FIG.4.

GRE and TATA2 mutations affect endogenous IMD2 transcriptional regulation and resistance of cells to MPA. (A) Sequence of the IMD2 promoter region, with the GRE and AT-rich sequences shown in bold type. (B) Yeast strains carrying the indicated IMD2 sequences were treated or not treated with guanine (Gua) or MPA. Total cellular RNA was extracted, and the levels of transcripts of IMD2 and DED1 (as an internal control) were measured by S1 nuclease analysis. wt, wild type; mut, mutant. (C) The 5′ end of the IMD2 transcripts was characterized by 5′ extension with the indicated yeast strains under various growth conditions. The oligonucleotide used in the extension reactions was also used for sequencing to identify the position of the start site. (D) The indicated yeast strains were grown to exponential phase and plated in serial dilutions on plates containing or not containing Gua, MPA, or 6-AU. Plates were incubated at 30°C for 3 days. (E) S1 nuclease analysis of induction of IMD2 transcripts by MPA and 6-AU.