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Biomedical subjects

E N Olson

Publications and source records attributed to E N Olson.

At least 37 records · Page 2Linked to original sources
PubMed2001

Control of muscle development by dueling HATs and HDACs.

Skeletal muscle cells have provided an especially auspicious system in which to dissect the roles of chromatin structure in the control of cell growth, differentiation, and development. The MyoD and MEF2 families of transcription factors act cooperatively to regulate the expression of skeletal muscle-specific genes. Recent studies have shown that these two classes of transcription factors associate with histone acetyltransferases and histone deacetylases to control the activation and repression, respectively, of the muscle differentiation program. Signaling systems that regulate the growth and differentiation of muscle cells act, at least in part, by regulating the intracellular localization and associations of these chromatin remodeling enzymes with myogenic transcription factors. We describe the molecules and mechanisms involved in chromatin remodeling during skeletal muscle development.

Acetyltransferases↗
PubMed2001

Antisocial, an intracellular adaptor protein, is required for myoblast fusion in Drosophila.

Somatic muscle formation in Drosophila requires fusion of muscle founder cells with fusion-competent myoblasts. In a genetic screen for genes that control muscle development, we identified antisocial (ants), a gene that encodes an ankyrin repeat-, TPR repeat-, and RING finger-containing protein, required for myoblast fusion. In ants mutant embryos, founder cells and fusion-competent myoblasts are properly specified and patterned, but they are unable to form myotubes. ANTS, which is expressed specifically in founder cells, interacts with the cytoplasmic domain of Dumbfounded, a founder cell transmembrane receptor, and with Myoblast city, a cytoskeletal protein, both of which are also required for myoblast fusion. These findings suggest that ANTS functions as an intracellular adaptor protein that relays signals from Dumbfounded to the cytoskeleton during myoblast fusion.

Amino Acid Motifs↗
PubMed2001

Identification of a signal-responsive nuclear export sequence in class II histone deacetylases.

Activation of muscle-specific genes by the MEF2 transcription factor is inhibited by class II histone deacetylases (HDACs) 4 and 5, which contain carboxy-terminal deacetylase domains and amino-terminal extensions required for association with MEF2. The inhibitory action of HDACs is overcome by myogenic signals which disrupt MEF2-HDAC interactions and stimulate nuclear export of these transcriptional repressors. Nucleocytoplasmic trafficking of HDAC5 is mediated by binding of the chaperone protein 14-3-3 to two phosphoserine residues (Ser-259 and Ser-498) in its amino-terminal extension. Here we show that HDAC4 and -5 each contain a signal-responsive nuclear export sequence (NES) at their extreme carboxy termini. The NES is conserved in another class II HDAC, HDAC7, but is absent in class I HDACs and the HDAC-related corepressor, MEF2-interacting transcription repressor. Our results suggest that this conserved NES is inactive in unphosphorylated HDAC5, which is localized to the nucleus, and that calcium-calmodulin-dependent protein kinase (CaMK)-dependent binding of 14-3-3 to phosphoserines 259 and 498 activates the NES, with consequent export of the transcriptional repressor to the cytoplasm. A single amino acid substitution in this NES is sufficient to retain HDAC5 in the nucleus in the face of CaMK signaling. These findings provide molecular insight into the mechanism by which extracellular cues alter chromatin structure to promote muscle differentiation and other MEF2-regulated processes.

Amino Acid Sequence↗
PubMed2001

Analysis of the tendon cell fate using Scleraxis, a specific marker for tendons and ligaments.

Little is known about the genesis and patterning of tendons and other connective tissues, mostly owing to the absence of early markers. We have found that Scleraxis, a bHLH transcription factor, is a highly specific marker for all the connective tissues that mediate attachment of muscle to bone in chick and mouse, including the limb tendons, and show that early scleraxis expression marks the progenitor cell populations for these tissues. In the early limb bud, the tendon progenitor population is found in the superficial proximomedial mesenchyme. Using the scleraxis gene as a marker we show that these progenitors are induced by ectodermal signals and restricted by bone morphogenetic protein (BMP) signaling within the mesenchyme. Application of Noggin protein antagonizes this endogenous BMP activity and induces ectopic scleraxis expression. However, the presence of excess tendon progenitors does not lead to the production of additional or longer tendons, indicating that additional signals are required for the final formation of a tendon. Finally, we show that the endogenous expression of noggin within the condensing digit cartilage contributes to the induction of distal tendons.

Animals↗
PubMed2001

The Mef2c gene is a direct transcriptional target of myogenic bHLH and MEF2 proteins during skeletal muscle development.

Members of the MEF2 family of transcription factors are upregulated during skeletal muscle differentiation and cooperate with the MyoD family of myogenic basic helix-loop-helix (bHLH) transcription factors to control the expression of muscle-specific genes. To determine the mechanisms that regulate MEF2 gene expression during skeletal muscle development, we analyzed the mouse Mef2c gene for cis-regulatory elements that direct expression in the skeletal muscle lineage in vivo. We describe a skeletal muscle-specific control region for Mef2c that is sufficient to direct lacZ reporter gene expression in a pattern that recapitulates that of the endogenous Mef2c gene in skeletal muscle during pre- and postnatal development. This control region is a direct target for the binding of myogenic bHLH and MEF2 proteins. Mutagenesis of the Mef2c control region shows that a binding site for myogenic bHLH proteins is essential for expression at all stages of skeletal muscle development, whereas an adjacent MEF2 binding site is required for maintenance but not for initiation of Mef2c transcription. Our findings reveal the existence of a regulatory circuit between these two classes of transcription factors that induces, amplifies and maintains their expression during skeletal muscle development.

Animals↗
PubMed2000

Calsarcins, a novel family of sarcomeric calcineurin-binding proteins.

The calcium- and calmodulin-dependent protein phosphatase calcineurin has been implicated in the transduction of signals that control the hypertrophy of cardiac muscle and slow fiber gene expression in skeletal muscle. To identify proteins that mediate the effects of calcineurin on striated muscles, we used the calcineurin catalytic subunit in a two-hybrid screen for cardiac calcineurin-interacting proteins. From this screen, we discovered a member of a novel family of calcineurin-interacting proteins, termed calsarcins, which tether calcineurin to alpha-actinin at the z-line of the sarcomere of cardiac and skeletal muscle cells. Calsarcin-1 and calsarcin-2 are expressed in developing cardiac and skeletal muscle during embryogenesis, but calsarcin-1 is expressed specifically in adult cardiac and slow-twitch skeletal muscle, whereas calsarcin-2 is restricted to fast skeletal muscle. Calsarcins represent a novel family of sarcomeric proteins that link calcineurin with the contractile apparatus, thereby potentially coupling muscle activity to calcineurin activation.

Actinin↗
PubMed2000

Activation of the myocyte enhancer factor-2 transcription factor by calcium/calmodulin-dependent protein kinase-stimulated binding of 14-3-3 to histone deacetylase 5.

Skeletal muscle differentiation is controlled by interactions between myocyte enhancer factor-2 (MEF2) and myogenic basic helix-loop-helix transcription factors. Association of MEF2 with histone deacetylases (HDAC) -4 and -5 results in repression of MEF2 target genes and inhibition of myogenesis. Calcium/calmodulin-dependent protein kinase (CaMK) signaling promotes myogenesis by disrupting MEF2-HDAC complexes and stimulating HDAC nuclear export. To further define the mechanisms that confer CaMK responsiveness to HDAC4 and -5, we performed yeast two-hybrid screens to identify HDAC-interacting factors. These screens revealed interactions between HDAC4 and members of the 14-3-3 family of proteins, which function as signal-dependent intracellular chaperones. HDAC4 binds constitutively to 14-3-3 in yeast and mammalian cells, whereas HDAC5 binding to 14-3-3 is largely dependent on CaMK signaling. CaMK phosphorylates serines -259 and -498 in HDAC5, which subsequently serve as docking sites for 14-3-3. Our studies suggest that 14-3-3 binding to HDAC5 is required for CaMK-dependent disruption of MEF2-HDAC complexes and nuclear export of HDAC5, and implicate 14-3-3 as a signal-dependent regulator of muscle cell differentiation.

14-3-3 Proteins↗
PubMed2000

Independent signals control expression of the calcineurin inhibitory proteins MCIP1 and MCIP2 in striated muscles.

Calcineurin, a calcium/calmodulin-regulated protein phosphatase, modulates gene expression in cardiac and skeletal muscles during development and in remodeling responses such as cardiac hypertrophy that are evoked by environmental stresses or disease. Recently, we identified two genes encoding proteins (MCIP1 and MCIP2) that are enriched in striated muscles and that interact with calcineurin to inhibit its enzymatic activity. In the present study, we show that expression of MCIP1 is regulated by calcineurin activity in hearts of mice with cardiac hypertrophy, as well as in cultured skeletal myotubes. In contrast, expression of MCIP2 in the heart is not altered by activated calcineurin but responds to thyroid hormone, which has no effect on MCIP1. A approximately 900-bp intragenic segment located between exons 3 and 4 of the MCIP1 gene functions as an alternative promoter that responds to calcineurin. This region includes a dense cluster of 15 consensus binding sites for NF-AT transcription factors. Because MCIP proteins can inhibit calcineurin, these results suggest that MCIP1 participates in a negative feedback circuit to diminish potentially deleterious effects of unrestrained calcineurin activity in cardiac and skeletal myocytes. Inhibitory effects of MCIP2 on calcineurin activity may be pertinent to gene switching events driven by thyroid hormone in striated muscles. The full text of this article is available at http://www. circresaha.org.

Animals↗
PubMed2000

Members of the HRT family of basic helix-loop-helix proteins act as transcriptional repressors downstream of Notch signaling.

The Hairy-related transcription-factor (HRT) genes encode three related basic helix-loop-helix transcription factors that show sequence similarity to the Hairy and Enhancer of split family of transcriptional repressors. HRT proteins are expressed in specific regions of the developing heart, vasculature, pharyngeal arches and somites, and the periodicity of their expression in somitic precursors mirrors that of Notch signaling-related molecules. In the present study, we show that the intracellular domain of the Notch1 receptor (Notch1 IC), which is constitutively active, up-regulates HRT2 expression in 10T(1/2) fibroblasts. Luciferase reporter assays using the regulatory regions of the mouse HRT genes revealed that transcription of all three genes is stimulated by Notch1 IC. The promoters of the HRT genes share homology in a binding site for Suppressor of Hairless [Su(H)], a transcriptional mediator of Notch signaling. A dominant-negative Su(H) mutant abolished Notch-activated HRT2 expression, and mutation of the conserved Su(H) consensus site in the HRT2 promoter attenuated transcriptional activation by Notch. Ectopic expression of HRT proteins also blocked activation of HRT2 expression by Notch1 IC through a mechanism requiring the basic region, but not the conserved carboxyl-terminal YQPW-TEVGAF motif of HRT2. These findings identify HRT genes as downstream targets for Notch signaling and reveal a negative autoregulatory loop whereby HRT proteins repress their own expression through interference with Notch signaling.

Animals↗
PubMed2000

Signal-dependent nuclear export of a histone deacetylase regulates muscle differentiation.

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.

Animals↗
PubMed2000

A genetic blueprint for cardiac development.

Congenital heart disease is the leading non-infectious cause of death in children. It is becoming increasingly clear that many cardiac abnormalities once thought to have multifactorial aetiologies are attributable to mutations in developmental control genes. The consequences of these mutations can be manifest at birth as life-threatening cardiac malformations or later as more subtle cardiac abnormalities. Understanding the genetic underpinnings of cardiac development has important implications not only for understanding congenital heart disease, but also for the possibility of cardiac repair through genetic reprogramming of non-cardiac cells to a cardiogenic fate.

Animals↗
PubMed2000

Regulation of microtubule dynamics and myogenic differentiation by MURF, a striated muscle RING-finger protein.

The RING-finger domain is a novel zinc-binding Cys-His protein motif found in a growing number of proteins involved in signal transduction, ubiquitination, gene transcription, differentiation, and morphogenesis. We describe a novel muscle-specific RING-finger protein (MURF) expressed specifically in cardiac and skeletal muscle cells throughout pre- and postnatal mouse development. MURF belongs to the RING-B-box-coiled-coil subclass of RING-finger proteins, characterized by an NH(2)-terminal RING-finger followed by a zinc-finger domain (B-box) and a leucine-rich coiled-coil domain. Expression of MURF is required for skeletal myoblast differentiation and myotube fusion. The leucine-rich coiled-coil domain of MURF mediates association with microtubules, whereas the RING-finger domain is required for microtubule stabilization and an additional region is required for homo-oligomerization. Expression of MURF establishes a cellular microtubule network that is resistant to microtubule depolymerization induced by alkaloids, cold and calcium. These results identify MURF as a myogenic regulator of the microtubule network of striated muscle cells and reveal a link between microtubule organization and myogenesis.

3T3 Cells↗
PubMed2000

The basic helix-loop-helix transcription factor capsulin controls spleen organogenesis.

Formation of numerous internal organs involves reciprocal epithelial-mesenchymal signaling and subsequent patterning and growth of the organ primordium. Capsulin is a basic helix-loop-helix transcription factor expressed in mesenchymal cells that encapsulate the epithelial primordia of internal organs, including the kidney and lung, as well as the epicardium, which gives rise to the coronary arteries. Capsulin is also expressed in the mesothelium that gives rise to the spleen. We demonstrate that mice homozygous for a capsulin null mutation fail to form a spleen. The homeobox genes Hox11 and Bapx1, shown previously to be essential regulators of spleen organogenesis, and a lacZ reporter introduced into the capsulin locus, were expressed in the early splenic primordium, derived from the splanchnic mesoderm, of homozygous mutant embryos. However, this primordium failed to develop beyond an initial group of precursor cells and underwent rapid apoptosis. The phenotype of capsulin mutant mice demonstrates that capsulin acts within a subpopulation of splanchnic mesodermal cells to control an essential early step in spleen organogenesis that is likely to represent a point of regulatory convergence of the capsulin, Hox11, and Bapx1 genes.

Animals↗
PubMed2000

Thoracic skeletal defects in myogenin- and MRF4-deficient mice correlate with early defects in myotome and intercostal musculature.

Myogenin and MRF4 are skeletal muscle-specific bHLH transcription factors critical for muscle development. In addition to a variety of skeletal muscle defects, embryos homozygous for mutations in myogenin or MRF4 display phenotypes in the thoracic skeleton, including rib fusions and sternal defects. These skeletal defects are likely to be secondary because myogenin and MRF4 are not expressed in the rib cartilage or sternum. In this study, the requirement for myogenin and MRF4 in thoracic skeletal development was further examined. When a hypomorphic allele of myogenin and an MRF4-null mutation were placed together, the severity of the thoracic skeletal defects was greatly increased and included extensive rib cartilage fusion and fused sternebrae. Additionally, new rib defects were observed in myogenin/MRF4 compound mutants, including a failure of the rib cartilage to contact the sternum. These results suggested that myogenin and MRF4 share overlapping functions in thoracic skeletal formation. Spatial expression patterns of skeletal muscle-specific markers in myogenin- and MRF4-mutant embryos revealed early skeletal muscle defects not previously reported. MRF4-/- mice displayed abnormal intercostal muscle morphology, including bifurcation and fusion of adjacent intercostals. myogenin/MRF4-mutant combinations displayed ventral myotome defects, including a failure to express normal levels of myf5. The results suggested that the early muscle defects observed in myogenin and MRF4 mutants may cause subsequent thoracic skeletal defects, and that myogenin and MRF4 have overlapping functions in ventral myotome differentiation and intercostal muscle morphogenesis.

Alleles↗
PubMed2000

MEF2 responds to multiple calcium-regulated signals in the control of skeletal muscle fiber type.

Different patterns of motor nerve activity drive distinctive programs of gene transcription in skeletal muscles, thereby establishing a high degree of metabolic and physiological specialization among myofiber subtypes. Recently, we proposed that the influence of motor nerve activity on skeletal muscle fiber type is transduced to the relevant genes by calcineurin, which controls the functional activity of NFAT (nuclear family of activated T cell) proteins. Here we demonstrate that calcineurin-dependent gene regulation in skeletal myocytes is mediated also by MEF2 transcription factors, and is integrated with additional calcium-regulated signaling inputs, specifically calmodulin-dependent protein kinase activity. In skeletal muscles of transgenic mice, both NFAT and MEF2 binding sites are necessary for properly regulated function of a slow fiber-specific enhancer, and either forced expression of activated calcineurin or motor nerve stimulation up-regulates a MEF2-dependent reporter gene. These results provide new insights into the molecular mechanisms by which specialized characteristics of skeletal myofiber subtypes are established and maintained.

Animals↗
PubMed2000

Signal-dependent activation of the MEF2 transcription factor by dissociation from histone deacetylases.

Myocyte enhancer factor-2 (MEF2) transcription factors control muscle-specific and growth factor-inducible genes. We show that hypertrophic growth of cardiomyocytes in response to phenylephrine and serum is accompanied by activation of MEF2 through a posttranslational mechanism mediated by calcium, calmodulin-dependent protein kinase (CaMK), and mitogen-activated protein kinase (MAPK) signaling. CaMK stimulates MEF2 activity by dissociating class II histone deacetylases (HDACs) from the DNA-binding domain. MAPKs, which activate MEF2 by phosphorylation of the transcription activation domain, maximally stimulate MEF2 activity only when repression by HDACs is relieved by CaMK signaling to the DNA-binding domain. These findings identify MEF2 as an endpoint for hypertrophic stimuli in cardiomyocytes and demonstrate that MEF2 mediates synergistic transcriptional responses to the CaMK and MAPK signaling pathways by signal-dependent dissociation from HDACs.

Animals↗
PubMed2000

Failure of Myf5 to support myogenic differentiation without myogenin, MyoD, and MRF4.

The basic helix-loop-helix (bHLH) transcription factors-MyoD, Myf5, myogenin, and MRF4-can each activate the skeletal muscle-differentiation program in transfection assays. However, their functions during embryogenesis, as revealed by gene-knockout studies in mice, are distinct. MyoD and Myf5 have redundant functions in myoblast specification, whereas myogenin and either MyoD or MRF4 are required for differentiation. Paradoxically, myoblasts from myogenin mutant or MyoD/MRF4 double-mutant neonates differentiate normally in vitro, despite their inability to differentiate in vivo, suggesting that the functions of the myogenic bHLH factors are influenced by the cellular environment and that the specific myogenic defects observed in mutant mice do not necessarily reflect essential functions of these factors. Understanding the individual roles of these factors is further complicated by their ability to cross-regulate one another's expression. To investigate the functions of Myf5 in the absence of contributions from other myogenic bHLH factors, we generated triple-mutant mice lacking myogenin, MyoD, and MRF4. These mice appear to contain a normal number of myoblasts, but in contrast to myogenin or MyoD/MRF4 mutants, differentiated muscle fibers fail to form in vivo and myoblasts from neonates of this triple-mutant genotype are unable to differentiate in vitro. These results suggest that physiological levels of Myf5 are insufficient to activate the myogenic program in the absence of other myogenic factors and suggest that specialized functions have evolved for the myogenic bHLH factors to switch on the complete program of muscle gene expression.

Animals↗