Building a metal-responsive promoter with synthetic regulatory elements

Area of Science:

• Molecular Biology
• Gene Regulation

Background:

• The mouse metallothionein-I gene promoter contains multiple metal regulatory elements (MREs).
• These MREs are located between -176 and -44 base pairs from the cap site.

Purpose of the Study:

• To investigate the function of MREs in gene regulation.
• To determine the role of MRE-a in zinc-inducible gene expression.

Main Methods:

• Constructing fusion genes with the metallothionein-I promoter and herpes simplex virus thymidine kinase gene.
• Transfecting plasmids into baby hamster kidney cells for transient assays.
• Inserting synthetic DNA fragments containing MRE-a into the thymidine kinase promoter.

Main Results:

• Multiple MREs are present in the metallothionein-I promoter region.
• Two copies of MRE-a significantly increased zinc inducibility.
• Inducibility was enhanced by additional MRE-a sequences.
• MRE-a demonstrated synergistic function with thymidine kinase promoter elements and acted as a positive, zinc-dependent element with the TATA box.

Conclusions:

• MRE-a functions as a potent, zinc-dependent regulatory element.
• Multiple MREs contribute to the efficient regulation of gene expression by zinc.
• The number and arrangement of MREs influence the degree of gene induction.