En masse nascent transcription analysis to elucidate regulatory transcription factors

Fan, Jinshui; Zhan, Ming; Shen, Jikui; Martindale, Jennifer L.; Yang, Xiaoling; Kawai, Tomoko; Gorospe, Myriam

doi:10.1093/nar/gkj510

Cited by 13 publications

(12 citation statements)

References 34 publications

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“…One study used low pH as a stress agent and identified selective Oct-binding site enrichment in a large group of coordinately up-regulated mRNAs (Duggan et al 2006). Another study identified sites in >90% of genes, with significant changes in transcription rate following exposure of HeLa cells to camptothecin (Fan et al 2006). Oct1 also appears to be a mediator of the response to heavy metals (Glahn et al 2008).…”

Section: Genes and Development 217mentioning

confidence: 99%

A general mechanism for transcription regulation by Oct1 and Oct4 in response to genotoxic and oxidative stress

Kang¹,

Gemberling²,

Nakamura³

et al. 2009

Genes Dev.

118

150

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Oct1 and Oct4 are homologous transcription factors with similar DNA-binding specificities. Here we show that Oct1 is dynamically phosphorylated in vivo following exposure of cells to oxidative and genotoxic stress. We further show that stress regulates the selectivity of both proteins for specific DNA sequences. Mutation of conserved phosphorylation target DNA-binding domain residues in Oct1, and Oct4 confirms their role in regulating binding selectivity. Using chromatin immunoprecipitation, we show that association of Oct4 and Oct1 with a distinct group of in vivo targets is inducible by stress, and that Oct1 is essential for a normal post-stress transcriptional response. Finally, using an unbiased Oct1 target screen we identify a large number of genes targeted by Oct1 specifically under conditions of stress, and show that several of these inducible Oct1 targets are also inducibly bound by Oct4 in embryonic stem cells following stress exposure. Oct1 and Oct4 (products of the Pou2f1 and Pou5f1 genes) are members of the POU (Pit-1, Oct1/2, Unc-86) domain transcription factor family (Herr et al. 1988;Ryan and Rosenfeld 1997). This family is defined by the presence of a bipartite DNA-binding domain in which two subdomains, covalently connected by a flexible linker, typically recognize DNA through major groove interactions on opposite sides of the helix (Klemm et al. 1994). The classical DNA recognition sequence is known as an octamer motif (59-ATGCAAAT-39, hereafter called a ''simple'' octamer). However, we demonstrated recently that native binding sites for Oct4 frequently exist in complex paired, overlapping, and nonconsensus configurations (Tantin et al. 2008).Oct4 is a master regulator of the stem cell state and has recently been shown to be one of three proteins sufficient to reprogram differentiated adult mouse and human cells to the embryonic stem (ES) cell lineage (Okita et al. 2007;Takahashi et al. 2007;Nakagawa et al. 2008). The biological function of Oct1 is more enigmatic. Oct1 is known to interact with regulatory sites in interleukin, immunoglobulin, and histone genes (Garrity et al. 1994;Zheng et al. 2003;Ushmorov et al. 2004;Murayama et al. 2006). Oct1 also moderately stimulates gene expression reporter constructs linked to target sequences in transient transfection assays (Sive et al. 1986;LeBowitz et al. 1988). However, we showed that Oct1 is nonessential for native H2B, IgH, and Igk expression (V.E. V.E.H. Wang et al. 2004). Oct1-deficient cells appear morphologically normal in light microscopy and divide at normal rates. Oct1-deficient mice die in mid-late gestation (embryonic days 12,5-18.5 [E12.5-E18.5]) (V.E.H. .We determined previously that Oct1 À/À mouse embryonic fibroblasts (MEFs) are hypersensitive to oxidative and genotoxic stress (Tantin et al. 2005). One explanation for this result is that constitutive products of Oct1-mediated transcription participate in stress response pathways. Support for an alternative hypothesis, namely that Oct1 directly senses cellular stress, comes from the f...

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Section: Genes and Development 217mentioning

confidence: 99%

A general mechanism for transcription regulation by Oct1 and Oct4 in response to genotoxic and oxidative stress

Kang¹,

Gemberling²,

Nakamura³

et al. 2009

Genes Dev.

118

150

View full text Add to dashboard Cite

show abstract

“…Since 2002, several genomic upgrades of TRO have appeared for different organisms. The most comprehensive studies were published by the groups of Myriam Gorospe [12,13] and Jack D. Keene [14] in mammalian cells, and by Legen et al in plants [15]. In 2004 [16], a genomic upgrade for the yeast S. cerevisiae called "genomic run-on" (GRO) was developed.…”

Section: Methods To Measure Rna Polymerase Density: the Nascent Trmentioning

confidence: 99%

Genome-wide studies of mRNA synthesis and degradation in eukaryotes

Pérez-Ortín

Miguel-Jiménez

2012

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanism

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Keywords:Transcription rate mRNA turnover RNA polymerase II Transcription elongation mRNA stabilityIn recent years, the use of genome-wide technologies has revolutionized the study of eukaryotic transcription producing results for thousands of genes at every step of mRNA life. The statistical analyses of the results for a single condition, different conditions, different transcription stages, or even between different techniques, is outlining a totally new landscape of the eukaryotic transcription process. Although most studies have been conducted in the yeast Saccharomyces cerevisiae as a model cell, others have also focused on higher eukaryotes, which can also be comparatively analyzed. The picture which emerges is that transcription is a more variable process than initially suspected, with large differences between genes at each stage of the process, from initiation to mRNA degradation, but with striking similarities for functionally related genes, indicating that all steps are coordinately regulated. This article is part of a Special Issue entitled: Nuclear Transport and RNA Processing.

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“…For construction of plasmids for RNA interference against RFX1, we selected two target sites spanning nucleotides 2021-2039 and 1657-1675 in the mouse Rfx1 mRNA (GenBank TM accession number X76088) (22) and designated them RFX1-siRNA1 2 and RFX1-siRNA2, respectively. RFX1-siRNA1 was designed according a previous report (25). Forward and reverse oligonucleotides corresponding to these sites were synthesized, annealed, and subcloned into the pSUPER.puro vector (OligoEngine, Seattle, WA), resulting in pSUPER.puro-RFX1-1 and pSUPER.puro-RFX1-2, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…For this, two siRNAs (RFX1-siRNA1 and RFX1-siRNA2) were designed according to a previous report (25), and the respective short hairpin RNA vectors were constructed based on pSUPER.puro. Their efficacies were confirmed at the RNA and protein levels by transiently expressing them in NIH3T3 cells.…”

Section: Reduction Of Rfx1 Leads To the Impaired Immediate Early Respmentioning

confidence: 99%

RFX1 Mediates the Serum-induced Immediate Early Response of Id2 Gene Expression

Wang

Nemoto

Yokota

2007

Journal of Biological Chemistry

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En masse nascent transcription analysis to elucidate regulatory transcription factors

Cited by 13 publications

References 34 publications

A general mechanism for transcription regulation by Oct1 and Oct4 in response to genotoxic and oxidative stress

A general mechanism for transcription regulation by Oct1 and Oct4 in response to genotoxic and oxidative stress

Genome-wide studies of mRNA synthesis and degradation in eukaryotes

RFX1 Mediates the Serum-induced Immediate Early Response of Id2 Gene Expression

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