Figure 4.

Recruitment of Swi3, Med15, and Spt3 is zinc responsive and Zap1 dependent. Untagged wild-type (BY4741), and isogenic Swi3-TAP, Med15-TAP, and Spt3-TAP tagged strains were grown in LZM supplemented with either 1 μM (−, lanes 1–4) or 1000 μM (+, lanes 5–8) ZnCl₂. Isogenic zap1Δ cells grown in low zinc were used in lanes 9–12. The cells were cross-linked with formaldehyde, harvested, and chromatin immunoprecipitation analysis was performed using IgG-Sepharose to immunoprecipitate the TAP-tagged proteins. Coprecipitation of specific DNA fragments was then assessed by PCR using primers flanking the ZREs of the ZRT1 and ZPS1 promoters. Primers specific for the CMD1 promoter were used as a negative control. PCR amplification of 10-fold serial dilutions of input samples was used to confirm the quantitative nature of the assay.