FIG. 5.

P58^IPK represses PKR function in vivo. (A) Yeast growth analysis. Yeast strain RY1-1 was transformed with the 2μm expression plasmid pEMBLYex4 (vector; position 3), pYex-PKR Δ295–300 (PKR Δ295–300; position 2), or pYex-P58^IPK (P58^IPK; position 4) and streaked onto uracil-deficient medium containing 2% dextrose (SD) or 10% galactose (SGAL). To control for the specificity of growth effects due to PKR or P58^IPK expression, the gcn2Δ isogenic control strain, J110, harboring vector alone (position 1) was included in all analyses. (B) Protein expression. Lanes 1 to 3 contain extracts from the RY1-1 strains shown in panel A harboring pYex-P58^IPK (lane 1), pEMBLYex4 alone (lane 2), or pYex-PKR Δ295–300 (lane 3). An extract from the J110 parental control strain harboring pEMBLYex4 is shown in lane 4. Strains were grown for 5 h in galactose-containing liquid medium, and extracts were prepared as described in Materials and Methods. Proteins (25 μg) were separated by SDS-PAGE and subjected to immunoblot analysis. Panels show the same blot probed sequentially with antibodies specific to PKR, P58^IPK, or actin.