Shen Hsi Yang

SUMO promotes HDAC-mediated transcriptional repression.

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.

Dynamic Interplay of the SUMO and ERK Pathways in Regulating Elk-1 Transcriptional Activity

The ETS domain transcription factor Elk-1 is a direct target of the MAP kinase pathways. Phosphorylation of the Elk-1 transcriptional activation domain by MAP kinases triggers its activation. However, Elk-1 also contains two domains with repressive activities. One of these, the R motif, appears to function by suppressing the activity of the activation domain. Here, we demonstrate that SUMO modification of the R motif is required for this repressive activity. A dynamic interplay exists between the activating ERK MAP kinase pathway and the repressive SUMO pathway. ERK pathway activation leads to both phosphorylation of Elk-1 and loss of SUMO conjugation and, hence, to the loss of the repressive activity of the R motif. Thus, the reciprocal regulation of the activation and repressive activities are coupled by MAP kinase modification of Elk-1.

MAP kinase signalling cascades and transcriptional regulation

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.

An extended consensus motif enhances the specificity of substrate modification by SUMO

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.

MAP kinase phosphorylation-dependent activation of Elk-1 leads to activation of the co-activator p300.

CBP/p300 recruitment to enhancer‐bound complexes is a key determinant in promoter activation by many transcription factors. We present a novel mechanism of activating such complexes and show that pre‐assembled Elk‐1–p300 complexes become activated following Elk‐1 phosphorylation by changes in Elk‐1–p300 interactions rather than recruitment. It is known that Elk‐1 binds to promoter in the absence of stimuli. However, it is unclear how activation of Elk‐1 by mitogen‐acivated protein kinase (MAPK)‐mediated phosphorylation leads to targeted gene transactivation. We show that Elk‐1 can interact with p300 in vitro and in vivo in the absence of a stimulus through the Elk‐1 C‐terminus and the p300 N‐terminus. Phosphorylation on Ser383 and Ser389 of Elk‐1 by MAPK enhances this basal binding but, most importantly, Elk‐1 exhibits new interactions with p300. These interaction changes render a strong histone acetyltransferase activity in the Elk‐1‐associated complex that could play a critical role in chromatin remodeling and gene activation. The pre‐assembly mechanism may greatly accelerate transcription activation, which is important in regulation of expression of immediate‐early response genes, in particular those involved in stress responses.

Temporal Recruitment of the mSin3A-Histone Deacetylase Corepressor Complex to the ETS Domain Transcription Factor Elk-1

ABSTRACT The transcriptional status of eukaryotic genes is determined by a balance between activation and repression mechanisms. The nuclear hormone receptors represent classical examples of transcription factors that can regulate this balance by recruiting corepressor and coactivator complexes in a ligand-dependent manner. Here, we demonstrate that the equilibrium between activation and repression via a single transcription factor, Elk-1, is altered following activation of the Erk mitogen-activated protein kinase cascade. In addition to its C-terminal transcriptional activation domain, Elk-1 contains an N-terminal transcriptional repression domain that can recruit the mSin3A-histone deacetylase 1 corepressor complex. Recruitment of this corepressor is enhanced in response to activation of the Erk pathway in vivo, and this recruitment correlates kinetically with the shutoff of one of its target promoters, c-fos. Elk-1 therefore undergoes temporal activator-repressor switching and contributes to both the activation and repression of target genes following growth factor stimulation.

THE MECHANISM OF PHOSPHORYLATION-INDUCIBLE ACTIVATION OF THE ETS-DOMAIN TRANSCRIPTION FACTOR ELK-1

Protein phosphorylation represents one of the major mechanisms for transcription factor activation. Here we demonstrate a molecular mechanism by which phosphorylation by mitogen‐activated protein (MAP) kinases leads to changes in transcription factor activity. MAP kinases stimulate DNA binding and transcriptional activation mediated by the mammalian ETS‐domain transcription factor Elk‐1. Phosphorylation of the C‐terminal transcriptional activation domain induces a conformational change in Elk‐1, which accompanies the stimulation of DNA binding. C‐terminal phosphorylation is coupled to activation of DNA binding by the N‐terminal DNA‐binding domain via an additional intermediary domain. Activation of DNA binding is mediated by an allosteric mechanism involving the key phosphoacceptor residues. Together, these results provide a molecular model for how phosphorylation induces changes in Elk‐1 activity.

PIASx acts as an Elk-1 coactivator by facilitating derepression.

The ETS‐domain transcription factor Elk‐1 is a MAP kinase‐inducible transcriptional activator protein. However, in the basal state, its activity is repressed by SUMO‐dependent histone deacetylase (HDAC) recruitment. Relief of this repression accompanies the activation process. Here, we demonstrate that PIASxα acts to facilitate this derepression process. Members of the PIAS family of proteins can act as E3 enzymes that enhance the sumoylation status of a variety of substrates. However, PIASx‐mediated coactivation of Elk‐1 occurs in an E3 activity‐independent manner. PIASxα binds to Elk‐1 in vivo and enhances its transcriptional activity. The coactivating properties of PIASxα require Elk‐1 to be modified with SUMO and the integrity of the SUMO binding motif in PIASxα. PIASxα activates Elk‐1 through alterations in the HAT/HDAC activities associated with Elk‐1. In particular, PIASxα facilitates the loss of the repressive HDAC‐2 from sumoylated Elk‐1, a key event in the activation of Elk‐1 in response to signalling through the ERK MAP kinase pathway. Our data therefore reveal a novel coactivator function for PIASxα through reversing SUMO‐mediated repression of transcription factor activity.

Otx2 and Oct4 drive early enhancer activation during embryonic stem cell transition from naive pluripotency.

Summary 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.

The ETS Domain Transcription Factor Elk-1 Contains a Novel Class of Repression Domain

ABSTRACT The ETS domain transcription factor Elk-1 serves as an integration point for different mitogen-activated protein (MAP) kinase pathways. Phosphorylation of Elk-1 by MAP kinases triggers its activation. However, while the activation process is well understood, its downregulation-inactivation is less well characterized. The ETS DNA-binding domain plays a role in the downregulation of Elk-dependent promoter activity following mitogenic activation by recruiting the mSin3A-HDAC complex. Here we have identified a novel evolutionarily conserved repression domain in Elk-1, termed the R motif, which serves to reduce the basal transcriptional activity of Elk-1 and dampen its response to mitogenic signals. This domain is highly potent and portable and can repress transcription in trans. The R motif is related to the CRD1 repression domain in p300 and can functionally replace this domain and confer p21waf1/cip1 inducibility on p300. However, the R motif acts in a context-dependent manner and is not p21waf1/cip1 responsive in Elk-1. Thus, the Elk-1 R motif and the p300 CRD1 motif represent a new class of repression domains that are regulated in a context-dependent manner.