<p>A) Confirmation of the <i>ice2Δ</i> low zinc growth defect. Wild type (BY4743, <i>filled columns</i>) and <i>ice2Δ</i> (BY4743 <i>ice2Δ</i>, <i>open columns</i>) cells were inoculated into LZM supplemented with either 1 or 3 µM ZnCl<sub>2</sub> and grown overnight prior to measuring the culture optical densities at 600 nm (OD<sub>600</sub>). B) Loss of Ice2 causes a zinc-suppressible hyper-induction of the unfolded protein response (UPR). Wild type (BY4743) and homozygous <i>ice2Δ</i> mutant (BY4743 <i>ice2Δ</i>) cells were transformed with the UPRE-lacZ reporter pMCZ-Y and inoculated into low zinc medium (LZM) supplemented with 0.3, 1, 3 or 10 µM ZnCl<sub>2</sub>. These cells were then grown overnight prior to measuring β–galactosidase activity. C) Loss of Ice2 exacerbates the zinc-suppressible hyper-induction of the UPR in <i>msc2Δ zrg17Δ zrc1Δ cot1Δ</i> quadruple mutants. JSY5 (<i>msc2Δ zrg17Δ zrc1Δ cot1Δ</i>) and JSY5 <i>ice2Δ</i> (<i>msc2Δ zrg17Δ zrc1Δ cot1Δ ice2Δ</i>) cells were transformed with the UPRE-lacZ reporter pMCZ-Y and inoculated into low zinc medium (LZM) supplemented with 1, 3, 10 or 100 µM ZnCl<sub>2</sub>. These cells were then grown overnight prior to measuring β–galactosidase activity. The wild-type strain used was the isogenic CM100 strain. D) Zinc treatment does not inhibit the UPR induction in response to tunicamycin. Wild-type BY4743 cells bearing the UPRE-lacZ reporter were grown to exponential phase in LZM supplemented with the indicated concentration of zinc, then treated for 2 hours with 2 µg/ml tunicamycin prior to β–galactosidase activity assay. Data presented are the averages of triplicate cultures for each condition and the error bars indicate ±1 S.D.</p
