TRANSCRIPTIONAL REGULATION OF THE HUMAN ALPHA-2(I) COLLAGEN GENE - COMBINED ACTION OF UPSTREAM STIMULATORY AND INHIBITORY CIS-ACTING ELEMENTS

This study identifies three regions of the human alpha 2(I) collagen p romoter involved in the binding of nuclear factors. These regions incl ude sequences from -173 to -155 (footprint I), -133 to -119 (footprint II), and -101 to -72 (footprint III). A novel positive cis-element co ntaining a TCCTCC motif was identified within footprint II. In additio n, we demonstrated that a pyrimidine-rich region within footprint I is a binding site for a transcriptional repressor, and a CCAAT motif wit hin footprint III is a binding site for a transcriptional activator, C omparative functional analysis of the cis-acting elements within the p roximal 350 base pairs of this promoter, including previously characte rized Sp1 binding sites at -300, indicates that constitutive activity of this promoter is regulated equivalently by the three positive cis-a cting elements at -300, -125, and -80. Mutations in the repressor site at -160 increase constitutive activity by 4-6 fold. However, simultan eous mutations of the repressor site and the cis-regulatory element at either the -300 or -125 sites result in no increase in constitutive t ranscription activity suggesting interaction between the activators an d repressor elements, In contrast, simultaneous mutation of the CCAAT motif and the repressor site results in about a 4-fold increase, sugge sting that activation via the CCAAT motif may be independent of this r epressor.