The metallothioneins provide a useful model for studying the environmental and developmental regulation of eukaryotic gene expression. The DNA sequences and cellular factors involved in the heavy metal induction of mammalian metallothionein gene transcription have been investigated by in vitro mutagenesis, gene transfer and factor titration experiments. The results indicate that cellular components interact with two distinct regions of the upstream flanking DNA and activate transcription by a positive regulatory mechanism. Cells from patients with Menkes' disease, an inherited disorder of copper metabolism, are defective in some step of the regulatory pathway. Analysis of three human metallothionein genes has shown that regulation also occurs during development, possibly associated with changes in methylation. Genetic strategies have been developed to study the regulation of a metallothionein-like protein in a lower eukaryotic, and metallothionein-based expression vectors have been utilized to overproduce useful proteins such as growth hormone and hepatitis surface antigen.