600 nm. B, GFP-Atg8 localization to the PAS and transport to the vacuole. Wild-type, atg2Δ, or atg11Δ cells expressing GFP-Atg8 were cultured as in A, and GFP-Atg8 was analyzed by fluorescence microscopy. C, processing of GFP-Atg8. Cells were cultured as in B. At the indicated time points, lysates were prepared and analyzed by Western blotting. D, zinc depletion by TPEN. Wild-type or atg2Δ cells were pre-grown in SD to log phase (A₆₀₀ = 1), and then inoculated into fresh SD with or without TPEN (5 or 100 μm) at an initial A₆₀₀ of 0.05. At the indicated time points, cell density was measured. E, microscopic observation of GFP-Atg8 expressed in atg11Δ cells in the absence (−TPEN) or presence of TPEN (+100 μm) in SD. F, growth of several types of atg mutants in SD or SD(−zinc). Wild-type, atg1Δ, atg2Δ, atg11Δ, atg17Δ, atg19Δ, or pep4Δ cells were cultured as in A. G, electron microscopic analyses. To observe autophagic bodies in the vacuoles, pep4Δprb1Δ or pep4Δprb1Δatg2Δ cells were used. The cells were cultured in SD(−zinc) for 0 or 28 h and then examined by transmission electron microscopy. Scale bar, 500 nm.