TRANSCRIPTIONAL REPRESSION OF THE ALPHA-1(I) COLLAGEN GENE BY RAS IS MEDIATED IN PART BY AN INTRONIC AP1 SITE

We have previously shown that transformation of fibroblasts by ras res ults in transcriptional inhibition of the alpha 1(I) gene. An alpha 1( I)-hGH chimeric plasmid containing 3.7 kb of 5' flanking and 4.4 kb of alpha 1(I) transcribed sequence was regulated appropriately by ras in a transient transfection assay. in contrast, a similar plasmid contai ning alpha 1(I) DNA from -220 to +500 was virtually unresponsive to ra s. The regions from -3700 to -220 and +500 to +4400 contributed equall y to the ras-mediated inhibition of the parental plasmid. Deletion ana lysis indicated that a short fragment, between +500 and +890 in the fi rst intron of the alpha 1(I) gene, was recognized differently in ras-t ransformed and wild-type cells. A previously described AP1 site in thi s fragment stimulated al (I) transcription in Rat1 fibroblasts but was inactive in ras-transformed cells. Mobility shift assays using nuclea r extracts from the two cell types demonstrated differences in binding to the alpha 1(I) AP1 site. We conclude that ras transformation suppr esses the function of a cell-specific enhancer in the first intron of the alpha 1(I) collagen gene. (C) 1995 Wiley-Liss, Inc.