Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The abundance of delta-crystallin in the chicken eye lens provides an advantageous marker for tissue-specific gene expression during cellular differentiation. The lens-specific expression of the delta 1-crystallin gene is governed by an enhancer in the third intron, which binds a positive (delta EF2) and negative (delta EF1) factor in its core region. Here we show by DNase I footprinting, electrophoretic mobility-shift assays, and cotransfection experiments with the delta 1-promoter/enhancer fused to the chloramphenicol acetyltransferase reporter gene that the delta 1-crystallin enhancer has two adjacent functional Pax-6 binding sites. We also demonstrate by DNase I footprinting that the delta EF1 site can bind the transcription factor USF, raising the possibility that USF may cooperate with Pax-6 in activation of the chicken delta 1- and alpha A-crystallin genes. These data, coupled with our recent demonstration that Pax-6 activates the alpha A-crystallin gene, suggest that Pax-6 may have been used extensively throughout evolution to recruit and express crystallin genes in the lens.
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PMID:Pax-6 and lens-specific transcription of the chicken delta 1-crystallin gene. 775 64

We have demonstrated previously that a transgene comprising the -164/+44 fragment of the murine alphaB-crystallin gene fused to the bacterial chloramphenicol acetyltransferase (cat) gene is lens-specific in transgenic mice. The -147 to -118 sequence was identified as a lens-specific regulatory region and is called here LSR1 for lens-specific region 1. In the present experiments, a -115/+44-cat transgene was also lens-specific in transgenic mice, although the average activity was 30 times lower than that derived from the -164/+44-cat transgene. The -115/+44 alphaB-crystallin fragment contains a highly conserved region (-78 to -46) termed here LSR2. A -68/+44-cat transgene, in which LSR2 is truncated, was inactive in transgenic mice. DNase I footprinting indicated that LSR1 and LSR2 bind partially purified nuclear proteins from either alphaTN4-1 lens cells or the mouse lens as well as the purified paired domain of Pax-6. Site-specific mutation of LSR1 eliminated both Pax-6 binding and promoter activity of the -164/+44-cat transgene in transgenic mice. Finally antibody/electrophoretic mobility shift assays and cotransfection experiments indicated that Pax-6 can activate the alphaB-crystallin promoter via LSR1 and LSR2. Our data strengthen the idea that Pax-6 has had a major role in recruiting genes for high expression in the lens.
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PMID:Pax-6 and alphaB-crystallin/small heat shock protein gene regulation in the murine lens. Interaction with the lens-specific regions, LSR1 and LSR2. 879 91

There are two tandemly linked delta-crystallin genes [5' delta 1 -delta 2 3'] in the chicken, with the delta 1-crystallin gene being expressed much more highly (50-100-fold) in the embryonic lens than the delta 2-crystallin gene. Previous transfection experiments have shown that a lens-preferred enhancer exists in the third intron of each chicken delta-crystallin gene. In the present investigation we have used transgenic mice to establish that both the chicken delta 1- and delta 2-crystallin enhancers are preferentially active in the mouse lens in combination with their homologous promoter and the chloramphenicol acetyltransferase (CAT) reporter gene. The promoter/ CAT constructs lacking the enhancers were inactive in the transgenic mice. In one case, a truncated delta 2-crystallin promoter (-308/+24) in combination with the enhancer was also active in the Purkinje cells of the cerebellum of the transgenic mice, which could prove useful in future experiments. Finally, retinoic acid receptors (RAR beta) activated the delta 1-crystallin, but not the delta 2-crystallin enhancer in teh recombinant plasmids in cotransfected embryonic chicken lens epithelial cells treated with retinoic acid. This activation did not occur when using the care enhancer (fragment B4) lacking surrounding flanking sequences (fragment B3 and B5) of the enhancer. Together these experiments show that the chicken delta-crystallin enhancers show lens-preference in transgenic mice despite the absence of delta-crystallin in this species and add retinoic acid nuclear receptors to the growing list of transcription factors (including Pax-6, Sox-2, and delta EF3) that directly or indirectly contribute to the high expression of the delta 1-crystallin gene in the lens.
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PMID:Lens-preferred activity of chicken delta 1- and delta 2-crystallin enhancers in transgenic mice and evidence for retinoic acid-responsive regulation of the delta 1-crystallin gene. 921 65

The B cell-specific transcription factor Pax-5 has been shown previously to interact with the promoter of the blk gene in vitro. blk encodes a tyrosine kinase associated with the B cell receptor, which is expressed during the early but not the final stages of B cell development. To investigate whether Pax-5 regulates expression of the blk gene in vivo during B cell development and/or activation, Pax-5a was overexpressed in B cell lines. Increases in blk promoter activity using a chloramphenicol acetyltransferase reporter gene system suggested a role for Pax-5a as a transcriptional activator. Subsequent site-specific mutagenesis studies showed that mutations of the Pax-5 binding site on blk significantly alter promoter activity, although results suggested that other factors could bind to this region as well. Using mobility shift assays, we detected an inducible transcription factor that interacts strongly with a sequence overlapping the Pax-5 site on the blk promoter and identified this as a homodimer of NF-kappaB/p50, a member of the NF-kappaB/Rel family of transcription factors. This factor was present at high levels in lipopolysaccharide-activated normal B cells and in plasma cell lines but either at low levels or undetectable levels in resting normal B cells or pre-B or mature B cell lines. In contrast, lipopolysaccharide induction of a pre-B cell line (703/Z) induced a complex that contained both NF-kappaB/p50 and p65. These studies suggest that different NF-kappaB complexes are able to interact with a sequence overlapping the Pax-5 site on the blk promoter and that the relative levels of "bound" factor influence levels of blk expression. Since p50 homodimers and p50/p65 heterodimers of the NF-kappaB complex should have opposing effects on blk transcription, this could provide a mechanism to differentially regulate blk expression during B cell development and activation.
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PMID:The transcription factor NF-kappaB/p50 interacts with the blk gene during B cell activation. 966 Aug 39

It has been demonstrated previously that Pax-6, a paired domain (PD)/homeodomain (HD) transcription factor critical for eye development, contributes to the activation of the alphaB-, alphaA-, delta1-, and zeta-crystallin genes in the lens. Here we have examined the possibility that the inverse relationship between the expression of Pax-6 and beta-crystallin genes within the developing chicken lens reflects a negative regulatory role of Pax-6. Cotransfection of a plasmid containing the betaB1-crystallin promoter fused to the chloramphenicol acetyltransferase reporter gene and a plasmid containing the full-length mouse Pax-6 coding sequences into primary embryonic chicken lens epithelial cells or fibroblasts repressed the activity of this promoter by as much as 90%. Pax-6 constructs lacking the C-terminal activation domain repressed betaB1-crystallin promoter activity as effectively as the full-length protein, but the PD alone or Pax-6 (5a), a splice variant with an altered PD affecting its DNA binding specificity, did not. DNase footprinting analysis revealed that truncated Pax-6 (PD+HD) binds to three regions (-183 to -152, -120 to -48, and -30 to +1) of the betaB1-crystallin promoter. Earlier experiments showed that the betaB1-crystallin promoter sequence from -120 to -48 contains a cis element (PL2 at -90 to -76) that stimulates the activity of a heterologous promoter in lens cells but not in fibroblasts. In the present study, we show by electrophoretic mobility shift assay and cotransfection that Pax-6 binds to PL2 and represses its ability to activate promoter activity; moreover, mutation of PL2 eliminated binding by Pax-6. Taken together, our data indicate that Pax-6 (via its PD and HD) represses the betaB1-crystallin promoter by direct interaction with the PL2 element. We thus suggest that the relatively high concentration of Pax-6 contributes to the absence of betaB1-crystallin gene expression in lens epithelial cells and that diminishing amounts of Pax-6 in lens fiber cells during development allow activation of this gene.
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PMID:Dual roles for Pax-6: a transcriptional repressor of lens fiber cell-specific beta-crystallin genes. 971 Jun 41

Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) is selectively expressed in islet beta cells and is a major autoantigen in a mouse model of type I diabetes. The analysis of IGRP-chloramphenicol acetyltransferase (CAT) fusion gene expression through transient transfection of islet-derived betaTC-3 cells revealed that a promoter region, located between -273 and -254, is essential for high IGRP-CAT fusion gene expression. The sequence of this promoter region does not match that for any known islet-enriched transcription factor. However, data derived from gel retardation assays, a modified ligation-mediated polymerase chain reaction in situ footprinting technique and a SDS-polyacrylamide separation/renaturation procedure led to the hypothesis that this protein might be Pax-6, a conclusion that was confirmed by gel supershift assays. Additional experiments revealed a second non-consensus Pax-6 binding site in the -306/-274 IGRP promoter region. Pax-6 binding to these elements is unusual in that it appears to require both its homeo and paired domains. Interestingly, loss of Pax-6 binding to the -273/ -246 element is compensated by Pax-6 binding to the -306/-274 element and vice versa. Gel retardation assays revealed that another islet-enriched transcription factor, namely Pdx-1, binds four non-consensus elements in the IGRP promoter. However, mutation of these elements has little effect on IGRP fusion gene expression. Although chromatin immunoprecipitation assays show that both Pax-6 and Pdx-1 bind to the IGRP promoter within intact cells, in contrast to the critical role of these factors in beta cell-specific insulin gene expression, IGRP gene transcription appears to require Pax-6 but not Pdx-1.
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PMID:Differential regulation of islet-specific glucose-6-phosphatase catalytic subunit-related protein gene transcription by Pax-6 and Pdx-1. 1518 Sep 90