Functional analysis of DNA sequences located within a cluster of DNase I hypersensitive sites colocalizing with a MAR element at the upstream border of the chicken alpha-globin gene domain

We have cloned and sequenced a genomic DNA fragment of chicken containing a cluster of DNase I hypersensitive sites (DHS) located 11-15 kb upstream fr om the first gene of the a-globin gene domain and including a constitutive DHS flanked by two erythroid-specific ones. A 1.2-kb subfragment of the DNA fragment under study located upstream to the constitutive DHS and colocali zing roughly with one of the erythroid-specific DHS was shown to possess th e properties of a matrix association region (MAR). The cloned DNA sequences were tested for their ability to serve as promoters and/or influence trans cription from the promoter of the alpha(D) globin gene. In the region studi ed, we did not find any promoters or enhancers that were active in erythroi d cells. The whole DNase I hypersensitive region and some of its subfragmen ts showed a silencing effect when placed downstream from the reporter gene. The expression of the reporter gene was completely abolished, however, whe n these DNA fragments were placed between the aD promoter and the reporter gene. Thus, they seem to act as transcription "terminators." Numerous polya denylation signals (AATAAA) and an AT-rich palindrome were found within the sequenced DNA fragment. These observations are discussed within the frame of the hypothesis postulating that continuous transcription is essential fo r maintaining the active status of genomic domains. Furthermore, it is sugg ested that the DNA fragment studied contains a negative control element tha t keeps globin genes silent within the chromatin domain permanently open in nonerythroid cells. J. Cell. Biochem. 74:38-49, 1999. (C) 1999 Wiley-Liss, Inc.