scholarly journals Dissecting transcriptional amplification by MYC

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Zuqin Nie ◽  
Chunhua Guo ◽  
Subhendu K Das ◽  
Carson C Chow ◽  
Eric Batchelor ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Large Scale ◽  
Statistical Parameter ◽  
Genome Wide ◽  
E Box ◽  
Amplifier Gain ◽  
Series Of Experiments ◽  
E Boxes ◽  
Transcriptional Amplification ◽  
Theory And Experiments

Supraphysiological MYC levels are oncogenic. Originally considered a typical transcription factor recruited to E-boxes (CACGTG), another theory posits MYC a global amplifier increasing output at all active promoters. Both models rest on large-scale genome-wide ”-omics’. Because the assumptions, statistical parameter and model choice dictates the ‘-omic’ results, whether MYC is a general or specific transcription factor remains controversial. Therefore, an orthogonal series of experiments interrogated MYC’s effect on the expression of synthetic reporters. Dose-dependently, MYC increased output at minimal promoters with or without an E-box. Driving minimal promoters with exogenous (glucocorticoid receptor) or synthetic transcription factors made expression more MYC-responsive, effectively increasing MYC-amplifier gain. Mutations of conserved MYC-Box regions I and II impaired amplification, whereas MYC-box III mutations delivered higher reporter output indicating that MBIII limits over-amplification. Kinetic theory and experiments indicate that MYC activates at least two steps in the transcription-cycle to explain the non-linear amplification of transcription that is essential for global, supraphysiological transcription in cancer.

scholarly journals Wild-Type Circadian Rhythmicity Is Dependent on Closely Spaced E Boxes in the Drosophila timelessPromoter

2001 ◽  
Vol 21 (4) ◽  
pp. 1207-1217 ◽  
Author(s):  
Michael J. McDonald ◽  
Michael Rosbash ◽  
Patrick Emery
Keyword(s):  
Transcription Factor ◽  
Transcriptional Regulation ◽  
Locomotor Activity ◽  
Circadian Rhythms ◽  
High Amplitude ◽  
Wild Type ◽  
Activity Rhythms ◽  
E Box ◽  
E Boxes ◽  
Circadian Transcription

ABSTRACT Transcriptional regulation plays an important role inDrosophila melanogaster circadian rhythms. The period promoter has been well studied, but the timeless promoter has not been analyzed in detail. Mutagenesis of the canonical E box in the timelesspromoter reduces but does not eliminate timeless mRNA cycling or locomotor activity rhythms. This is because there are at least two other cis-acting elements close to the canonical E box, which can also be transactivated by the circadian transcription factor dCLOCK. These E-box-like sequences cooperate with the canonical E-box element to promote high-amplitude transcription, which is necessary for wild-type rhythmicity.

scholarly journals The MNT transcription factor autoregulates its expression and supports proliferation in MYC-associated factor X (MAX)-deficient cells

2020 ◽  
Vol 295 (7) ◽  
pp. 2001-2017 ◽  
Author(s):  
M. Carmen Lafita-Navarro ◽  
Judit Liaño-Pons ◽  
Andrea Quintanilla ◽  
Ignacio Varela ◽  
Rosa Blanco ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Transcription Factor ◽  
Transcriptional Regulator ◽  
Bhlh Transcription Factor ◽  
Factor X ◽  
Associated Factor ◽  
Helix Loop Helix ◽  
E Box ◽  
E Boxes ◽  
Cycle Regulation

The MAX network transcriptional repressor (MNT) is an MXD family transcription factor of the basic helix-loop-helix (bHLH) family. MNT dimerizes with another transcriptional regulator, MYC-associated factor X (MAX), and down-regulates genes by binding to E-boxes. MAX also dimerizes with MYC, an oncogenic bHLH transcription factor. Upon E-box binding, the MYC–MAX dimer activates gene expression. MNT also binds to the MAX dimerization protein MLX (MLX), and MNT–MLX and MNT–MAX dimers co-exist. However, all MNT functions have been attributed to MNT–MAX dimers, and no functions of the MNT–MLX dimer have been described. MNT's biological role has been linked to its function as a MYC oncogene modulator, but little is known about its regulation. We show here that MNT localizes to the nucleus of MAX-expressing cells and that MNT–MAX dimers bind and repress the MNT promoter, an effect that depends on one of the two E-boxes on this promoter. In MAX-deficient cells, MNT was overexpressed and redistributed to the cytoplasm. Interestingly, MNT was required for cell proliferation even in the absence of MAX. We show that in MAX-deficient cells, MNT binds to MLX, but also forms homodimers. RNA-sequencing experiments revealed that MNT regulates the expression of several genes even in the absence of MAX, with many of these genes being involved in cell cycle regulation and DNA repair. Of note, MNT–MNT homodimers regulated the transcription of some genes involved in cell proliferation. The tight regulation of MNT and its functionality even without MAX suggest a major role for MNT in cell proliferation.

scholarly journals Runx1 Shapes the Chromatin Landscape Via a Cascade of Direct and Indirect Targets

2020 ◽  
Author(s):  
Matthew R. Hass ◽  
Daniel Brisette ◽  
Sreeja Parameswaran ◽  
Mario Pujato ◽  
Omer Donmez ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Large Scale ◽  
Chromatin Accessibility ◽  
Open Chromatin ◽  
Genome Wide ◽  
Related Transcription Factor ◽  
Indirect Consequence ◽  
Genome Scale ◽  
Upregulated Genes ◽  
Transcription Factor 1

AbstractRunt-related transcription factor 1 (Runx1) can act as both an activator and a repressor. Here we show that CRISPR-mediated deletion of Runx1 in an embryonic kidney-derived cell (mK4) results in large-scale genome-wide changes to chromatin accessibility and gene expression. Open chromatin regions near down-regulated loci are enriched for Runx sites, remain bound by Runx2, but lose chromatin accessibility and expression in Runx1 knockout cells. Unexpectedly, regions near upregulated genes are depleted of Runx sites and are instead enriched for Zeb transcription factor binding sites. Re-expressing Zeb2 in Runx1 knockout cells restores suppression. These data confirm that Runx1 activity is uniquely needed to maintain open chromatin at many loci, and demonstrate that genome-scale derepression is an indirect consequence of losing Runx1-dependent Zeb expression.

Transcription factor AP-4 is a ligand for immunoglobulin-κ promoter E-box elements

2001 ◽  
Vol 354 (2) ◽  
pp. 431-438 ◽  
Author(s):  
Alaitz ARANBURU ◽  
Robert CARLSSON ◽  
Christine PERSSON ◽  
Tomas LEANDERSON
Keyword(s):  
Transcription Factor ◽  
Gene Families ◽  
Sequence Motifs ◽  
Mobility Shift ◽  
Conserved Sequence ◽  
Nuclear Extracts ◽  
E Box ◽  
Variable Gene ◽  
E Boxes ◽  
Electrophoretic Mobility Shift Assays

Immunoglobulin (Ig)-κ promoters from humans and mice share conserved sequences. The octamer element is common to all Ig promoters and pivotal for their function. However, other conserved sequence motifs, that differ between Ig variable gene families, are required for normal promoter function. These conserved motifs do not stimulate transcription in the absence of an octamer. One example is an E-box of the E47/E12 type (5′-CAGCTG-3′), which is found in all promoters of the human and murine Ig-κ gene subgroups/families, with the exception of subgroups II and VI and their related murine families. In the present study we show that the ubiquitously expressed transcription factor AP-4, and not E47, interacts specifically with the κ promoter E-boxes when tested in electrophoretic mobility-shift assays using nuclear extracts derived from human and murine B-cell lines. Furthermore, AP-4, unlike E47, did not act as a transactivator, which is in agreement with previous studies on intact κ promoters, showing that transcription is absent when the octamer element has been mutated. Based on these data, and the conservation of the 5′-CAGCTG-3′ motif among human and murine κ promoters, we propose that AP-4 is the major ligand for Ig-κ promoter E-boxes.

scholarly journals CRISPR/Cas9 screen for functional MYC binding sites reveals MYC-dependent vulnerabilities in K562 cells

2021 ◽  
Author(s):  
Marta Kazimierska ◽  
Marta Podralska ◽  
Magdalena Zurawek ◽  
Tomasz Wozniak ◽  
Marta Elzbieta Kasprzyk ◽  
...  
Keyword(s):  
Binding Sites ◽  
Target Genes ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Cancer Type ◽  
Genome Wide ◽  
A Genome ◽  
E Box ◽  
A Cell ◽  
E Boxes

The transcription factor MYC is a proto-oncogene with a well-documented essential role in the pathogenesis and maintenance of several types of cancer. MYC binds to specific E-box sequences in the genome to regulate expression of adjacent genes in a cell type- and developmental stage-specific manner. To date, a comprehensive analysis of direct MYC targets with essential roles in different types of cancer is missing. To enable identification of functional MYC binding sites and corresponding target genes, we designed a CRISPR/Cas9 library to destroy E-box sequences in a genome-wide fashion. As a proof of principle, using this library we identified several known and novel MYC targets critical for K562 chronic myelogenous leukemia cells and uncovered specific features of essential E-boxes. Our unique, well-validated tool opens new possibilities to gain novel insights into MYC-dependent vulnerabilities in any cancer type.

scholarly journals Runx1 shapes the chromatin landscape via a cascade of direct and indirect targets

PLoS Genetics ◽  
2021 ◽  
Vol 17 (6) ◽  
pp. e1009574
Author(s):  
Matthew R. Hass ◽  
Daniel Brissette ◽  
Sreeja Parameswaran ◽  
Mario Pujato ◽  
Omer Donmez ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Large Scale ◽  
Chromatin Accessibility ◽  
Open Chromatin ◽  
Metanephric Mesenchyme ◽  
Genome Wide ◽  
Related Transcription Factor ◽  
Subsequent Activation ◽  
Genome Scale ◽  
Upregulated Genes

Runt-related transcription factor 1 (Runx1) can act as both an activator and a repressor. Here we show that CRISPR-mediated deletion of Runx1 in mouse metanephric mesenchyme-derived mK4 cells results in large-scale genome-wide changes to chromatin accessibility and gene expression. Open chromatin regions near down-regulated loci enriched for Runx sites in mK4 cells lose chromatin accessibility in Runx1 knockout cells, despite remaining Runx2-bound. Unexpectedly, regions near upregulated genes are depleted of Runx sites and are instead enriched for Zeb transcription factor binding sites. Re-expressing Zeb2 in Runx1 knockout cells restores suppression, and CRISPR mediated deletion of Zeb1 and Zeb2 phenocopies the gained expression and chromatin accessibility changes seen in Runx1KO due in part to subsequent activation of factors like Grhl2. These data confirm that Runx1 activity is uniquely needed to maintain open chromatin at many loci, and demonstrate that Zeb proteins are required and sufficient to maintain Runx1-dependent genome-scale repression.

scholarly journals Evaluation of Myc E-Box Phylogenetic Footprints in Glycolytic Genes by Chromatin Immunoprecipitation Assays

2004 ◽  
Vol 24 (13) ◽  
pp. 5923-5936 ◽  
Author(s):  
Jung-whan Kim ◽  
Karen I. Zeller ◽  
Yunyue Wang ◽  
Anil G. Jegga ◽  
Bruce J. Aronow ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Chromatin Immunoprecipitation ◽  
Binding Sites ◽  
Phylogenetic Footprinting ◽  
Regulatory Sequences ◽  
Full Spectrum ◽  
E Box ◽  
Important Regulator ◽  
E Boxes ◽  
Genomic Regions

ABSTRACT Prediction of gene regulatory sequences using phylogenetic footprinting has advanced considerably but lacks experimental validation. Here, we report whether transcription factor binding sites predicted by dot plotting or web-based Trafac analysis could be validated by chromatin immunoprecipitation assays. MYC overexpression enhances glycolysis without hypoxia and hence may contribute to altered tumor metabolism. Because the full spectrum of glycolytic genes directly regulated by Myc is not known, we chose Myc as a model transcription factor to determine whether it binds target glycolytic genes that have conserved canonical Myc binding sites or E boxes (5′-CACGTG-3′). Conserved canonical E boxes in ENO1, HK2, and LDHA occur in 31- to 111-bp islands with high interspecies sequence identity (>65%). Trafac analysis revealed another region in ENO1 that corresponds to a murine region with a noncanonical E box. Myc bound all these conserved regions well in the human P493-6 B lymphocytes. We also determined whether Myc could bind nonconserved canonical E boxes found in the remaining human glycolytic genes. Myc bound PFKM, but it did not significantly bind GPI, PGK1, and PKM2. Binding to BPGM, PGAM2, and PKLR was not detected. Both GAPD and TPI1 do not have conserved E boxes but are induced and bound by Myc through regions with noncanonical E boxes. Our results indicate that Myc binds well to conserved canonical E boxes, but not nonconserved E boxes. However, the binding of Myc to unpredicted genomic regions with noncanonical E boxes reveals a limitation of phylogenetic footprinting. In aggregate, these observations indicate that Myc is an important regulator of glycolytic genes, suggesting that MYC plays a key role in a switch to glycolytic metabolism during cell proliferation or tumorigenesis.

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