Fig. 4.

Z-DNA formation demarcates the boundaries of neighboring nucleosomes. (A) Reporter gene activities are shown for the promoter constructs with either wild-type (+) or mutant (−) TATA box in the presence (+) or absence (−) of the upstream d(CG)₉ repeat sequence. (B) Nucleosome-scanning assay. (Upper) Nucleosome occupancy is measured for four overlapping regions (≈100–140 bp) surrounding the d(CG)₉ repeat sequence at the mutant TATA box promoter (regions 1 to 4). DNA of the promoter region is depicted as a line with 10-bp intervals (vertical lines). Location of d(CG)₉ repeat sequence is denoted as an insertion at 0 bp. In plasmids without this Z-forming sequence, it is replaced by a random sequence of approximately equal length (shortened by 3 bp). Regions 1 through 4 are indicated by horizontal lines below the DNA. (Lower) Nucleosome occupancy of the four overlapping regions is measured by the relative amounts of PCR products generated in the absence (−) or presence (+) of the upstream promoter-proximal d(CG)₉ repeat sequence at the mutant TATA box promoter. Relative to the ends of the d(CG)₉ repeat sequence, the locations of the regions 1 to 4 determined by PCR primers are as given in parentheses: region 1 (−109 to −5), region 2 (−71 to +26), region 3 (−28 to +72), and region 4 (+3 to +145). Specific PCR products of interest in the electrophoresis gels are indicated by arrowheads on the right. The asterisk in region 4 indicates a nonspecific PCR product also observed in the absence of reporter plasmid.