Previous studies in our laboratory have shown that a trans-acting factor, which binds to a 106 bp sequence in the mouse metallothionein-I (MT-I) gene, is responsible for the relatively high level of MT-I gene transcription in the liver. Using electrophoretic mobility shift assay, we have now identified a 26 bp sequence within the 106 bp region, which interacts with a trans-activating factor in the liver nuclear extract. This sequence, designated MRE-c', is located between positions -135 and -110 with respect to the transcription start site and comprises the metal regulatory element MRE-c and part of its 5' and 3' flanking sequences. UV cross-linking and Southwestern analysis showed that a protein of an apparent molecular mass of 33,000 specifically interacts with MRE-c'. Deletion of the MRE-c' region resulted in a six- to sevenfold decrease in the MT-I promoter activity, as measured by reduction in chloramphenicol acetyltransferase activity. A comparison of other regulatory domains of the MT-I gene and the potential factors interacting with these sequences indicates that MRE-c' and probably the 33 kDa polypeptide are involved in the constitutive transcription of the MT-I gene.
Animals Base Sequence