A Triplex-forming Sequence from the Human c-MYC Promoter Interferes with DNA Transcription

Belotserkovskii, Boris P.; Silva, Erandi De; Tornaletti, Silvia; Wang, Guliang; Vásquez, Karen M.; Hanawalt, Philip C.

doi:10.1074/jbc.m704618200

Cited by 128 publications

(124 citation statements)

References 44 publications

(16 reference statements)

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“…We have recently reported that an S1-sensitive element from the c-MYC promoter, which has the potential to form either a H-DNA or a quadruplex structure, blocks T7 RNA polymerase in an in vitro assay, resulting in partial transcription product arrest. Further, various nucleotide substitutions were introduced to specifically destabilize either the triplex structure or the quadruplex structure, and the result suggested that the triplex structure, but not quadruplex, was responsible for the transcription arrest [155].…”

Section: H-dna Is Implicated In Transcription Regulationmentioning

confidence: 99%

DNA triple helices: Biological consequences and therapeutic potential

Jain

Wang²,

Vásquez³

2008

Biochimie

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262

199

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DNA structure is a critical element in determining its function. The DNA molecule is capable of adopting a variety of non-canonical structures, including three-stranded (i.e. triplex) structures, which will be the focus of this review. The ability to selectively modulate the activity of genes is a longstanding goal in molecular medicine. DNA triplex structures, either intermolecular triplexes formed by binding of an exogenously applied oligonucleotide to a target duplex sequence, or naturally occurring intramolecular triplexes (H-DNA) formed at endogenous mirror repeat sequences, present exploitable features that permit site-specific alteration of the genome. These structures can induce transcriptional repression and site-specific mutagenesis or recombination. Triplex-forming oligonucleotides (TFOs) can bind to duplex DNA in a sequence specific fashion with high affinity, and can be used to direct DNA-modifying agents to selected sequences. H-DNA plays important roles in vivo and is inherently mutagenic and recombinogenic, such that elements of the H-DNA structure may be pharmacologically exploitable. In this review we discuss the biological consequences and therapeutic potential of triple helical DNA structures. We anticipate that the information provided will stimulate further investigations aimed toward improving DNA triplexrelated gene targeting strategies for biotechnological and potential clinical applications.

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Section: H-dna Is Implicated In Transcription Regulationmentioning

confidence: 99%

DNA triple helices: Biological consequences and therapeutic potential

Jain

Wang²,

Vásquez³

2008

Biochimie

Self Cite

262

199

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“…Although the in vivo existence of DNA:RNA triplexes and their role in epigenetic programming still need to be validated, our study provides novel mechanistic insights into the mode of action of ncRNA in chromatin-based processes, implicating a direct and possibly widespread role of RNA:DNA structures in epigenetic regulation. In support of this view, triple-helix target sites were found to be overrepresented at human gene promoters, highlighting the potential of naturally occurring hydrogen bond interactions of ncRNA with the major groove of DNA to control gene expression (Goñ i et al 2004;Belotserkovskii et al 2007). A triplex-mediated mechanism of DNA methylation and gene silencing is particularly intriguing considering that the vast majority of the transcriptional output of the human genome represents ncRNA (Prasanth and Spector 2007;Bü hler 2009;Mattick 2009).…”

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confidence: 78%

Interaction of noncoding RNA with the rDNA promoter mediates recruitment of DNMT3b and silencing of rRNA genes

Schmitz¹,

Mayer²,

Postepska³

et al. 2010

Genes Dev.

449

390

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Noncoding RNAs are important components of regulatory networks controlling the epigenetic state of chromatin. We analyzed the role of pRNA (promoterassociated RNA), a noncoding RNA that is complementary to the rDNA promoter, in mediating de novo CpG methylation of rRNA genes (rDNA). We show that pRNA interacts with the target site of the transcription factor TTF-I, forming a DNA:RNA triplex that is specifically recognized by the DNA methyltransferase DNMT3b. The results reveal a compelling new mechanism of RNA-dependent DNA methylation, suggesting that recruitment of DNMT3b by DNA:RNA triplexes may be a common and generally used pathway in epigenetic regulation.Supplemental material is available at http://www.genesdev.org.Received April 28, 2010; revised version accepted August 26, 2010. Noncoding RNAs (ncRNAs) play a profound and complex role in regulating gene expression (Goodrich and Kugel 2009). While the mechanistic details of how RNA and chromatin are connected remain unclear, there is increasing evidence that epigenetic regulation likely represents a balanced interplay of both RNA and chromatin fields (Bernstein and Allis 2005;Mattick 2009). Previous studies have established that ncRNA plays a key role in epigenetic silencing of rRNA genes. Mammalian genomes contain several clusters of tandemly arrayed rRNA genes (rDNA), of which only a subset is transcribed. Silent rDNA repeats are marked by heterochromatic histone modifications and CpG methylation of the rDNA promoter . The establishment and maintenance of the heterochromatic state is mediated by NoRC (Strohner et al. 2001), a chromatin remodeling complex comprising the ATPase SNF2h and a large subunit, termed TIP5 (TTF-Iinteracting protein #5). NoRC silences rRNA genes by recruiting enzymes that mediate heterochromatin formation and silencing (Zhou et al. 2002;Zhou and Grummt 2005). Transcriptional silencing involves DNA methylation at a critical CpG residue (CpG-133) within the upstream control element (UCE) of the rDNA promoter, thereby impairing binding of the transcription factor UBF and abrogating transcription complex formation . Importantly, NoRC function requires the association of TIP5 with RNA that originates from an RNA polymerase I (Pol I) promoter located in the intergenic spacer ;2 kb upstream of the pre-rRNA transcription start site (Mayer et al. 2006). These intergenic transcripts are of low abundance and usually do not accumulate in vivo because they are rapidly degraded or processed into 150-to 250-nucleotide (nt) RNAs that are shielded from degradation by binding to NoRC. These NoRC-associated transcripts are termed pRNA (for promoter-associated RNA), as their sequence matches the rDNA promoter. TIP5 recognizes the secondary structure of pRNA, and the interaction of TIP5 with pRNA changes the structure of both pRNA and NoRC in an induced fit mechanism (Mayer et al. 2008). Antisensemediated depletion of pRNA leads to nucleoplasmic distribution of NoRC, hypomethylation of rDNA, and enhanced Pol I transcription. ''RNA refeeding'' a...

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“…Purification of transcription substrates was as described in ref. 31. Unless otherwise stated, in transcription experiments with linear DNA we used plasmid templates linearized by Dra III restriction digestion 0.96 kb downstream from the promoter.…”

Section: Methodsmentioning

confidence: 99%

Mechanisms and implications of transcription blockage by guanine-rich DNA sequences

Belotserkovskii

Liu

Tornaletti

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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135

143

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Various DNA sequences that interfere with transcription due to their unusual structural properties have been implicated in the regulation of gene expression and with genomic instability. An important example is sequences containing G-rich homopurinehomopyrimidine stretches, for which unusual transcriptional behavior is implicated in regulation of immunogenesis and in other processes such as genomic translocations and telomere function. To elucidate the mechanism of the effect of these sequences on transcription we have studied T7 RNA polymerase transcription of G-rich sequences in vitro. We have shown that these sequences produce significant transcription blockage in an orientation-, lengthand supercoiling-dependent manner. Based upon the effects of various sequence modifications, solution conditions, and ribonucleotide substitutions, we conclude that transcription blockage is due to formation of unusually stable RNA/DNA hybrids, which could be further exacerbated by triplex formation. These structures are likely responsible for transcription-dependent replication blockage by G-rich sequences in vivo.R-loops | DNA supercoiling | Hoogsteen base pairing | inosine | 7-deazaquanosine S equence-specific modulation of transcription, including transcription blockage or impediment, plays an important role in DNA transactions, for example, transcription-related mutagenesis and recombination (reviewed in refs. 1 and 2) and could also be responsible for several severe genetic diseases (reviewed in refs. 3-5).Among the DNA sequences that could affect transcription are GC-rich homopurine-homopyrimidine (hPu/hPy) stretches. These sequences could form unusual DNA structures, including triplexes and G quadruplexes (reviewed in refs. 3-5), which have been implicated in several transcription-dependent phenomena (for example, see refs. 6-9).Another important property of these sequences is a dramatic asymmetry in the stabilities of RNA/DNA duplexes: The rPu/dPy duplex is significantly more stable, whereas the rPy/dPu duplex is less stable than a DNA/DNA duplex of the same sequence (10). The increased stability of rPu/dPy duplexes is likely responsible for stable R-loop formation by these sequences (11), although alternative DNA structures might also be involved (8,12,13).The simplest example of GC-rich hPu/hPy sequences, the G n ∕C n repeats, is abundant in various genomes, including transcribed domains (14, 15).The G 32 ∕C 32 stretch was previously shown to stall DNA replication in Escherichia coli plasmids in vivo (16). Remarkably, this effect was observed only when the sequence was transcribed, which led to a model stipulating that this sequence stalled an elongating RNA polymerase, and the stalled transcription complex, in turn, blocked the replication machinery (16).To elucidate the mechanism of transcription blockage by this sequence, we have studied its effect on T7 RNA polymerase (T7 RNAP) transcription in vitro, using various sequence modifications and solution conditions that allowed us to discriminate between possible D...

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A Triplex-forming Sequence from the Human c-MYC Promoter Interferes with DNA Transcription

Cited by 128 publications

References 44 publications

DNA triple helices: Biological consequences and therapeutic potential

DNA triple helices: Biological consequences and therapeutic potential

Interaction of noncoding RNA with the rDNA promoter mediates recruitment of DNMT3b and silencing of rRNA genes

Mechanisms and implications of transcription blockage by guanine-rich DNA sequences

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