Members of the myocyte enhancer factor-2 (MEF2) family of transcription factors associate with myogenic basic helix-loop-helix transcription factors such as MyoD to activate skeletal myogenesis. MEF2 proteins also interact with the class II histone deacetylases HDAC4 and HDAC5, resulting in repression of MEF2-dependent genes. Execution of the muscle differentiation program requires release of MEF2 from repression by HDACs, which are expressed constitutively in myoblasts and myotubes. Here we show that HDAC5 shuttles from the nucleus to the cytoplasm when myoblasts are triggered to differentiate. Calcium/calmodulin-dependent protein kinase (CaMK) signalling, which stimulates myogenesis and prevents formation of MEF2-HDAC complexes, also induces nuclear export of HDAC4 and HDAC5 by phosphorylation of these transcriptional repressors. An HDAC5 mutant lacking two CaMK phosphorylation sites is resistant to CaMK-mediated nuclear export and acts as a dominant inhibitor of skeletal myogenesis, whereas a cytoplasmic HDAC5 mutant is unable to block efficiently the muscle differentiation program. Our results highlight a mechanism for transcriptional regulation through signal- and differentiation-dependent nuclear export of a chromatin-remodelling enzyme, and suggest that nucleo-cytoplasmic trafficking of HDACs is involved in the control of cellular differentiation.

The heart responds to stress signals by hypertrophic growth, which is accompanied by activation of the MEF2 transcription factor and reprogramming of cardiac gene expression. We show here that class II histone deacetylases (HDACs), which repress MEF2 activity, are substrates for a stress-responsive kinase specific for conserved serines that regulate MEF2-HDAC interactions. Signal-resistant HDAC mutants lacking these phosphorylation sites are refractory to hypertrophic signaling and inhibit cardiomyocyte hypertrophy. Conversely, mutant mice lacking the class II HDAC, HDAC9, are sensitized to hypertrophic signals and exhibit stress-dependent cardiomegaly. Thus, class II HDACs act as signal-responsive suppressors of the transcriptional program governing cardiac hypertrophy and heart failure.

The eukaryotic transcription factor NF-B plays a central role in the induced expression of human immunodeficiency virus type 1 and in many aspects of the genetic program mediating normal T-cell activation and growth. The nuclear activity of NF-B is tightly regulated from the cytoplasmic compartment by an inhibitory subunit called IB␣. This cytoplasmic inhibitor is rapidly phosphorylated and degraded in response to a diverse set of NF-B-inducing agents, including T-cell mitogens, proinflammatory cytokines, and viral transactivators such as the Tax protein of human T-cell leukemia virus type 1. To explore these IB␣-dependent mechanisms for NF-B induction, we identified novel mutants of IB␣ that uncouple its inhibitory and signal-transducing functions in human T lymphocytes. Specifically, removal of the N-terminal 36 amino acids of IB␣ failed to disrupt its ability to form latent complexes with NF-B in the cytoplasm. However, this deletion mutation prevented the induced phosphorylation, degradative loss, and functional release of IB␣ from NF-B in Tax-expressing cells. Alanine substitutions introduced at two serine residues positioned within this N-terminal regulatory region of IB␣ also yielded constitutive repressors that escaped from Tax-induced turnover and that potently inhibited immune activation pathways for NF-B induction, including those initiated from antigen and cytokine receptors. In contrast, introduction of a phosphoserine mimetic at these sites rectified this functional defect, a finding consistent with a causal linkage between the phosphorylation status and proteolytic stability of this cytoplasmic inhibitor. Together, these in vivo studies define a critical signal response domain in IB␣ that coordinately controls the biologic activities of IB␣ and NF-B in response to viral and immune stimuli.

No abstract

ABSTRACT

Previous work has identified alterations in histone acetylation in animal models of drug addiction and depression. However, the mechanisms which integrate drugs and stress with changes in chromatin structure remain unclear. Here, we identify the activity-dependent class II histone deacetylase, HDAC5, as a central integrator of these stimuli with changes in chromatin structure and gene expression. Chronic, but not acute, exposure to cocaine or stress decreases HDAC5 function in the nucleus accumbens (NAc), a major brain reward region, which allows for increased histone acetylation and transcription of HDAC5 target genes. This regulation is behaviorally important, as loss of HDAC5 causes hypersensitive responses to chronic, not acute, cocaine or stress. These findings suggest that proper balance of histone acetylation is a crucial factor in the saliency of a given stimulus and that disruption of this balance is involved in the transition from an acute adaptive response to a chronic psychiatric illness.

A variety of stress signals stimulate cardiac myocytes to undergo hypertrophy. Persistent cardiac hypertrophy is associated with elevated risk for the development of heart failure. Recently, we showed that class II histone deacetylases (HDACs) suppress cardiac hypertrophy and that stress signals neutralize this repressive function by triggering phosphorylation-and CRM1-dependent nuclear export of these chromatin-modifying enzymes. However, the identities of cardiac HDAC kinases have remained unclear. Here, we demonstrate that signaling by protein kinase C (PKC) is sufficient and, in some cases, necessary to drive nuclear export of class II HDAC5 in cardiomyocytes. Inhibition of PKC prevents nucleocytoplasmic shuttling of HDAC5 in response to a subset of hypertrophic agonists. Moreover, a nonphosphorylatable HDAC5 mutant is refractory to PKC signaling and blocks cardiomyocyte hypertrophy mediated by pharmacological activators of PKC. We also demonstrate that protein kinase D (PKD), a downstream effector of PKC, directly phosphorylates HDAC5 and stimulates its nuclear export. These findings reveal a novel function for the PKC/PKD axis in coupling extracellular cues to chromatin modifications that control cellular growth, and they suggest potential utility for small-molecule inhibitors of this pathway in the treatment of pathological cardiac gene expression.Coordinated changes in gene transcription during cell growth and differentiation require mechanisms for coupling intracellular signaling pathways with the genome. The acetylation of nucleosomal histones has emerged as a central mechanism in the control of gene transcription during such cellular transitions (20). Acetylation of histones by histone acetyltransferases promotes transcription by relaxing chromatin structure, whereas histone deacetylation by histone deacetylases (HDACs) reverses this process, resulting in transcriptional repression. How these chromatin-modifying enzymes are linked to, and controlled by, intracellular signaling is only beginning to be understood.There are two classes of HDACs that can be distinguished by their structures and expression patterns. Class I HDACs (HDAC1, HDAC2, and HDAC3) are expressed ubiquitously and are composed mainly of a catalytic domain (13). In contrast, class II HDACs (HDAC4, HDAC5, HDAC7, and HDAC9) display more restricted expression patterns and contain an N-terminal extension, which mediates interactions with other transcriptional cofactors and confers responsiveness to calcium-dependent signaling (12,25,33). Signaling by calcium/ calmodulin-dependent protein kinase (CaMK) results in phosphorylation of the N termini of class II HDACs, which govern their intracellular localization and interactions with other factors (29, 32). Phosphorylation of signal-responsive serine residues creates docking sites for the 14-3-3 family of chaperone proteins, which promote shuttling of HDACs from the nucleus to the cytoplasm in a CRM1-dependent fashion (14,21,30,31,48).CaMK signaling to class II HDACs governs the activity of th...

The adult heart responds to stress signals by hypertrophic growth, which is often accompanied by activation of a fetal cardiac gene program and eventual cardiac demise. We showed previously that histone deacetylase 9 (HDAC9) acts as a suppressor of cardiac hypertrophy and that mice lacking HDAC9 are sensitized to cardiac stress signals. Here we report that mice lacking HDAC5 display a similar cardiac phenotype and develop profoundly enlarged hearts in response to pressure overload resulting from aortic constriction or constitutive cardiac activation of calcineurin, a transducer of cardiac stress signals. In contrast, mice lacking either HDAC5 or HDAC9 show a hypertrophic response to chronic ␤-adrenergic stimulation identical to that of wild-type littermates, suggesting that these HDACs modulate a specific subset of cardiac stress response pathways. We also show that compound mutant mice lacking both HDAC5 and HDAC9 show a propensity for lethal ventricular septal defects and thin-walled myocardium. These findings reveal central roles for HDACs 5 and 9 in the suppression of a subset of cardiac stress signals as well as redundant functions in the control of cardiac development.Postnatal growth of the heart occurs primarily through hypertrophy, in which cardiac myocytes increase in size but not in number (reviewed in reference 34). Hypertrophy can occur in response to physiological stimuli, such as exercise, or pathological stimuli, such as myocardial infarction, hypertension, aortic stenosis, or valve dysfunction. While stress-induced hypertrophy serves initially to normalize ventricular wall stress, this form of hypertrophy, when prolonged, can progress to dilated cardiomyopathy and sudden death. Pathological cardiac hypertrophy is a major predictor of human morbidity and mortality and a major cause of heart failure (17,18,25).Numerous intracellular signaling pathways have been implicated in the transduction of hypertrophic signals from the cardiomyocyte cell surface to the nucleus (reviewed in references 2, 8, 22, and 34). Many hypertrophic agonists acting through cell surface receptors coupled to G␣q mobilize intracellular calcium, which activates downstream kinases and the calcium-and calmodulin-dependent phosphatase calcineurin. Activation of these effectors is sufficient and, in many cases, necessary for hypertrophic growth of the heart (14, 31, 35, 49). Elevation of cyclic AMP in response to ␤-adrenergic agonists also stimulates cardiac hypertrophy via protein kinase A and other downstream effectors (36). The identification of nodal points in hypertrophic signaling pathways and the mechanisms that link signaling in the cytoplasm with changes in gene expression that contribute to maladaptive growth of the heart represent major challenges in the field.Pathological cardiac hypertrophy is coupled to the activation of a fetal cardiac gene program, which results in the expression of fetal proteins involved in contractility, metabolism, and calcium handling that are incompatible with sustained function of the adult...