HMG-CoA reductase (HMGR) catalyzes the rate-controlling step in cholesterol production. This enzyme is highly expressed in the liver, where it is subject to extensive hormonal and dietary regulation. Although much is known about the regulation of the HMGR promoter in cultured cells, this issue has not been directly addressed in liver. The technique of in vivo electroporation was utilized to perform the first functional analysis of the HMGR promoter in live animals. Analysis of a series of deletion constructs showed that deletion of the region containing the cyclic AMP response element (CRE) at -104 to -96 and an NF-Y site at -70 to -65 resulted in marked reduction of promoter activity. Sterol regulation of this promoter was investigated by raising tissue cholesterol levels by feeding cholesterol and by decreasing them through administration of a statin (lovastatin). Using this approach, we found that HMGR promoter constructs were sterol responsive in live animals, adding in vivo relevance to previous findings in cultured cells. We also conclude that in vivo electroporation is a convenient and powerful technique for the analysis of promoter elements in the livers of live animals.