Restriction enzyme digestions identify discrete domains in the chromatin around the promoter of the XXXX alpha 2(I) collagen gene.

@entity19 @entity9828 @entity1582
We have examined the chromatin structure around the +1 transcriptional start site of the @entity19 alpha 2(I) collagen gene by studying the accessibility of DNA to several restriction enzymes as well as to @entity9828 . In NIH 3T3 cells, which express high levels of alpha 2(I) collagen mRNA, we detect a @entity9828 -hypersensitive site from -240 to +110 relative to the start site of transcription at +1. By digesting chromatin with restriction enzymes, which cleave naked DNA at multiple sites within the -2000 to +1000 region, a considerably more complex picture was revealed. DNA sequences upstream of around -550 and downstream of +150 are much less accessible to restriction enzymes than the region between these sites and are, therefore, probably packaged in a more compact conformation. The region from around -550 to -240 although not within the @entity9828 -hypersensitive domain is nevertheless accessible to restriction enzymes and, therefore, presumably in a relatively "open" conformation. In addition, beginning 5' to -100 there is a gradual decrease in restriction enzyme accessibility as one approaches +150. Of particular interest is the finding that although sites at +65 and +126 are relatively accessible, a HinfI site at +113 is resistant in chromatin. In v-mos transformed NIH 3T3 cells which express alpha 2(I) collagen at much lower levels than untransformed NIH 3T3 cells, the @entity9828 -hypersensitive site as well as the majority of the chromatin restriction enzyme accessibility patterns are similar to those found in untransformed NIH 3T3 cells. However, a SphI site at +58 appears less accessible in the transformed cells. We also examined the chromatin of a @entity1582 cell line which does not synthesize alpha 2(I) collagen at detectable levels. In the nuclei of these cells the DNA of the alpha 2(I) collagen promoter is inaccessible to @entity9828 and to all restriction enzymes.
