BIOCHEMICAL AND FUNCTIONAL ANALYSES OF CHROMATIN CHANGES AT THE TCR-BETA GENE LOCUS DURING CD4(-)CD8(-) TO CD4(+)CD8(+) THYMOCYTE DIFFERENTIATION

Allelic exclusion is the process wherein lymphocytes express Ag recept ors from only one of two possible alleles, and is effected through a f eedback inhibition of further rearrangement of the second allele, The feedback signal is thought to cause chromatin changes that block acces sibility of the second allele to the recombinase, To identify the puta tive chromatin changes associated with allelic exclusion, we assayed f or DNase I hypersensitivity, DNA methylation, and transcription in 100 kb of the TCR-beta locus, Contrary to current models, we identified c hromatin changes indicative of an active and accessible locus associat ed with the occurrence of allelic exclusion, Of 11 DNase I hypersensit ive sites identified, 3 were induced during CD4(-)CD8(-) to CD4(+)CD8( +) thymocyte differentiation, and demethylation and increased germline transcription of the locus were evident, We further examined the role of the most prominently induced site near the TCR-beta enhancer (E be ta) in allelic exclusion by targeted mutagenesis. Two other sites were also examined in New Zealand White (NZW) mice that have a natural del etion in the TCR-beta locus, TCR-beta gene recombination and allelic e xclusion were normal in both mutant mice, negating dominant roles for the three hypersensitive sites in the control of allelic exclusion, Th e data suggest that alternative cis-regulatory elements, perhaps conta ined in the E beta enhancer and/or in the upstream V beta region, are involved in the control of TCR-beta allelic exclusion.