A Targeted Mutation at the T-Cell Receptor α/δ Locus Impairs T-Cell Development and Reveals the Presence of the Nearby Antiapoptosis Gene <i>Dad1</i>

Hong, Nam-Doo; Cado, Dragana; Mitchell, James R.; Ortiz, Benjamin; Hsieh, Shu-Ching; Winoto, Astar

doi:10.1128/mcb.17.4.2151

Cited by 44 publications

(59 citation statements)

References 37 publications

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“…TCR␣ and TCR␦ genes are expressed by the ␣␤ and ␥␦ lineages of T cells, respectively. In contrast, Dad1 is expressed ubiquitously (1). In addition to the appropriate V-D-J rearrangements that generate functional TCR␣ and TCR␦ chains, this locus is often involved in aberrant gene rearrangements that can lead to T cell malignancies (8).…”

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confidence: 99%

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Function and Factor Interactions of a Locus Control Region Element in the Mouse T Cell Receptor-α/Dad1 Gene Locus

Ortiz

Harrow

Cado

et al. 2001

The Journal of Immunology

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Locus control regions (LCRs) refer to cis-acting elements composed of several DNase I hypersensitive sites, which synergize to protect transgenes from integration-site dependent effects in a tissue-specific manner. LCRs have been identified in many immunologically important gene loci, including one between the TCRδ/TCRα gene segments and the ubiquitously expressed Dad1 gene. Expression of a transgene under the control of all the LCR elements is T cell specific. However, a subfragment of this LCR is functional in a wide variety of tissues. How a ubiquitously active element can participate in tissue-restricted LCR activity is not clear. In this study, we localize the ubiquitously active sequences of the TCR-α LCR to an 800-bp region containing a prominent DNase hypersensitive site. In isolation, the activity in this region suppresses position effect transgene silencing in many tissues. A combination of in vivo footprint examination of this element in widely active transgene and EMSAs revealed tissue-unrestricted factor occupancy patterns and binding of several ubiquitously expressed transcription factors. In contrast, tissue-specific, differential protein occupancies at this element were observed in the endogenous locus or full-length LCR transgene. We identified tissue-restricted AML-1 and Elf-1 as proteins that potentially act via this element. These data demonstrate that a widely active LCR module can synergize with other LCR components to produce tissue-specific LCR activity through differential protein occupancy and function and provide evidence to support a role for this LCR module in the regulation of both TCR and Dad1 genes.

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“…1A). This locus contains three differentially regulated genes, TCR␣, TCR␦, and Dad1 (1). The first two genes are components of ␣␤ and ␥␦ T cell Ag receptors, respectively.…”

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confidence: 99%

Function and Factor Interactions of a Locus Control Region Element in the Mouse T Cell Receptor-α/Dad1 Gene Locus

Ortiz

Harrow

Cado

et al. 2001

The Journal of Immunology

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“…One explanation for the ectopic transgene expression is that the TCR § chain locus is at least partially open in non-T cells since HS from the TCR § LCR are detectable in tissues that do not contain T cells [26]. The TCR § LCR controls the ubiquitously expressed anti-apoptotic gene Dad1 contained in the TCR § locus.…”

Section: Discussionmentioning

confidence: 99%

Ectopic T cell receptor expression causes B cell immunodeficiency in transgenic mice

2004

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Mice expressing transgenic T cell receptors (TCR) are used to explore important questions in immunity. However, transgene expression may have unexpected effects. We previously reported a B cell immunodeficiency, comprising decreased B cell numbers and diminished antibody responses, in mice that express a transgenic TCR specific for nicotinic acetylcholine receptor; the mice were generated using cassette vectors designed specifically for transgenic TCR expression [see Kouskoff et al. J. Immunol. Methods 1995. 180: 273-280]. We now show data suggesting that this defect is due to the expression and accumulation of TCR § and g chains inside B cells and induction of an endoplasmic reticulum stress response, causing apoptosis at the pre B-I and later B cell stage. Thus, inappropriate transgene expression can profoundly affect B cells, leading to a previously undescribed mechanism of immunodeficiency.

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“…Knowledge of the cis-regulatory features of this locus derives largely from functional studies of the homologous mouse domain 19 , including the identification of a locus control region 20 and an insulator activity 21 . In human cells, (HS1-HS5); the HBE promoter together with an upstream element; the HBG1 promoter; the HBG2 promoter; the HBG2 3¢ enhancer; the HBD promoter; the HBB promoter; and the HBB 3¢ enhancer.…”

Section: Analysis Of the Human Tcra Regulatory Domainmentioning

confidence: 99%

“…Primers were designed using Primer3 (ref. 24) restricting several parameters, including target amplimer size (250 bp 7 50 bases), primer T m (optimal, 60 1C 7 2 1C), %GC (50% optimal, range 40-80%) and length (optimal 24, range [19][20][21][22][23][24][25][26][27], and the poly X (maximum 4). Primers were then scanned for repetitive sequences by alignment with a database of repetitive sequences.…”

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confidence: 99%

High-throughput localization of functional elements by quantitative chromatin profiling

et al. 2004

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Identification of functional, noncoding elements that regulate transcription in the context of complex genomes is a major goal of modern biology. Localization of functionality to specific sequences is a requirement for genetic and computational studies. Here, we describe a generic approach, quantitative chromatin profiling, that uses quantitative analysis of in vivo chromatin structure over entire gene loci to rapidly and precisely localize cis-regulatory sequences and other functional modalities encoded by DNase I hypersensitive sites. To demonstrate the accuracy of this approach, we analyzed B300 kilobases of human genome sequence from diverse gene loci and cleanly delineated functional elements corresponding to a spectrum of classical cis-regulatory activities including enhancers, promoters, locus control regions and insulators as well as novel elements. Systematic, highthroughput identification of functional elements coinciding with DNase I hypersensitive sites will substantially expand our knowledge of transcriptional regulation and should simplify the search for noncoding genetic variation with phenotypic consequences.Understanding the human genome will require comprehensive delineation of functional elements within the 98% of genomic terrain that does not encode protein. In vivo, cis-regulatory modalities colocalize with focal alterations in chromatin structure [1][2][3][4] , and this governs the accessibility of genomic sequences to critical regulatory factors. Exploitation of the close connection between functional elements and chromatin structure should offer a powerful and generic approach for de novo identification of cis-regulatory sequences in the context of complex gene domains.Active regulatory elements within complex genomes are distinguished by pronounced sensitivity to the nonspecific endonuclease DNase I 3-5 when exposed in the context of intact nuclei. DNase I hypersensitive sites are the sine qua non of a diverse spectrum of classical transcriptional and chromosomal regulatory activities including enhancers, promoters, silencers, insulators, boundary elements and locus control regions 1,3,6 . Indeed, in the human genome, many functional elements were first identified as major DNase I hypersensitive sites and only later were found to have specific regulatory roles. Analysis of chromatin structure may enable generic delineation of functional elements across the genome, provided it exhibits direct sequence specificity, quantitative data output that permits automated analysis, and adaptability to a high-throughput format.DNase I hypersensitive sites in native genomic domains have traditionally been localized by an approach relying on Southern transfer followed by indirect end-labeling 5 . Although widely applied, this technique is not quantitative and has numerous technical and resource-related limitations that prevent its application on a genome-wide scale. The major limitations of conventional hypersensitivity assays are the low throughput and the lack of sequence specificity. Conventional So...

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A Targeted Mutation at the T-Cell Receptor α/δ Locus Impairs T-Cell Development and Reveals the Presence of the Nearby Antiapoptosis Gene Dad1

Cited by 44 publications

References 37 publications

Function and Factor Interactions of a Locus Control Region Element in the Mouse T Cell Receptor-α/Dad1 Gene Locus

Function and Factor Interactions of a Locus Control Region Element in the Mouse T Cell Receptor-α/Dad1 Gene Locus

Ectopic T cell receptor expression causes B cell immunodeficiency in transgenic mice

High-throughput localization of functional elements by quantitative chromatin profiling

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