Regulation of alpha-tropomyosin and N5 genes by a shared enhancer. Modular structure and hierarchical organization.

We have identified a 99-base pair (bp) tandem repeat consisting of two modules/repeat: a 40-bp-long d(CA.GT)20 potential Z-DNA sequence and a 59-bp-long unique sequence. This repeat is located between and differentially regulates two transcription units, rat alpha-tropomyosin and N5, organized in "head-to-head" orientation. Transient expression experiments showed orientation- and position-independent enhancer activity of the 99-bp repeat in a tissue-specific and developmental manner. Deletion analysis revealed a hierarchical organization establishing the two modules as protoenhancers. Southwestern blot analysis of crude nuclear extracts identified at least six C2C12 myotube nuclear proteins binding to the 99-bp repeat. These observations demonstrate a mammalian gene regulatory sequence structurally and functionally organized like the simian virus 40 viral enhancer and confirm in a specific gene context the contribution of a d(CA.GT)20 tract in transcriptional regulatory mechanisms as a protoenhancer.