Myoblast fusion: When it takes more to make one

Cell-cell fusion is a crucial and highly regulated event in the genesis of both form and function of many tissues. One particular type of cell fusion, myoblast fusion, is a key cellular process that shapes the formation and repair of muscle. Despite its importance for human health, the mechanisms underlying this process are still not well understood. The purpose of this review is to highlight the recent literature pertaining to myoblast fusion and to focus on a comparison of these studies across several model systems, particularly the fly, zebrafish and mouse. Advances in technical analysis and imaging have allowed identification of new fusion genes and propelled further characterization of previously identified genes in each of these systems. Among the cellular steps identified as critical for myoblast fusion are migration, recognition, adhesion, membrane alignment and membrane pore formation and resolution. Importantly, striking new evidence indicates that orthologous genes govern several of these steps across these species. Taken together, comparisons across three model systems are illuminating a once elusive process, providing exciting new insights and a useful framework of genes and mechanisms.     

ANKRD13C acts as a molecular chaperone for G protein-coupled receptors

Although the mechanisms that regulate folding and maturation of newly synthesized G protein-coupled receptors are crucial for their function, they remain poorly characterized. By yeast two-hybrid screening, we have isolated ANKRD13C, a protein of unknown function, as an interacting partner for the DP receptor for prostaglandin D(2). In the present study we report the characterization of this novel protein as a regulator of DP biogenesis and trafficking in the biosynthetic pathway. Co-localization by confocal microscopy with an endoplasmic reticulum (ER) marker, subcellular fractionation experiments, and demonstration of the interaction between ANKRD13C and the cytoplasmic C terminus of DP suggest that ANKRD13C is a protein associated with the cytosolic side of ER membranes. Co-expression of ANKRD13C with DP initially increased receptor protein levels, whereas siRNA-mediated knockdown of endogenous ANKRD13C decreased them. Pulse-chase experiments indicated that ANKRD13C can promote the biogenesis of DP by inhibiting the degradation of newly synthesized receptors. However, a prolonged interaction between ANKRD13C and DP resulted in ER retention of misfolded/unassembled forms of the receptor and to their proteasome-mediated degradation. ANKRD13C also regulated the expression of other GPCRs tested (CRTH2, thromboxane A(2) (TPα), and β2-adrenergic receptor), whereas it did not affect the expression of green fluorescent protein, GRK2 (G protein-coupled receptor kinase 2), and VSVG (vesicular stomatitis virus glycoprotein), showing specificity toward G protein-coupled receptors. Altogether, these results suggest that ANKRD13C acts as a molecular chaperone for G protein-coupled receptors, regulating their biogenesis and exit from the ER.     

Sympathetic control after cardiac resynchronization therapy: responders versus nonresponders

Cardiac resynchronization therapy (CRT) decreases muscle sympathetic nerve activity (MSNA) in patients with severe congestive heart failure (CHF) and cardiac asynchrony. Whether this affects equally patients who clinically respond or not to CRT is unknown. We tested the hypothesis that the favorable effects of CRT on MSNA disappear on CRT interruption only in those who respond to CRT. Twenty-three consecutive CHF patients participated in the study, among whom 16 presented a symptomatic improvement by one or more New York Heart Association (NYHA) functional classes 15 +/- 5 mo after CRT (responders), and seven had not improved after 12 +/- 4 mo of CRT (nonresponders). MSNA and echocardiographic recordings were obtained in random order during atrio-right ventricular pacing (ARV), without stimulation in patients who were not pacemaker dependent (OFF, n = 17), and during atrio-biventricular pacing (BIV). Responders had a longer 6-min walking distance, a lower NYHA class and brain natriuretic peptide levels, and a better quality of life than did nonresponders (all P < 0.05). MSNA increased by 25 +/- 7% in the responders, whereas it remained unchanged in the nonresponders, when shifting from the BIV mode to a nonsynchronous condition (ARV and OFF modes) (P < 0.01). Cardiac output decreased by 0.7 +/- 0.2 l/min in the responders but did not change when shifting from the BIV mode to the nonsynchronous pacing mode in the nonresponders (P < 0.01). In conclusion, reversible sympathoinhibition is a marker of the clinical response to CRT.     

RGS2 is upregulated by and attenuates the hypertrophic effect of alpha1-adrenergic activation in cultured ventricular myocytes

Regulator of G protein signaling (RGS) proteins counter the effects of G protein-coupled receptors (GPCRs) by limiting the abilities of G proteins to propagate signals, although little is known concerning their role in cardiac pathophysiology. We investigated the potential role of RGS proteins on alpha1-adrenergic receptor signals associated with hypertrophy in primary cultures of neonatal rat cardiomyocytes. Levels of mRNA encoding RGS proteins 1-5 were examined, and the alpha1-adrenergic agonist phenylephrine (PE) significantly increased RGS2 gene expression but had little or no effect on the others. The greatest changes in RGS2 mRNA occurred within the first hour of agonist addition. We next investigated the effects of RGS2 overexpression produced by infecting cells with an adenovirus encoding RGS2-cDNA on cardiomyocyte responses to PE. As expected, PE increased cardiomyocyte size and also significantly upregulated alpha-skeletal actin and ANP expression, the markers of hypertrophy, as well as the Na-H exchanger 1 isoform. These effects were blocked in cells infected with the adenovirus expressing RGS2. We also examined hypertrophy-associated MAP kinase pathways, and RGS2 overexpression completely prevented the activation of ERK by PE. In contrast, the activation of both JNK and p38 unexpectedly were increased by RGS2, although the ability of PE to further activate the p38 pathway was reduced. These results indicate that RGS2 is an important negative-regulatory factor in cardiac hypertrophy produced by alpha1-adrenergic receptor stimulation through complex mechanisms involving the modulation of mitogen-activated protein kinase signaling pathways.     

An initial genetic linkage map of the rhesus macaque (Macaca mulatta) genome using human microsatellite loci

Rhesus macaques (Macaca mulatta) are the most widely used nonhuman primate species in biomedical research. To create new opportunities for genetic and genomic studies using rhesus monkeys, we constructed a genetic linkage map of the rhesus genome. This map consists of 241 microsatellite loci, all previously mapped in the human genome. These polymorphisms were genotyped in five pedigrees of rhesus monkeys totaling 865 animals. The resulting linkage map covers 2048 cM including all 20 rhesus autosomes, with average spacing between markers of 9.3 cM. Average heterozygosity among those markers is 0.73. This linkage map provides new comparative information concerning locus order and interlocus distances in humans and rhesus monkeys. The map will facilitate whole-genome linkage screens to locate quantitative trait loci (QTLs) that influence individual variation in phenotypic traits related to basic primate anatomy, physiology, and behavior, as well as QTLs relevant to risk factors for human disease.     

Dibutyl phthalate, the bioactive compound produced by Streptomyces albidoflavus 321.2

It was found that the bioactive compound, dibutyl phthalate, was produced by a new soil isolate Streptomyces albidoflavus 321.2. Once this active compound was recovered by ethyl acetate from the fermented broth, being possible to isolate 13.4 mg/l, it was purified by paper, silica gel column, thin layer and gas chromatography. Structure was determined by analysing UV, IR and GC-MS spectra. During analysis, such active compound showed strong activity against gram-positive and gram-negative bacteria, as well as unicellular and filamentous fungi. The antimicrobial activity of the compound was reversed by the amino acid proline. No acute toxicity was observed.     

Cyclic strain increases fibroblast proliferation, matrix accumulation, and elastic modulus of fibroblast-seeded polyurethane constructs

Rapid induction of matrix production and mechanical strengthening is essential to the development of bio-artificial constructs for repair and replacement of load-bearing connective tissues. Toward this end, we describe the development of a mechanical bioreactor and its application to investigate the influence of cyclic strain on fibroblast proliferation, matrix accumulation, and the mechanical properties of fibroblast-seeded polyurethane constructs (FSPC). Human fibroblasts were cultured in 10% serum-containing conditions within three-dimensional, porous elastomeric substrates under static conditions and a model regime of cyclic strain (10% strain, 0.25 Hz, 8 h/day), with and without ascorbic acid supplementation. After one week, the combination of cyclic strain and ascorbic acid resulted in significantly increased construct elastic modulus (>110%) relative to either condition alone. In contrast, cyclic strain alone was sufficient to stimulate significant increases in fibroblast proliferation. Mechanical strengthening of FSPCs was accompanied by increased type I collagen and fibronectin matrix accumulation and distribution, and significantly increased gene expression for type I collagen, TGFbeta-1, and CTGF. These results suggest that strain-induced conditioning in vitro leads to mechanical strengthening of fibroblast/material constructs, most likely resulting from increased collagen matrix deposition, secondary to strain-induced increases in cytokine production.     

Improving the biological control of leafminers (Diptera: Agromyzidae) using the sterile insect technique

The leafminer Liriomyza trifolii (Burgess) (Diptera: Agromyzidae) is a worldwide pest of ornamental and vegetable crops. The most promising nonchemical approach for controlling Liriomyza leafminers in greenhouses is regular releases of the parasitoid Diglyphus isaea (Walker) (Hymenoptera: Eulophidae). In the current study, we examine the hypothesis that the use of D. isaea for biological control of leafminers in greenhouse crops may be more practical and efficient when supplemented with additional control strategies, such as the sterile insect technique (SIT). In small cages, our SIT experiments suggest that release of sterile L. trifolii males in three sterile-to-fertile male ratios (3:1, 5:1, and 10:1) can significantly reduce the numbers of the pest offspring. In large cage experiments, when both parasitoids and sterile males were released weekly, the combined methods significantly reduced mine production and the adult leafminer population size. Moreover, a synergistic interaction effect between these two methods was found, and a model based on our observed data predicts that because of this effect, only the use of both methods can eradicate the pest population. Our study indicates that an integrated pest management approach that combines the augmentative release of the parasitoid D. isaea together with sterile leafminer males is more efficient than the use of either method alone. In addition, our results validate previous theoretical models and demonstrate synergistic control with releases of parasitoids and sterile insects.     

Neural tube derived signals and Fgf8 act antagonistically to specify eye versus mandibular arch muscles

Recent knockout experiments in the mouse generated amazing craniofacial skeletal muscle phenotypes. Yet none of the genes could be placed into a molecular network, because the programme to control the development of muscles in the head is not known. Here we show that antagonistic signals from the neural tube and the branchial arches specify extraocular versus branchiomeric muscles. Moreover, we identified Fgf8 as the branchial arch derived signal. However, this molecule has an additional function in supporting the proliferative state of myoblasts, suppressing their differentiation, while a further branchial arch derived signal, namely Bmp7, is an overall negative regulator of head myogenesis.