Technical Abstract: We are studying the mechanisms regulating the expression of the genes in the casein gene cluster region. Multi-species comparative sequence analysis showed conserved linkage in this genomic domain, which contains the casein gene cluster, consisting of three (or four depending on the species) evolutionarily related genes and one physically and functionally linked gene, the three-member HTN/STATH gene-family, the proline-rich protein gene family PROL-1, -3 &-5, MUC-7, NYD-SP26, and four other genes. This region appears to be mammalian specific. All genes in this region encode, or are predicted to encode, secreted proteins and are expressed in tissues with epithelial components. The caseins, expressed in breast epithelium in a developmental-stage specific fashion, are essential for the growth and survival of the offspring and constitute markers for terminal differentiation of the breast epithelial cells. In addition, the other genes are expressed in salivary and/or mammary gland tissue. Despite high sequence divergence in the coding regions of the genes in this domain 23 conserved non-coding elements were identified based on comparative analysis and their presence in all species studied. One of these regions includes a beta-casein gene enhancer, 6.3 kb 5' to the start site of transcription in the mouse, which displays lactogenic hormone-dependent histone acetylation. These conserved elements and their potential role in gene regulation in this genomic domain are being studied using computational approaches as well as approaches to study the chromatin structure of this domain.