The MAP kinase (MAPK) signalling pathways play fundamental roles in a wide range of cellular processes and are often deregulated in disease states. One major mode of action for these pathways is in ...controlling gene expression, in particular through regulating transcription. In this review, we discuss recent significant advances in this area. In particular we focus on the mechanisms by which MAPKs are targeted to the nucleus and chromatin, and once there, how they impact on chromatin structure and subsequent gene regulation. We also discuss how systems biology approaches have contributed to our understanding of MAPK signaling networks, and also how the MAPK pathways intersect with other regulatory pathways in the nucleus. Finally, we summarise progress in studying the physiological functions of key MAPK transcriptional targets.
► Targeting mechanisms of MAPKs to the nucleus and chromatin ► MAPKs in controlling chromatin structure ► Systems analysis of nuclear MAPK signaling networks ► Complexities of MAPK signaling in the nucleus ► Physiological functions of MAPK transcriptional targets
Protein modification by SUMO conjugation is an important regulatory event. Sumoylation usually takes place on a lysine residue embedded in the core consensus motif ψKxE. However, this motif confers ...limited specificity on the sumoylation process. Here, we have probed the roles of clusters of acidic residues located downstream from the core SUMO modification sites in proteins such as the transcription factor Elk‐1. We demonstrate that these are functionally important in SUMO‐dependent transcriptional repression of Elk‐1 transcriptional activity. Mechanistically, the acidic residues are important in enhancing the efficiency of Elk‐1 sumoylation by Ubc9. Similar mechanisms operate in other transcription factors and phosphorylation sites can functionally substitute for acidic residues. Thus, an extended sumoylation motif, termed the NDSM (negatively charged amino acid‐dependent sumoylation motif), helps define functional SUMO targets. We demonstrate that this extended motif can be used to correctly predict novel targets for SUMO modification.
Embryonic stem cells (ESCs) must transition through a series of intermediate cell states before becoming terminally differentiated. Here, we investigated the early events in this transition by ...determining the changes in the open chromatin landscape as naive mouse ESCs transition to epiblast-like cells (EpiLCs). Motif enrichment analysis of the newly opening regions coupled with expression analysis identified ZIC3 as a potential regulator of this cell fate transition. Chromatin binding and genome-wide transcriptional profiling following Zic3 depletion confirmed ZIC3 as an important regulatory transcription factor, and among its targets are genes encoding a number of transcription factors. Among these is GRHL2, which acts through enhancer switching to maintain the expression of a subset of genes from the ESC state. Our data therefore place ZIC3 upstream of a set of pro-differentiation transcriptional regulators and provide an important advance in our understanding of the regulatory factors governing the early steps in ESC differentiation.
Cancer is driven by both genetic and epigenetic changes that impact on gene expression profiles and the resulting tumourigenic phenotype. Enhancers are transcriptional regulatory elements that are ...key to our understanding of how this rewiring of gene expression is achieved in cancer cells. Here, we have harnessed the power of RNA-seq data from hundreds of patients with oesophageal adenocarcinoma (OAC) or its precursor state Barrett's oesophagus coupled with open chromatin maps to identify potential enhancer RNAs and their associated enhancer regions in this cancer. We identify ~1000 OAC-specific enhancers and use these data to uncover new cellular pathways that are operational in OAC. Among these are enhancers for
,
, and
, and we show that their activity is required for cancer cell viability. We also demonstrate the clinical utility of our dataset for identifying disease stage and patient prognosis. Our data therefore identify an important set of regulatory elements that enhance our molecular understanding of OAC and point to potential new therapeutic directions.
Embryonic stem cells (ESC) are able to give rise to any somatic cell type. A lot is known about how ESC pluripotency is maintained, but comparatively less is known about how differentiation is ...promoted. Cell fate decisions are regulated by interactions between signaling and transcriptional networks. Recent studies have shown that the overexpression or downregulation of the transcription factor Jun can affect the ESC fate. Here we have focussed on the role of the Jun in the exit of mouse ESCs from ground state pluripotency and the onset of early differentiation. Transcriptomic analysis of differentiating ESCs reveals that Jun is required to upregulate a programme of genes associated with cell adhesion as ESCs exit the pluripotent ground state. Several of these Jun‐regulated genes are shown to be required for efficient adhesion. Importantly this adhesion is required for the timely regulated exit of ESCs from ground state pluripotency and the onset of early differentiation events. Stem Cells 2016;34:1213–1224
Schematic model for the role of Jun in promoting the loss of pluripotency. Jun levels increase as ESCs exit from ground state pluripotency (upon "2i" withdrawal; ‐2i), leading to the activation of a program of genes associated with cell adhesion. This in turn contributes to the loss of pluripotency and the onset of the commitment and differentiation process. The heatmap shown below demonstrates the reciprocal effects of Jun depletion (2 days after 2i withdrawal; generally decreased gene activation) and overexpression (for 36 hours in 2i +LIF conditions; generally increased gene activation) on genes whose expression changes following Jun depletion.
Embryonic stem cells (ESCs) are unique in that they have the capacity to differentiate into all of the cell types in the body. We know a lot about the complex transcriptional control circuits that ...maintain the naive pluripotent state under self-renewing conditions but comparatively less about how cells exit from this state in response to differentiation stimuli. Here, we examined the role of Otx2 in this process in mouse ESCs and demonstrate that it plays a leading role in remodeling the gene regulatory networks as cells exit from ground state pluripotency. Otx2 drives enhancer activation through affecting chromatin marks and the activity of associated genes. Mechanistically, Oct4 is required for Otx2 expression, and reciprocally, Otx2 is required for efficient Oct4 recruitment to many enhancer regions. Therefore, the Oct4-Otx2 regulatory axis actively establishes a new regulatory chromatin landscape during the early events that accompany exit from ground state pluripotency.
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•Transcription factor Otx2 drives enhancer activation in differentiating mouse ESCs•Oct4 controls Otx2 expression levels•Otx2 collaborates with Oct4 in enhancer activation•Otx2 contributes to enhancer maintenance and de novo activation
The transcription factor Otx2 plays an important role in neural development and in the exit of embryonic stem cells from the pluripotent ground state. In this study, Yang et al. demonstrate that Otx2 drives enhancer activation and maintenance during the early cell-fate transition away from ground state pluripotency. Otx2 is involved in a regulatory partnership with Oct4 wherein Oct4 initially promotes Otx2 expression. Otx2 subsequently recruits Oct4 to a subset of enhancers and establishes a regulatory chromatin landscape.
Transcriptional activation is accompanied by multiple molecular events that remodel the local chromatin environment in promoter regions. These molecular events are often orchestrated in response to ...the activation of signalling pathways, as exemplified by the response of immediate early genes such as FOS to ERK MAP kinase signalling. Here, we demonstrate that inducible NFI recruitment permits PARP1 binding to the FOS promoter by a mutually reinforcing loop. PARP1 and its poly(ADP‐ribosyl)ation activity are required for maintaining FOS activation kinetics. We also show that the histone variant H2A.Z associates with the FOS promoter and acts in a transcription‐suppressive manner. However, in response to ERK pathway signalling, H2A.Z is replaced by H2A; PARP1 activity is required to promote this exchange. Thus, our work has revealed an additional facet of PARP1 function in promoting dynamic remodelling of promoter‐associated nucleosomes to allow transcriptional activation in response to cellular signalling.
This study reveals an essential role for PARP1 in the exchange of the histone variant H2A.Z by H2A at the ‐1 nucleosome of the FOS promoter, which promotes signal‐induced transcriptional activation.
Embryonic stem cells and induced pluripotent stem cells represent potentially important therapeutic agents in regenerative medicine. Complex interlinked transcriptional and signaling networks control ...the fate of these cells towards maintenance of pluripotency or differentiation. In this study we have focused on how mouse embryonic stem cells begin to differentiate and lose pluripotency and, in particular, the role that the ERK MAP kinase and GSK3 signaling pathways play in this process. Through a genome-wide siRNA screen we have identified more than 400 genes involved in loss of pluripotency and promoting the onset of differentiation. These genes were functionally associated with the ERK and/or GSK3 pathways, providing an important resource for studying the roles of these pathways in controlling escape from the pluripotent ground state. More detailed analysis identified MAP kinase phosphatases as a focal point of regulation and demonstrated an important role for these enzymes in controlling ERK activation kinetics and subsequently determining early embryonic stem cell fate decisions.
Recently, SUMO modification has been shown to impart repressive properties on several transcriptional regulatory proteins. Indeed, the ETS domain transcription factor Elk-1 is modified by SUMO, and ...this modification is reversed by ERK MAP kinase pathway activation. This causes a switch from a repressive to activated state. However, the mechanism(s) of SUMO-mediated transcriptional repression is unclear. Here, we have investigated how sumoylation of Elk-1 leads to transcriptional repression. We demonstrate that sumoylation of Elk-1 results in the recruitment of histone deacetylase activity to promoters. In particular, our data point to a key role for HDAC-2. This recruitment leads to decreased histone acetylation and hence transcriptional repression at Elk-1 target genes. Thus, our data demonstrate an important integration point for two protein-modifying pathways in the cell, the SUMO and deacetylation pathways, that combine to promote transcriptional repression.
Abstract
High levels of histone acetylation are associated with the regulatory elements of active genes, suggesting a link between acetylation and gene activation. We revisited this model, in the ...context of EGF-inducible gene expression and found that rather than a simple unifying model, there are two broad classes of genes; one in which high lysine acetylation activity is required for efficient gene activation, and a second group where the opposite occurs and high acetylation activity is inhibitory. We examined the latter class in more detail using EGR2 as a model gene and found that lysine acetylation levels are critical for several activation parameters, including the timing of expression onset, and overall amplitudes of the transcriptional response. In contrast, DUSP1 responds in the canonical manner and its transcriptional activity is promoted by acetylation. Single cell approaches demonstrate heterogenous activation kinetics of a given gene in response to EGF stimulation. Acetylation levels modify these heterogenous patterns and influence both allele activation frequencies and overall expression profile parameters. Our data therefore point to a complex interplay between acetylation equilibria and target gene induction where acetylation level thresholds are an important determinant of transcriptional induction dynamics that are sensed in a gene-specific manner.
Graphical Abstract
Graphical Abstract
Model showing how acetylation levels driven by the KDAC-KAT equilibrium differentially affects gene activation.
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