Figure 9.

Transcriptional Regulation of MSD3 by Poor Fe Nutrition or by H₂O₂ Stress.

(A) Schematic of reporter gene construct. White segments represent the ARS reporter gene, black the minimal TUB2 promoter, red the MSD3 5′ flanking DNA. The numbers beneath the MSD3 5′ flanking segment indicate positions relative to the transcript start site determined by 5′-rapid amplification of cDNA ends.

(B) Arylsulfatase activity on plates. Single representative transformants growing on agar-solidified TAP medium with the indicated Fe supplement were stained for arylsulfatase activity (visualized as a blue halo around colonies) by spraying with a solution of the chromogenic substrate 5-bromo-4-chloro-3-indoxyl sulfate.

(C) Coordinate accumulation of mRNAs for MSD3 (closed symbols) and the ARS2 reporter gene driven by the MSD3 promoter (open symbols) in cells challenged with H₂O₂. Transformants carrying either the MSD3 5′ flanking DNA fused to the ARS2 reporter gene (circles), or the ARS2 reporter gene alone (squares), were grown to early log phase (2 × 10⁶ cells/mL) in TAP medium, and H₂O₂ was added to 1 mM. Cells were collected at the time points shown and RNA analyzed by real-time PCR for the abundance of transcripts encoding MSD3 and ARS2. Data shown are for representative individual transformants.