AN INTRICATE ARRANGEMENT OF BINDING-SITES FOR THE ETS FAMILY OF TRANSCRIPTION FACTORS REGULATES ACTIVITY OF THE ALPHA-4 INTEGRIN GENE PROMOTER

alpha 4 integrins mediate cell-cell and cell-extracellular matrix inte ractions that are critical for maturation and function of the immune s ystem as well as differentiation of skeletal muscle. Here we examine m olecular mechanisms controlling the pattern of alpha 4 expression. The activity of constructs containing 5' deletion mutants of the alpha 4 gene promoter was compared in transfection assays into cell lines that express alpha 4 and cell lines that do not. The sequence between posi tion -42 and -76 base pairs (bp) was required for efficient transcript ion in cells that express alpha 4, but it showed no activity in HeLa c ells, which do not express alpha 4. Three binding sites for the Ets fa mily of transcription factors are found in this region: two adjacent s ites at positions -50 and -54 bp and a more 5' site at position -67 bp . Using a series of constructs containing deletions and mutations in t his region, we found that the 3'-most site alone was sufficient for bi nding GA-binding protein alpha (GABP alpha)/GABP beta and for a low le vel of transcriptional activation. When all three sites were present, a second complex ''a'' was detected, which contains an unknown member of the Ets family. Formation of complex a was cell-type specific and c orrelated with a high level of transcription. Deletion of the 5'-most Ets site had no effect on binding to GABP alpha/GABP beta, but it elim inated a. Concomitant with this loss of a, a new Ets-1-containing comp lex ''c'' appeared. Complex c substituted efficiently for complex a in transcriptional activation. We conclude that although neither of the two 5'-most Ets sites alone binds nuclear protein, they appear to act as modulators which control the pattern of Ets proteins that bind the alpha 4 gene promoter. This arrangement of Ets sites, coupled with the tissue- and developmental-specific expression of Ets members, likely play a key role in defining the pattern of alpha 4 integrin.