Figure 4.—

Effect of mutations blocking both the IMP and the histidine biosynthesis pathways from accumulating AICAR. (A–C). Yeast strains transformed with a plasmid carrying the ADE1-lacZ fusion (P115) were grown for 6 hr in SD casa medium containing adenine at either low (0.025 mm) or high (0.3 mm) concentration. β-Gal activity was then measured as described in materials and methods. Strains were: (A) BY4742 (wild type), Y16591 (ade3), Y10190 (his1), Y13388 (his7) Y1166 (ade3 his1), and Y1661 (ade3 his7); (B) BY4742 (wild type), Y16591 (ade3), Y1551 (ade3 ade5,7), and Y1554 (ade3 ade6); and (C) BY4742 (wild type), Y16591 (ade3), Y1166 (ade3 his1), Y1551 (ade3 ade5,7), and Y1657 (ade3 ade5,7 his1). (D) Northern blot analysis of ADE17 gene expression in the ade2 ade3 ade13 mutant strain. Strains BY4742 (wild type), Y10190 (his1), Y14601 (ade5,7), Y16591 (ade3), Y1166 (ade3 his1), Y1551 (ade3 ade5,7), and Y1657 (ade3 ade5,7 his1) were grown to an OD₆₀₀ of 0.5 in SD casa medium with or without adenine as indicated. Extraction of RNA and hybridization were performed as described in materials and methods. Radioactivity was detected using a phosphorimager and signal was quantified using the ImageQuant software. The quantification is presented as the ADE17/ACT1 ratio, which was arbitrarily set up as “1” in the wild-type “+ade” control lane.