The regulation of acetylation is central for the epigenetic control of lineage-specific gene expression and determines cell fate decisions. We provide evidence that the inhibition of histone deacetylases (HDACs) blocks the endothelial differentiation of adult progenitor cells. To define the mechanisms by which HDAC inhibition prevents endothelial differentiation, we determined the expression of homeobox transcription factors and demonstrated that HoxA9 expression is down-regulated by HDAC inhibitors. The causal involvement of HoxA9 in the endothelial differentiation of adult progenitor cells is supported by the finding that HoxA9 overexpression partially rescued the endothelial differentiation blockade induced by HDAC inhibitors. Knockdown and overexpression studies revealed that HoxA9 acts as a master switch to regulate the expression of prototypical endothelial-committed genes such as endothelial nitric oxide synthase, VEGF-R2, and VE-cadherin, and mediates the shear stress–induced maturation of endothelial cells. Consistently, HoxA9-deficient mice exhibited lower numbers of endothelial progenitor cells and showed an impaired postnatal neovascularization capacity after the induction of ischemia. Thus, HoxA9 is regulated by HDACs and is critical for postnatal neovascularization.

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Our data establish TWEAK as a positive regulator of cardiomyocyte proliferation.

In the isolated CNS, different modulatory inputs can enable one motor network to generate multiple output patterns. Thus far, however, few studies have established whether different modulatory inputs also enable a defined network to drive distinct muscle and movement patterns in vivo, much as they enable these distinctions in behavioral studies. This possibility is not a foregone conclusion, because additional influences present in vivo (e.g., sensory feedback, hormonal modulation) could alter the motor patterns. Additionally, rhythmic neuronal activity can be transformed into sustained muscle contractions, particularly in systems with slow muscle dynamics, as in the crab (Cancer borealis) stomatogastric system used here. We assessed whether two different versions of the biphasic (protraction, retraction) gastric mill (chewing) rhythm, triggered in the isolated stomatogastric system by the modulatory ventral cardiac neurons (VCNs) and postoesophageal commissure (POC) neurons, drive different muscle and movement patterns. One distinction between these rhythms is that the lateral gastric (LG) protractor motor neuron generates tonic bursts during the VCN rhythm, whereas its POC-rhythm bursts are divided into fast, rhythmic burstlets. Intracellular muscle fiber recordings and tension measurements show that the LGinnervated muscles retain the distinct VCN-LG and POC-LG neuron burst structures. Moreover, endoscope video recordings in vivo, during VCN-triggered and POC-triggered chewing, show that the lateral teeth protraction movements exhibit the same, distinct protraction patterns generated by LG in the isolated nervous system. Thus, the multifunctional nature of an identified motor network in the isolated CNS can be preserved in vivo, where it drives different muscle activity and movement patterns.

Chromatin modifying enzymes play a critical role in cardiac differentiation. Previously, it has been shown that the targeted deletion of the histone methyltransferase, Smyd1, the founding member of the SET and MYND domain containing (Smyd) family, interferes with cardiomyocyte maturation and proper formation of the right heart ventricle. The highly related paralogue, Smyd2 is a histone 3 lysine 4- and lysine 36-specific methyltransferase expressed in heart and brain. Here, we report that Smyd2 is differentially expressed during cardiac development with highest expression in the neonatal heart. To elucidate the functional role of Smyd2 in the heart, we generated conditional knockout (cKO) mice harboring a cardiomyocyte-specific deletion of Smyd2 and performed histological, functional and molecular analyses. Unexpectedly, cardiac deletion of Smyd2 was dispensable for proper morphological and functional development of the murine heart and had no effect on global histone 3 lysine 4 or 36 methylation. However, we provide evidence for a potential role of Smyd2 in the transcriptional regulation of genes associated with translation and reveal that Smyd2, similar to Smyd3, interacts with RNA Polymerase II as well as to the RNA helicase, HELZ.

Posttranslational histone modification by acetylation or methylation regulates gene expression. Here, we investigated the role of the histone lysine methyltransferase MLL for angiogenic functions in human umbilical vein endothelial cells. Suppression of MLL expression by siRNA or incubation with the pharmacologic methyltransferase inhibitor 5-deoxy-5-(methylthio)adenosine significantly decreased endothelial-cell migration and capillary sprout formation, indicating that methyltransferase activity is required for proangiogenic endothelialcell functions. Because the expression of homeodomain transcription factors (Hox) is regulated by MLL, we elucidated the role of Hox gene expression. MLL silencing was associated with reduced mRNA and protein expression of HoxA9 and HoxD3, whereas HoxB3, HoxB4, HoxB5, and HoxB9 were not altered. Overexpression of HoxA9 or HoxD3 partially compensated for impaired migration in MLL siRNA-transfected endothelial cells, suggesting that HoxA9 and HoxD3 both contribute to MLL-dependent migration. As a potential underlying mechanism, MLL siRNA down-regulated mRNA and protein levels of the HoxA9-dependent axon guidance factor EphB4. In contrast, MLL knockdown effects on capillary sprouting were not rescued by HoxA9 or HoxD3 overexpression, indicating that MLL affects additional targets required for 3-dimensional sprout formation. IntroductionMLL (mixed lineage leukemia, ALL-1, HRX) is a chromatin regulator homolog to the Drosophila trithorax gene. 1-3 MLL was originally discovered by its involvement in human acute leukemia through chromosomal translocation and fusion to a variety of genes. 4 MLL fusion proteins were shown to cause myeloid progenitor immortalization by inducing a characteristic expression profile of homeobox (Hox) transcription factors. 5,6 Physiologically, the SET domain of MLL catalyzes the methylation of lysine 4 of histone H3, thereby regulating gene expression. 7,8 In particular, native MLL is essential for the epigenetic control of gene expression of a set of homeobox proteins HoxA7, HoxA9, and HoxA10 during ex vivo differentiation of embryonic stem cells 9 and for physiologic hematopoiesis. 10-12 MLL Ϫ/Ϫ mice die around dpc 10.5 with aberrant rostrocaudal segmentation, 13 or, in a different model targeting exons 12 to 14, at dpc 11.5 to 14.5 with edematous bodies and petechiae. 12 Beyond segmental identity and hematopoiesis, MLL is crucial for the maintenance of helper T-lymphocyte function. 14 As yet, however, a function of MLL in postnatal angiogenesis has not been explored.Hox transcription factors play important roles during embryonic development of the cardiovascular system. [15][16][17] Certain members of the Hox family also regulate postnatal angiogenesis and endothelialcell adhesion and migration. HoxD3 promotes endothelial invasion and migration 18 and converts resting endothelium to an angiogenic phenotype by inducing proangiogenic genes including the integrin subunits ␣5 and ␤3. 19 HoxB3 is required for capillary morphogenesis of preformed vascular sprout...

The extracellular matrix is crucial for organogenesis. It is a complex and dynamic component that regulates cell behavior by modulating the activity, bioavailability and presentation of growth factors to cell surface receptors. Here, we determined the role of the extracellular matrix protein Nephronectin (Npnt) in heart development using the zebrafish model system. The vertebrate heart is formed as a linear tube in which myocardium and endocardium are separated by a layer of extracellular matrix termed the cardiac jelly. During heart development, the cardiac jelly swells at the atrioventricular (AV) canal, which precedes valve formation. Here, we show that Npnt expression correlates with this process. Morpholino-mediated knockdown of Npnt prevents proper valve leaflet formation and trabeculation and results in greater than 85% lethality at 7 days post-fertilization. The earliest observed phenotype is an extended tube-like structure at the AV boundary. In addition, the expression of myocardial genes involved in cardiac valve formation (cspg2, fibulin 1, tbx2b, bmp4) is expanded and endocardial cells along the extended tube-like structure exhibit characteristics of AV cells (has2, notch1b and Alcam expression, cuboidal cell shape). Inhibition of has2 in npnt morphants rescues the endocardial, but not the myocardial, expansion. By contrast, reduction of BMP signaling in npnt morphants reduces the ectopic expression of myocardial and endocardial AV markers. Taken together, our results identify Npnt as a novel upstream regulator of Bmp4-Has2 signaling that plays a crucial role in AV canal differentiation.

Hedrich UB, Diehl F, Stein W. Gastric and pyloric motor pattern control by a modulatory projection neuron in the intact crab Cancer pagurus. J Neurophysiol 105: 1671-1680, 2011. First published February 16, 2011 doi:10.1152/jn.01105.2010.-Neuronal release of modulatory substances provides motor pattern generating circuits with a high degree of flexibility. In vitro studies have characterized the actions of modulatory projection neurons in great detail in the stomatogastric nervous system, a model system for neuromodulatory influences on central pattern generators. Less is known about the activities and actions of modulatory neurons in fully functional and richly modulated network settings, i.e., in intact animals. It is also unknown whether their activities contribute to the motor patterns in different behavioral conditions. Here, we show for the first time the activity and effects of the well-characterized modulatory projection neuron 1 (MCN1) in vivo and compare them to in vitro conditions. MCN1 was always spontaneously active, typically in a rhythmic fashion with its firing being interrupted by ascending inhibitions from the pyloric motor circuit. Its activity contributed to pyloric motor activity, because 1) the cycle period of the motor pattern correlated with MCN1 firing frequency and 2) stimulating MCN1 shortened the cycle period while 3) lesioning of the MCN1 axon reduced motor activity. In addition, gastric mill motor activity was elicited for the duration of the stimulation. Chemosensory stimulation of the antennae moved MCN1 away from baseline activity by increasing its firing frequency. Following this increase, a gastric mill rhythm was elicited and the pyloric cycle period decreased. Lesioning the MCN1 axon prevented these effects. Thus modulatory projection neurons such as MCN1 can control the motor output in vivo, and they participate in the processing of exteroceptive sensory information in behaviorally relevant conditions. stomatogastric nervous system; central pattern generator; sensorimotor; descending control; neuromodulation ONE OF THE BIGGEST CHALLENGES today is to determine how the neuromodulatory system contributes to the neuronal plasticity that allows the nervous system to respond adequately to different behavioral tasks (Gu 2002;Calabrese 2003;Krichmar 2008;Brezina 2010). In particular, motor pattern generating circuits show a high degree of flexibility due to neuromodulatory substances that trigger new patterns or modify ongoing activity (Morgan et al. 2002;Nusbaum and Beenhakker 2002;Marder et al. 2005;Dickinson 2006;Sakurai and Katz 2009). Recent studies (Chen et al. 2009) have addressed the complexity and variety of paracrine neuromodulator release in vivo, but much less is known about the in vivo activity of neuromodulatory neurons that control behavior. In general, motor networks are governed by descending neurons from higher order circuits that are involved in decision making or relay the appropriate decisive information. In lamprey, fish, and tadpole, reticulospinal neurons in...