Histone deacetylases (HDACs) modulate chromatin structure and transcription, but little is known about their function in mammalian development. HDAC1 was implicated previously in the repression of genes required for cell proliferation and differentiation. Here we show that targeted disruption of both HDAC1 alleles results in embryonic lethality before E10.5 due to severe proliferation defects and retardation in development. HDAC1-deficient embryonic stem cells show reduced proliferation rates, which correlate with decreased cyclin-associated kinase activities and elevated levels of the cyclin-dependent kinase inhibitors p21(WAF1/CIP1) and p27(KIP1). Similarly, expression of p21 and p27 is up-regulated in HDAC1-null embryos. In addition, loss of HDAC1 leads to significantly reduced overall deacetylase activity, hyperacetylation of a subset of histones H3 and H4 and concomitant changes in other histone modifications. The expression of HDAC2 and HDAC3 is induced in HDAC1-deficient cells, but cannot compensate for loss of the enzyme, suggesting a unique function for HDAC1. Our study provides the first evidence that a histone deacetylase is essential for unrestricted cell proliferation by repressing the expression of selective cell cycle inhibitors.

SummaryCell populations can be strikingly heterogeneous, composed of multiple cellular states, each exhibiting stochastic noise in its gene expression. A major challenge is to disentangle these two types of variability and to understand the dynamic processes and mechanisms that control them. Embryonic stem cells (ESCs) provide an ideal model system to address this issue because they exhibit heterogeneous and dynamic expression of functionally important regulatory factors. We analyzed gene expression in individual ESCs using single-molecule RNA-FISH and quantitative time-lapse movies. These data discriminated stochastic switching between two coherent (correlated) gene expression states and burst-like transcriptional noise. We further showed that the “2i” signaling pathway inhibitors modulate both types of variation. Finally, we found that DNA methylation plays a key role in maintaining these metastable states. Together, these results show how ESC gene expression states and dynamics arise from a combination of intrinsic noise, coherent cellular states, and epigenetic regulation.

Histone deacetylases (HDACs) catalyze the removal of acetyl groups from core histones. Because of their capacity to induce local condensation of chromatin, HDACs are generally considered repressors of transcription. In this report, we analyzed the role of the class I histone deacetylase HDAC1 as a transcriptional regulator by comparing the expression profiles of wild-type and HDAC1-deficient embryonic stem cells. A specific subset of mouse genes (7%) was deregulated in the absence of HDAC1. We identified several putative tumor suppressors (JunB, Prss11, and Plagl1) and imprinted genes (Igf2, H19, and p57) as novel HDAC1 targets. The majority of HDAC1 target genes showed reduced expression accompanied by recruitment of HDAC1 and local reduction in histone acetylation at regulatory regions. At some target genes, the related deacetylase HDAC2 partially masks the loss of HDAC1. A second group of genes was found to be downregulated in HDAC1-deficient cells, predominantly by additional recruitment of HDAC2 in the absence of HDAC1. Finally, a small set of genes (Gja1, Irf1, and Gbp2) was found to require HDAC activity and recruitment of HDAC1 for their transcriptional activation. Our study reveals a regulatory cross talk between HDAC1 and HDAC2 and a novel function for HDAC1 as a transcriptional coactivator.

The cyclin-dependent kinase inhibitor p21/WAF1/CIP1 is an important regulator of cell cycle progression, senescence, and differentiation. Genotoxic stress leads to activation of the tumor suppressor p53 and subsequently to induction of p21 expression. Here we show that the tumor suppressor p53 cooperates with the transcription factor Sp1 in the activation of the p21 promoter, whereas histone deacetylase 1 (HDAC1) counteracts p53-induced transcription from the p21 gene. The p53 protein binds directly to the C terminus of Sp1, a domain which was previously shown to be required for the interaction with HDAC1. Induction of p53 in response to DNA-damaging agents resulted in the formation of p53-Sp1 complexes and simultaneous dissociation of HDAC1 from the C terminus of Sp1. Chromatin immunoprecipitation experiments demonstrated the association of HDAC1 with the p21 gene in proliferating cells. Genotoxic stress led to recruitment of p53, reduced binding of HDAC1, and hyperacetylation of core histones at the p21 promoter. Our findings show that the deacetylase HDAC1 acts as an antagonist of the tumor suppressor p53 in the regulation of the cyclin-dependent kinase inhibitor p21 and provide a basis for understanding the function of histone deacetylase inhibitors as antitumor drugs.The tumor suppressor p53 can induce cell cycle arrest or apoptosis in response to a variety of stress signals, such as DNA damage, oncogenic stimuli, or hypoxia (reviewed in reference 49). Activation of p53 occurs by several mechanisms including protein stabilization and modification of the protein by phosphorylation and acetylation. p53 is a transcription factor that recognizes specific binding sites within numerous target genes including mdm2, cyclin G, bax, and p21/WAF1/CIP1 (for reviews see references 5 and 12). While multiple downstream targets are involved in the mediation of apoptotic effects, the main target for p53-induced cell cycle arrest seems to be the p21 gene. p21 has been identified by virtue of its activation by p53 (13), its association with cyclin/cyclin-dependent kinase (CDK) complexes (23, 66), and its up-regulation during senescence (47). Furthermore, the p21 protein was shown previously to interact with the proliferating cell nuclear antigen (PCNA), thereby preventing DNA replication (10). Induction of p21 expression by genotoxic stress and its role during terminal differentiation of various cell types have been investigated intensively. While p21 is activated by p53-dependent mechanisms in response to DNA damage to ensure cell cycle arrest and repair, a variety of agents that promote differentiation, like phorbol ester or okadaic acid, can up-regulate p21 independently of p53 (for a review see reference 16). Similarly, the p21 gene can be activated by transforming growth factor ␤, Ca 2ϩ , lovastatin, or nerve growth factor (16).Recently, a number of reports demonstrated the induction of p21 by inhibitors of histone deacetylases (HDACs), such as sodium butyrate (46), trichostatin A (TSA) (56), suberoylanilide hydroxamic a...

Prdm14 promotes germline fate and naive pluripotency by repressing FGF signalling and DNA methylationThe transcription factor Prdm14 is a critical regulator of pluripotency and germline development. This study shows that Prdm14 represses DNA methylation and FGF signaling, thereby promoting both germ cell fate and naive pluripotency in ESC.

Histone deacetylases (HDACs) are chromatin-modifying enzymes that are involved in the regulation of proliferation, differentiation and development. HDAC inhibitors induce cell cycle arrest, differentiation, or apoptosis in tumor cells and are therefore promising antitumor agents. Numerous genes were found to be deregulated upon HDAC inhibitor treatment; however, the relevant target enzymes are still unidentified. HDAC1 is required for mouse development and unrestricted proliferation of embryonic stem cells. We show here that HDAC1 reversibly regulates cellular proliferation and represses the cyclin-dependent kinase inhibitor p21 in embryonic stem cells. Disruption of the p21 gene rescues the proliferation phenotype of HDAC1 ؊/؊ embryonic stem cells but not the embryonic lethality of HDAC1 ؊/؊ mice. In the absence of HDAC1, mouse embryonic fibroblasts scarcely undergo spontaneous immortalization and display increased p21 expression. Chromatin immunoprecipitation assays demonstrate a direct regulation of the p21 gene by HDAC1 in mouse embryonic fibroblasts. Transformation with simian virus 40 large T antigen or ablation of p21 restores normal immortalization of primary HDAC1؊/؊ fibroblasts. Our data demonstrate that repression of the p21 gene is crucial for HDAC1-mediated control of proliferation and immortalization. HDAC1 might therefore be one of the relevant targets for HDAC inhibitors as anticancer drugs.Acetylation of core histones is linked to the opening of chromatin and transcriptional activation. Modification of lysine residues by acetylation is thought to affect gene expression either by altering the affinity of histones to the DNA, or by creating binding sites for detector proteins that regulate chromatin accessibility. The antagonistic activities of two types of enzymes, histone acetyltransferases and histone deacetylases (HDACs), control the reversible acetylation state at the Nterminal tail of histones. HDACs catalyze the removal of the acetyl moieties from acetylated histones and other proteins and are in general associated with transcriptional repression (17). Based on their homologies with yeast deacetylases mammalian HDACs have been classified into Rpd3-like (class I), Hda1-like (class II), and Sir2-like (class III) enzymes (19). HDAC11 seems to represent a class (class IV) on its own.HDACs have been shown to regulate many important biological processes, including cell cycle progression, differentiation, and development. In agreement with this idea, HDAC inhibitor treatment leads to cell cycle arrest, differentiation, and apoptosis in cultured tumor cells and tumors in animal models. Therefore, several HDAC inhibitors are currently tested as antitumor drugs in clinical trials. A variety of HDAC inhibitors, which target class I and class II enzymes have been identified (33), and it has been shown that they exert their antiproliferative effects via transcriptional and nontranscriptional mechanisms (32). Treatment of untransformed cells with HDAC inhibitors triggers a G 2 checkpoint resulting in arr...

An intricate link is becoming apparent between metabolism and cellular identities. Here, we explore the basis for such a link in an in vitro model for early mouse embryonic development: from naïve pluripotency to the specification of primordial germ cells (PGCs). Using single‐cell RNA‐seq with statistical modelling and modulation of energy metabolism, we demonstrate a functional role for oxidative mitochondrial metabolism in naïve pluripotency. We link mitochondrial tricarboxylic acid cycle activity to IDH2‐mediated production of alpha‐ketoglutarate and through it, the activity of key epigenetic regulators. Accordingly, this metabolite has a role in the maintenance of naïve pluripotency as well as in PGC differentiation, likely through preserving a particular histone methylation status underlying the transient state of developmental competence for the PGC fate. We reveal a link between energy metabolism and epigenetic control of cell state transitions during a developmental trajectory towards germ cell specification, and establish a paradigm for stabilizing fleeting cellular states through metabolic modulation.

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