Enhancer-blocking activity within the DNase I hypersensitive site 2 to 6 region between the TCR alpha and Dad1 genes

Although tightly linked, the TCR alpha and 6 genes are expressed specifical ly in T lymphocytes, whereas the Dad1 gene is ubiquitously expressed. Betwe en TCR alpha and Dad1 are eight DNase I hypersensitive sites (HS), HS1 colo calizes with the TCR alpha enhancer (E-alpha) and is T cell-specific; HS2, -3, -4, -5, and -6 map downstream of HS1 and are tissue-nonspecific. The re gion spanning HS2-6 was reported to display chromatin-opening activity and to confer copy number-dependent and integration site-independent transgene expression in transgenic mice. Here, we demonstrate that HS2-6 also display s enhancer-blocking activity, as it can block an enhancer from activating a promoter when located between the two in a chromatin-integrated context, a nd can do so without repressing either the enhancer or the promoter. Multip le enhancer-blocking elements are arrayed across HS2-6. We show that HS2-6 by itself does not activate transcription in chromatin context, but can syn ergize with an enhancer when located upstream of an enhancer and promoter. We propose that HS2-6 primarily functions as an insulator or boundary eleme nt that may be critical for the autonomous regulation of the TCR alpha and Dad1 genes.