Studies of H3K4me3 demethylation by KDM5B/Jarid1B/PLU1 reveals strong substrate recognition in vitro and identifies 2,4‐pyridine‐dicarboxylic acid as an in vitro and in cell inhibitor

Kristensen, Line H.; Nielsen, Anders L.; Helgstrand, Charlotte; Lees, Michael; Cloos, Paul A.; Kastrup, J.S.; Helin, Kristian; Olsen, Lars; Gajhede, Michael

doi:10.1111/j.1742-4658.2012.08567.x

Cited by 66 publications

(62 citation statements)

References 38 publications

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“…8A). Two studies have identified 2,4-pyridine-dicarboxylic acid (PDCA) as a small-molecule inhibitor of KDM5B (53,54). Following 24 h of OSKM overexpression, we observed a higher density and compaction of shKdm5b 4TF-MEFs than shLuc 4TF-MEFs (Fig.…”

Section: Kdm5b-depleted Es Cells Exhibit Reduced Self-renewalmentioning

confidence: 59%

Extended Self-Renewal and Accelerated Reprogramming in the Absence of Kdm5b

Kidder

et al. 2013

Molecular and Cellular Biology

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c Embryonic stem (ES) cell pluripotency is thought to be regulated in part by H3K4 methylation. However, it is unclear how H3K4 demethylation contributes to ES cell function and participates in induced pluripotent stem (iPS) cell reprogramming. Here, we show that KDM5B, which demethylates H3K4, is important for ES cell differentiation and presents a barrier to the reprogramming process. Depletion of Kdm5b leads to an extension in the self-renewal of ES cells in the absence of LIF. Transcriptome analysis revealed the persistent expression of pluripotency genes and underexpression of developmental genes during differentiation in the absence of Kdm5b, suggesting that KDM5B plays a key role in cellular fate changes. We also observed accelerated reprogramming of differentiated cells in the absence of Kdm5b, demonstrating that KDM5B is a barrier to the reprogramming process. Expression analysis revealed that mesenchymal master regulators associated with the epithelial-to-mesenchymal transition (EMT) are downregulated during reprogramming in the absence of Kdm5b. Moreover, global analysis of H3K4me3/2 revealed that enhancers of fibroblast genes are rapidly deactivated in the absence of Kdm5b, and genes associated with EMT lose H3K4me3/2 during the early reprogramming process. These findings provide functional insight into the role for KDM5B in regulating ES cell differentiation and as a barrier to the reprogramming process. E mbryonic stem (ES) cells have the unique ability to self-renew indefinitely and differentiate into the hundreds of cell types that exist in the mammalian developmental repertoire. Epigenetic regulation of transcription is critical to achieve defined cellular states that persist in development. ES cell self-renewal versus differentiation is regulated in part by external stimuli that signal to transcription factors (TFs) and chromatin modifiers to regulate the underlying chromatin structure. ES cells express high levels of TFs, such as Oct4, Sox2, Nanog, and Tbx3, that regulate pluripotency by associating with specific DNA sequences to drive expression of a network of pluripotency-related genes and to repress developmentally regulated genes (1-3). Disruption of these core regulatory factors results in a compromised self-renewal state leading to differentiation (4). While the functions of many TFs have been evaluated in ES cells, few studies have focused on the roles of chromatin modifiers in ES cell pluripotency (5-7). Chromatin regulation by way of posttranslational modification of histone tails creates an environment that is conducive or repressive for transcriptional activity, which is critical for propagating expression of networks of genes that maintain self-renewal or promote differentiation.The trithorax group (trxG) complex regulates methylation of lysine 4 of histone H3 (H3K4me), which is predominantly associated with active genes (8). H3K4me3 is highly enriched at transcriptional start sites (TSS) of highly active genes (9-12) where it is important for RNA polymerase II binding and activat...

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Section: Kdm5b-depleted Es Cells Exhibit Reduced Self-renewalmentioning

confidence: 59%

Extended Self-Renewal and Accelerated Reprogramming in the Absence of Kdm5b

Kidder

et al. 2013

Molecular and Cellular Biology

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“…S2 in the supplemental material). However, in contrast to all other small-molecule inhibitors used in this study, 2,4-PDCA, a histone demethylase inhibitor with high specificity for JARID1 and JMJD2 family demethylases, which are responsible for H3K4me3 and H3K36me3 demethylation, respectively (35), strongly enhanced both the direct repressive effect and stable silencing by antisense transcription during differentiation (Fig. 5E).…”

Section: Resultsmentioning

confidence: 99%

Chromatin-Mediated Reversible Silencing of Sense-Antisense Gene Pairs in Embryonic Stem Cells Is Consolidated upon Differentiation

Loos

Loda

Wijk

et al. 2015

Molecular and Cellular Biology

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Genome-wide gene expression studies have indicated that the eukaryotic genome contains many gene pairs showing overlapping sense and antisense transcription. Regulation of these coding and/or noncoding gene pairs involves intricate regulatory mechanisms. In the present study, we utilized an enhanced green fluorescent protein (EGFP)-tagged reporter plasmid cis linked to a doxycycline-inducible antisense promoter, generating antisense transcription that fully overlaps EGFP, to study the mechanism and dynamics of gene silencing after induction of noncoding antisense transcription in undifferentiated and differentiating mouse embryonic stem cells (ESCs). We found that EGFP silencing is reversible in ESCs but is locked into a stable state upon ESC differentiation. Reversible silencing in ESCs is chromatin dependent and is associated with accumulation of trimethylated lysine 36 on histone H3 (H3K36me3) at the EGFP promoter region. In differentiating ESCs, antisense transcription-induced accumulation of H3K36me3 was associated with an increase in CpG methylation at the EGFP promoter. Repression of the sense promoter was affected by small-molecule inhibitors which interfere with DNA methylation and histone demethylation pathways. Our results indicate a general mechanism for silencing of fully overlapping sense-antisense gene pairs involving antisense transcription-induced accumulation of H3K36me3 at the sense promoter, resulting in reversible silencing of the sense partner, which is stabilized during ESC differentiation by CpG methylation.

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“…1a). Previously, we and others have shown that recombinant protein containing the N-terminal half of KDM5C (residues 1-839), KDM5B (residues 1-769), or KDM5A (residues 1-797) is enzymatically active (43)(44)(45). Interestingly, the rice Jumonji demethylase JMJ703 has KDM5-like activity on trimethylated histone H3 Lys-4 but contains no insertion within the catalytic domain (47).…”

Section: The Atomic Coordinates and Structure Factors (Code 5e6h) Havmentioning

confidence: 99%

“…1a), we generated a hypothetical structural model for the N-terminal half of the KDM5 family, which is enzymatically active in vitro (43,44), using PHYRE2 (Protein Homology/analogy Recognition Engine) (57). With PHYRE2, we generated individual domain models based on known structural information.…”

Section: A Hypothetical Model Of the N-terminal Half Of Kdm5-tomentioning

confidence: 99%

Characterization of a Linked Jumonji Domain of the KDM5/JARID1 Family of Histone H3 Lysine 4 Demethylases

Horton¹,

Engstrom²,

Zoeller³

et al. 2016

Journal of Biological Chemistry

104

165

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The KDM5/JARID1 family of Fe(II)-and ␣-ketoglutarate-dependent demethylases remove methyl groups from tri-and dimethylated lysine 4 of histone H3. Accumulating evidence from primary tumors and model systems supports a role for KDM5A (JARID1A/RBP2) and KDM5B (JARID1B/PLU1) as oncogenic drivers. The KDM5 family is unique among the Jumonji domain-containing histone demethylases in that there is an atypical insertion of a DNA-binding ARID domain and a histone-binding PHD domain into the Jumonji domain, which separates the catalytic domain into two fragments (JmjN and JmjC). Here we demonstrate that internal deletion of the ARID and PHD1 domains has a negligible effect on in vitro enzymatic kinetics of the KDM5 family of enzymes. We present a crystal structure of the linked JmjN-JmjC domain from KDM5A, which reveals that the linked domain fully reconstitutes the cofactor (metal ion and ␣-ketoglutarate) binding characteristics of other structurally characterized Jumonji domain demethylases. Docking studies with GSK-J1, a selective inhibitor of the KDM6/ KDM5 subfamilies, identify critical residues for binding of the inhibitor to the reconstituted KDM5 Jumonji domain. Further, we found that GSK-J1 inhibited the demethylase activity of KDM5C with 8.5-fold increased potency compared with that of KDM5B at 1 mM ␣-ketoglutarate. In contrast, JIB-04 (a paninhibitor of the Jumonji demethylase superfamily) had the opposite effect and was ϳ8-fold more potent against KDM5B than against KDM5C. Interestingly, the relative selectivity of JIB-04 toward KDM5B over KDM5C in vitro translates to a ϳ10 -50-fold greater growth-inhibitory activity against breast cancer cell lines. These data define the minimal requirements for enzymatic activity of the KDM5 family to be the linked JmjNJmjC domain coupled with the immediate C-terminal helical zinc-binding domain and provide structural characterization of the linked JmjN-JmjC domain for the KDM5 family, which should prove useful in the design of KDM5 demethylase inhibitors with improved potency and selectivity.

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Studies of H3K4me3 demethylation by KDM5B/Jarid1B/PLU1 reveals strong substrate recognition in vitro and identifies 2,4‐pyridine‐dicarboxylic acid as an in vitro and in cell inhibitor

Cited by 66 publications

References 38 publications

Extended Self-Renewal and Accelerated Reprogramming in the Absence of Kdm5b

Extended Self-Renewal and Accelerated Reprogramming in the Absence of Kdm5b

Chromatin-Mediated Reversible Silencing of Sense-Antisense Gene Pairs in Embryonic Stem Cells Is Consolidated upon Differentiation

Characterization of a Linked Jumonji Domain of the KDM5/JARID1 Family of Histone H3 Lysine 4 Demethylases

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