Reversal of the silencing of tetracycline‐controlled genes requires the coordinate action of distinctly acting transcription factors

Pankiewicz, Renata; Karlen, Yann; Imhof, Markus O.; Mermod, Nicolas

doi:10.1002/jgm.644

Cited by 29 publications

(35 citation statements)

References 56 publications

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“…Moreover, mutations of the TRD domain that inhibit H3 interaction correspondingly decrease the ability of CTF-1 to achieve chromatin opening in mammalian cells (58). Irrespective of the precise molecular mechanism involved, these data give new insight into how the anti-silencing and boundary activities of CTF-1 may participate to its transcriptional activation function.…”

Section: Discussionmentioning

confidence: 91%

“…Alternatively, a variety of transcription activators have been further shown to alter chromatin structure through the recruitment of chromatin remodeling/modifying complexes (56,57). CTF-1 rather possesses a histone binding activity, which has been implicated in CTF-1 anti-silencing activity in mammalian cells (58). The GAL4 fusion proteins studied here support this conclusion, as the histone H3 binding activities of mutant proteins correlate well with transcriptional activation in a chromatin context, as illustrated by the boundary effect, but not with the transcriptional activation potential detected in the absence of native chromatin structure in transient assays.…”

Section: Discussionmentioning

confidence: 99%

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Chromatin Domain Boundaries Delimited by a Histone-binding Protein in Yeast

Ferrari

Simmen

Dusserre

et al. 2004

Journal of Biological Chemistry

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When located next to chromosomal elements such as telomeres, genes can be subjected to epigenetic silencing. In yeast, this is mediated by the propagation of the SIR proteins from telomeres toward more centromeric regions. Particular transcription factors can protect downstream genes from silencing when tethered between the gene and the telomere, and they may thus act as chromatin domain boundaries. Here we have studied one such transcription factor, CTF-1, that binds directly histone H3. A deletion mutagenesis localized the barrier activity to the CTF-1 histone-binding domain. A saturating point mutagenesis of this domain identified several amino acid substitutions that similarly inhibited the boundary and histone binding activities. Chromatin immunoprecipitation experiments indicated that the barrier protein efficiently prevents the spreading of SIR proteins, and that it separates domains of hypoacetylated and hyperacetylated histones. Together, these results suggest a mechanism by which proteins such as CTF-1 may interact directly with histone H3 to prevent the propagation of a silent chromatin structure, thereby defining boundaries of permissive and silent chromatin domains.The expression state of a eukaryotic gene depends in part on its location in the chromosome. This position effect results from the organization of eukaryotic genomes into discrete functional domains, defined by local differences in chromatin structure. The expression of genes within each domain appears to be defined and maintained by the concerted action of regulatory elements such as promoters, enhancers, silencers, and locus control regions. Individual domains may be bordered by boundary elements that separate regions of permissive and silent chromatin. Experimentally, boundary elements have been defined functionally by their ability to protect against position effect when flanking the assayed gene. Examples of boundary elements in metazoans include the subtelomeric anti-silencing regions at yeast telomeres, the scs and scsЈ elements flanking the 87A1 hsp70 locus in Drosophila, and the chicken ␤-globin boundary elements (1-4). Several proteins have been associated with barrier activities in yeast, Drosophila, and mammalian cells (5).Transcriptional repression of the yeast Saccharomyces cerevisiae subtelomeric regions is one of the best-studied examples of position-dependent gene expression. This phenomenon is referred to as telomere position effect and is similar to the position effect variegation (PEV) initially described in Drosophila (6). Transcriptional silencing at yeast telomeres is associated with a heterochromatin-like structure (7,8). DNA wrapped in transcriptionally silent chromatin replicates late in S phase and is refractory to some modifying agents. In addition, nucleosomes have reduced acetylation compared with nucleosomes from active region of the genome (9, 10).Transcriptional silencing at subtelomeric regions is mediated by a multiprotein nucleosome binding complex called the SIR complex, composed of the Sir2, Sir3, and...

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Section: Discussionmentioning

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Chromatin Domain Boundaries Delimited by a Histone-binding Protein in Yeast

Ferrari

Simmen

Dusserre

et al. 2004

Journal of Biological Chemistry

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“…This observation is consistent with previous studies where CTF/NF1 proteins act as barrier elements that block the propagation of silent chromatin structures, and thus protect transgenes from silencing effects mediated by repressive chromatin structures. 31,32,43 Hence, CTF/NF1 binding sites act both as enhancer-blockers and as barrier insulator elements. Maintenance of a euchromatic status of the provirus should favor the production of retroviral vectors.…”

Section: Discussionmentioning

confidence: 99%

“…30 The CAAT box-binding transcription factor/nuclear factor-1 (NF1, also called CTF/NF1) consists of a family of widely expressed transcription factors that possess a barrier function. 31,32 This family comprises NF1-A, NF1-B, NF1-C and NF1-X subtypes that share the same DNA-binding domain. 33,34 Of note, the NF1-C regulatory domain prevents the propagation of repressive chromatin structures that stem from the telomere, and thereby prevent transgene silencing at mammalian and yeast cells telomeres.…”

Section: Introductionmentioning

confidence: 99%

“…33,34 Of note, the NF1-C regulatory domain prevents the propagation of repressive chromatin structures that stem from the telomere, and thereby prevent transgene silencing at mammalian and yeast cells telomeres. 31,32 NF1-C regulates DNA replication by promoting the recruitment of the DNA polymerase to the adenovirus and SV40 origins of replication, and is also involved in promoter regulation. [35][36][37] Thus, CTF/NF1 proteins were described as barrier elements that act to control DNA transcription and replication, yet a potential enhancer-blocking function had not been assessed.…”

Section: Introductionmentioning

confidence: 99%

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CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

Gaussin

Modlich

Bauche³

et al. 2011

Gene Ther

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Gene transfer-based therapeutic approaches have greatly benefited from the ability of some viral vectors to efficiently integrate within the cell genome and ensure persistent transmission of newly acquired transgenes to the target cell progeny. However, integration of provirus has been associated with epigenetic repercussions that may influence the expression of both the transgene and cellular genes close to vector integration loci. The exploitation of genetic insulator elements may overcome both issues through their ability to act as barriers that limit transgene silencing and/or as enhancer-blockers preventing the activation of endogenous genes by the vector enhancer. We established quantitative plasmid-based assay systems to screen enhancerblocker and barrier genetic elements. Short synthetic insulators that bind to nuclear factor-I protein family transcription factors were identified to exert both enhancer-blocker and barrier functions, and were compared to binding sites for the insulator protein CTCF (CCCTC-binding factor). Gamma-retroviral vectors enclosing these insulator elements were produced at titers similar to their non-insulated counterparts and proved to be less genotoxic in an in vitro immortalization assay, yielding lower activation of Evi1 oncogene expression and reduced clonal expansion of bone marrow cells.

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A siRNA system based on HSP70 promoter results in controllable and powerful gene silencing by heat‐induction

Liao

Feng

Qian

et al. 2013

Biotechnology Progress

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RNAi is a powerful tool for gene-specific knockdown and gene therapy. However, the imprecise expression of siRNA limits the extensive application of RNAi in gene therapy. Here we report the development of a novel controllable siRNA expression vector pMHSP70psil that is initiated by HSP70 promoter. We determined the efficiency of the controllable siRNA system by targeting the gama-synuclein (SNCG) gene in breast cancer cells MCF-7. The results show that the controllable siRNA system can be induced to initiate siRNA expression by heat-induction. The silencing effect of SNCG occurs at a relatively low level (10.1%) at 37°C, while it is significantly increased to 69.4% after heat induction at 43°C. The results also show that the controllable siRNA system inhibits proliferation of cancer cells by heat-shock. Therefore, this RNAi strategy holds the promise of the high efficiency in gene knockdown at targeted times and locations, avoiding systemic side effects. It provides, for the first time, an approach to control siRNA expression by heat-shock.

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Reversal of the silencing of tetracycline‐controlled genes requires the coordinate action of distinctly acting transcription factors

Cited by 29 publications

References 56 publications

Chromatin Domain Boundaries Delimited by a Histone-binding Protein in Yeast

Chromatin Domain Boundaries Delimited by a Histone-binding Protein in Yeast

CTF/NF1 transcription factors act as potent genetic insulators for integrating gene transfer vectors

A siRNA system based on HSP70 promoter results in controllable and powerful gene silencing by heat‐induction

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