Identification and functional characterization of the human T-cell receptor beta gene transcriptional enhancer: common nuclear proteins interact with the transcriptional regulatory elements of the T-cell receptor alpha and beta genes.
A transcriptional enhancer has been mapped to a region 5.5 kilobases 3' of the C beta 2 gene in the human T-cell receptor (TCR) beta-chain locus. Transient transfections allowed localization of enhancer activity to a 480-base-pair HincII-XbaI restriction enzyme fragment. The TCR beta enhancer was active on both the minimal simian virus 40 promoter and a TCR beta variable gene promoter in both TCR alpha/beta + and TCR gamma/delta + T cells. It displayed significantly less activity in Epstein-Barr virus-transformed B cells and K562 chronic myelogenous leukemia cells and no activity in HeLa fibroblasts. DNA sequence analysis revealed that the enhancer contains a consensus immunoglobulin kappa E2 motif, as well as an AP-1-binding site and a cyclic AMP response element. DNase I footprint analyses using Jurkat T-cell nuclear extracts allowed the identification of five nuclear protein-binding sites, T beta 1 to T beta 5, within the enhancer element. Deletion and in vitro mutagenesis studies demonstrated that the T beta 2- and T beta 3- and T beta 4-binding sites are each required for full transcriptional enhancer activity. In contrast, deletion of the T beta 1- and T beta 5-binding sites had essentially no effect on enhancer function. Electrophoretic mobility shift assays demonstrated that TCR alpha/beta + and TCR gamma/delta + T cells expressed T beta 2-, T beta 3-, and T beta 4-binding activities. In contrast, non-T-cell lines, in which the enhancer was inactive, each lacked expression of at least one of these binding activities. TCR alpha and beta gene expression may be regulated by a common set of T-cell nuclear proteins in that the T beta 2 element binding a set of cyclic AMP response element-binding proteins that are also bound by the T alpha 1 element of the human TCR alpha enhancer and the decamer element present in a large number of human and murine TCR beta promoters. Similarly, the T beta 5 TCR beta-enhancer element and the T alpha 2 TCR alpha-enhancer element bind at least one common T-cell nuclear protein. Taken together, these results suggest that TCR beta gene expression is regulated by the interaction of multiple T cell nuclear proteins with a transcriptional enhancer element located 3' of the C beta 2 gene and that some of these proteins may be involved in the coordinate regulation of TCR alpha and beta gene expression.
Gottschalk
L R
LR
Howard Hughes Medical Institute, University of Michigan Medical Center, Ann Arbor 48109-0650.
Leiden
J M
JM
D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D004252:DNA Mutational Analysis; D004742:Enhancer Elements, Genetic; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000502:physiology; D009841:Oligonucleotides; D011401:Promoter Regions, Genetic; D011948:Receptors, Antigen, T-Cell / Q000235:genetics*; D016693:Receptors, Antigen, T-Cell, alpha-beta; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic
D009687:Nuclear Proteins; D009841:Oligonucleotides; D011948:Receptors, Antigen, T-Cell; D016693:Receptors, Antigen, T-Cell, alpha-beta