On a roll for new TRF targets
TL;DR: In this issue of Genes & Development, Isogai et al. (2007a) report that the TATA-less histone H1 promoter is regulated by TRF2, which provides a possible mechanism for earlier observations linking TRF3 with chromatin structure and helps to establish Drosophila TRF 2 as a broadly used core-promoter factor.
Abstract: In the early 1990s, one of us wrote in these pages a review entitled “TBP, a universal transcription factor?” (Hernandez 1993). At the time, it had become clear that the TATA-box-binding protein TBP was not a transcription factor exclusively involved in transcription from RNA polymerase II (pol II) promoters as had been thought before, but rather a factor involved in transcription by all three main types of eukaryotic nuclear RNA polymerases. In retrospect, however, the question mark at the end of the title was a wise touch! Indeed, shortly thereafter, the first TBP-related factor, TRF1, was described (Crowley et al. 1993). Since then, two more TRFs have been discovered (for review, see Berk 2000; Davidson 2003; Hochheimer and Tjian 2003), and it was found that some genes dispense with TBP and TRFs altogether (Wieczorek et al. 1998). This “expansion” of TBP into a TBP family of proteins begs the question of which promoters are targeted by which TBP family member. In this issue of Genes & Development, Isogai et al. (2007a) report that the TATA-less histone H1 promoter is regulated by TRF2. This provides a possible mechanism for earlier observations linking TRF2 with chromatin structure (Martianov et al. 2002; Kopytova et al. 2006). Furthermore, the identification by Isogai et al. (2007a) of a large number of TRF2-bound sites in the Drosophila genome helps to establish Drosophila TRF2 as a broadly used core-promoter factor. Among the three classes of TBP-related factors described so far, TRF2—also called TBP-like protein (TLP) or TBP-like factor (TLF)—is the only one to be widely present in metazoans (Ohbayashi et al. 1999; Kaltenbach et al. 2000; Veenstra et al. 2000). TRF1 has been found only in Drosophila and Anopheles (Crowley et al. 1993; Isogai et al. 2007b), and TRF3 is restricted to vertebrates (Persengiev et al. 2003). All three proteins contain a core domain related to the TBP C-terminal core domain, and some also contain variable Nand C-terminal domains.
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TL;DR: It is demonstrated that TRF2 knockdown regulates cell cycle progression and exerts distinct effects on G1 and specific mitotic phases, uncovering a critical and unanticipated role of a general transcription factor as a key regulator of cell cycle.
Abstract: TRF2 (TATA-box-binding protein-related factor 2) is an evolutionarily conserved general transcription factor that is essential for embryonic development of Drosophila melanogaster, C. elegans, zebrafish and Xenopus. Nevertheless, the cellular processes that are regulated by TRF2 are largely underexplored. Here, using Drosophila Schneider cells as a model, we discovered that TRF2 regulates cell cycle progression. Using flow cytometry, high-throughput microscopy and advanced imaging-flow cytometry, we demonstrate that TRF2 knockdown regulates cell cycle progression and exerts distinct effects on G1 and specific mitotic phases. RNA-seq analysis revealed that TRF2 regulates the expression of Cyclin E and the mitotic cyclins, Cyclin A, Cyclin B and Cyclin B3, but not Cyclin D or Cyclin C. To identify proteins that could account for the observed regulation of these cyclin genes, we searched for TRF2-interacting proteins. Interestingly, mass spectrometry analysis of TRF2-containing complexes identified GFZF, a nuclear glutathione S-transferase implicated in cell cycle regulation, and Motif 1 binding protein (M1BP). Furthermore, available ChIP-exo data revealed that TRF2, GFZF and M1BP co-occupy the promoters of TRF2-regulated genes. Using RNAi to knockdown the expression of either M1BP, GFZF, TRF2 or their combinations, we demonstrate that although GFZF and M1BP interact with TRF2, it is TRF2, rather than GFZF or M1BP, that is the main factor regulating the expression of Cyclin E and the mitotic cyclins. Taken together, our findings uncover a critical and unanticipated role of a general transcription factor as a key regulator of cell cycle.
4 citations
TL;DR: It is shown that TRF2 knockdown results in increased expression of distinct pro-apoptotic genes and induces apoptosis, uncovering a critical and unanticipated role of a general transcription factor as a key regulator of cell cycle and apoptosis.
Abstract: Background Diverse biological processes and transcriptional programs are regulated by RNA polymerase II (Pol II), which is recruited by the general transcription machinery to the core promoter to initiate transcription. TRF2 (TATA-box-binding protein-related factor 2) is an evolutionarily conserved general transcription factor that is essential for embryonic development of Drosophila melanogaster, C. elegans, zebrafish and Xenopus. Nevertheless, the cellular processes that are regulated by TRF2 are largely underexplored. Results Here, using Drosophila Schneider cells as a model, we discovered that TRF2 regulates apoptosis and cell cycle progression. We show that TRF2 knockdown results in increased expression of distinct pro-apoptotic genes and induces apoptosis. Using flow cytometry, high-throughput microscopy and advanced imaging-flow cytometry, we demonstrate that TRF2 regulates cell cycle progression and exerts distinct effects on G1 and specific mitotic phases. RNA-seq analysis revealed that TRF2 controls the expression of Cyclin E and the mitotic cyclins, Cyclin A, Cyclin B and Cyclin B3, but not Cyclin D or Cyclin C. To identify proteins that could account for the observed regulation of these cyclin genes, we searched for TRF2-interacting proteins. Interestingly, mass spectrometry analysis of TRF2-containing complexes identified GFZF, a nuclear glutathione S-transferase implicated in cell cycle regulation, and Motif 1 binding protein (M1BP). TRF2 has previously been shown to interact with M1BP and M1BP has been shown to interact with GFZF. Furthermore, available ChIP-exo data revealed that TRF2, GFZF and M1BP co-occupy the promoters of TRF2-regulated genes. Using RNAi to knockdown the expression of either M1BP, GFZF, TRF2 or their combinations, we demonstrate that although GFZF and M1BP interact with TRF2, it is TRF2, rather than GFZF or M1BP, that is the main factor regulating the expression of Cyclin E and the mitotic cyclins. Conclusions Our findings uncover a critical and unanticipated role of a general transcription factor as a key regulator of cell cycle and apoptosis.
2 citations
Dissertation•
07 Aug 2008
TL;DR: It is shown that the TAP-tagged RNAP II complex contains the hypophosphorylated form of RNAPII and is functionally active both in vitro and in vivo, and a role for RPAPI in the regulation ofRNAP II is proposed.
Abstract: The expression, replication and repair of the human genome involve the concerted action of various molecular machineries which form an intricate network of proteinprotein, protein-DNA and protein-RNA interactions. The first step of the gene expression process, the transcription of genes by RNA polymerase II (RNAP II), is regulated by a myriad of proteins, including the general transcription factors (GTFs). Previous studies have shown that, in the ceIl, these factors assemble with other proteins into complexes able to bind DNA to exert their regulatory role. However, very little is known about the organization of the transcription machinery in the soluble nuclear compartment of the ceIl, prior to or following its association with genomic DNA. In the following thesis, we have programmed HEK 293 cells to express RNAP II and GTFs subunits carrying tandem affinity purification (TAP) tags. In order to define the prote in interaction networks involved in the function of the RNAP II machinery, soluble prote in complexes were purified by double affinity chromatography. By using the Rpb 11-TAP subunit of RNAP II, we isolated for the first time a complex containing RNAP II, the GTFs TFIIB and TFIIF, the RNAP II phosphatase Fcpl, and a novel polypeptide of unknown function that we named RNA polymerase II-associated protein 1 (RPAP1). We showed that the TAP-tagged RNAP II complex contains the hypophosphorylated form of RNAP II and is functionally active both in vitro and in vivo. In addition, we have proposed a role for RPAPI in the regulation ofRNAP II. Our strategy for the purification of prote in complexes coupled to mass spectrometry allowed for the simple and efficient survey of soluble human prote in complexes containing components of the transcription machinery. Thirty-two tagged polypeptides yielded a network of 805 high-confidence interactions. Of note, aIl the tagged transcription factors that we purified and tested were active in transcription reactions in vitro and were found in association with transcribed regions using chromatin immunoprecipitation (ChIP) experiments in vivo, indicating that bon a fide, functional
2 citations
Dissertation•
01 Jan 2014
1 citations
Cites background from "On a roll for new TRF targets"
...encoding for TBP-related factors (TRFs) and variant forms of TAFs in higher eukaryotes [186]....
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TL;DR: A novel characteristic of TLP is shown, that is, interaction with TFIIA is essential to suppress proteasome‐dependent turnover of T LP, providing a further insight into TLP‐governed gene regulation.
Abstract: Although the majority of gene expression is driven by TATA-binding protein (TBP)-based transcription machinery, it has been reported that TBP-related factors (TRFs) are also involved in the regulation of gene expression. TBP-like protein (TLP), which is one of the TRFs and exhibits the highest affinity to TFIIA among known proteins, has recently been showed to have significant roles in gene regulation. However, how the level of TLP is maintained in vivo has remained unknown. In this study, we explored the mechanism by which TLP protein is turned over in vivo and the factor that maintains the amount of TLP. We showed that TLP is rapidly degraded by the ubiquitin-proteasome system and that tight interaction with TFIIA results in protection of TLP from ubiquitin-proteasome-dependent degradation. The half-life of TLP was shown to be less than a few hours, and the proteasome inhibitor MG132 specifically suppressed TLP degradation. Moreover, knockdown and over-expression experiments showed that TFIIA is engaged in stabilization of TLPin vivo. Thus, we showed a novel characteristic of TLP, that is, interaction with TFIIA is essential to suppress proteasome-dependent turnover of TLP, providing a further insight into TLP-governed gene regulation.
1 citations
Cites background from "On a roll for new TRF targets"
...TBP-related factors (TRFs), TRF1, TRF2 [also called TLP (TBP-like protein)] and TRF3, have also been shown to participate in transcription regulation (Reina & Hernandez 2007; Zehavi et al. 2015)....
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...Therefore, TLP has no TATA box-binding ability, whereas its affinity to TFIIA and TFIIB is preserved like other TRFs (Ohbayashi et al. 1999; Berk 2000; Reina & Hernandez 2007; Zehavi et al. 2015)....
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References
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TL;DR: Analyzing gene-expression patterns by in situ hybridization to whole-mount embryos provides an extremely rich dataset that can be used to identify genes involved in developmental processes that have been missed by traditional genetic analysis.
Abstract: Background: Cell-fate specification and tissue differentiation during development are largely achieved by the regulation of gene transcription. Results: As a first step to creating a comprehensive atlas of gene-expression patterns during Drosophila embryogenesis, we examined 2,179 genes by in situ hybridization to fixed Drosophila embryos. Of the genes assayed, 63.7% displayed dynamic expression patterns that were documented with 25,690 digital photomicrographs of individual embryos. The photomicrographs were annotated using controlled vocabularies for anatomical structures that are organized into a developmental hierarchy. We also generated a detailed time course of gene expression during embryogenesis using microarrays to provide an independent corroboration of the in situ hybridization results. All image, annotation and microarray data are stored in publicly available database. We found that the RNA transcripts of about 1% of genes show clear subcellular localization. Nearly all the annotated expression patterns are distinct. We present an approach for organizing the data by hierarchical clustering of annotation terms that allows us to group tissues that express similar sets of genes as well as genes displaying similar expression patterns. Conclusions: Analyzing gene-expression patterns by in situ hybridization to whole-mount embryos provides an extremely rich dataset that can be used to identify genes involved in developmental processes that have been missed by traditional genetic analysis. Systematic analysis of rigorously annotated patterns of gene expression will complement and extend the types of analyses carried out using expression microarrays.
740 citations
"On a roll for new TRF targets" refers background in this paper
...Interestingly, a computational analysis identified the DRE as one of the prevalent core-promoter motifs in the Drosophila genome, raising the possibility that the TRF2/DREF-containing complex controls a large group of promoters (Ohler et al. 2002; Tomancak et al. 2002)....
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TL;DR: A key step in retrieving the information stored in the complex genomes of eukaryotes involves the identification of transcription units and, more specifically, the recognition of promoter sequences by RNA polymerase.
Abstract: A key step in retrieving the information stored in the complex genomes of eukaryotes involves the identification of transcription units and, more specifically, the recognition of promoter sequences by RNA polymerase. In eukaryotes, the task of recognizing nuclear gene promoters and then transcribing the genes is divided among three highly related enzymes, RNA polymerases I, II, and III. Each of these RNA polymerases is dedicated to the transcription of specific sets of genes, and each depends on accessory factors, the so-called transcription factors, to recognize its cognate promoter sequences.
601 citations
"On a roll for new TRF targets" refers background in this paper
...In Drosophila cells, then, the TFIIIB activity, which is the key factor responsible for pol III recruitment (for a review, see Schramm and Hernandez 2002), contains Brf1 and TRF1, whereas in yeast it contains Brf1 and TBP....
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583 citations
Additional excerpts
...(Hernandez 1993)....
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TL;DR: Results indicate that linker histones can participate in epigenetic regulation of gene expression by contributing to the maintenance or establishment of specific DNA methylation patterns.
551 citations
"On a roll for new TRF targets" refers background in this paper
...Indeed, the observation that TRF2 depletion leads to both reduced amounts of histone H1 and altered polytene chromosomes provides further evidence that varying levels of histone H1 can have dramatic effects on chromatin structure (Fan et al. 2005)....
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TL;DR: The author’s views are based on personal experience, research, and interviews conducted at the 2016 USGS workshop on “Biology of infectious disease: Foundations of Natural Selection and Response to infectious disease .”
Abstract: AND PERSPECTIVE. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 827 ORGANIZATION OF HISTONE GENES-STRUCTURE OF HISTONE mRNAs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 828 OVERVIEW OF HIS TONE SYNTHESIS IN THE CELL CyCLE . . . . . . . . . . . . . . . . . . . . . . . . 829 TRANSCRIPTIONAL REGULATION . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 832 Higher Eukaryotes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ... . . . . . . . . . . . . . . . ... . . . . . . . . . . . . . . . . ... . . . . . . . . 832 Lower Eukaryotes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ..... . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . 841 POSTTRANSCRIPTIONAL REGULATION . . . . . .. . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . .. . . . . . . . .... 847 Higher Eukaryotes. . . . . .. . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ....... . . . . . . . . . . . . . . . .. . . . . . . . 847 Lower Eukaryotes . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ...... . . . . . . . . . . . . . . . . . . 853 MULTIPLE FORMS OF REGULATION MODULATE HISTONE mRNA LEVELS IN THE CELL CyCLE . . . . . . . . . . . . . . . . . . . ... . . . . . . . . . . . . . . . . . . . !l55 FUTURE PROSPECTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ..... . . .. . . . . . . . . . . . . . . . . . ... .. . . . . .. . . . . . . . . . 856
510 citations
"On a roll for new TRF targets" refers background in this paper
...In higher eukaryotes the organization is different in that the histone genes are more dispersed even though some clusters occur, and the number of copies per haploid genome is lower (for review, see Osley 1991)....
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