Expression Of Myf6 Research Articles

Replacement of fishmeal by yellow mealworm meal on the growth performance, feed utilisation and quality of large yellow croaker

An 80-day feeding trial was conducted to evaluate the effects of yellow mealworm (Tenebrio molitor, TM) meal as substitute for dietary fishmeal on the growth performance, feed utilisation and flesh quality of large yellow croaker (initial body weight: 189.18±0.13 g). The control diet (TM0) was designed to contain 56% of fishmeal. Based on the TM0, graded levels of TM meal (15, 30, 45, 60, 75 and 100%, respectively) were used to replace fishmeal to formulate the other six experimental diets (TM15, TM30, TM45, TM60, TM75 and TM100), respectively. The results showed that the survival was not significantly affected by dietary TM meal levels (P>0.05). Compared with control group, the final body weight, weight gain rate and protein efficiency ratio decreased significantly when the replacement level over 30%, while feed conversion ratio increased significantly as replacement level over 45% (P<0.05). The total protein-bound amino acid content in muscle was significantly increased with the increase of dietary TM meal inclusion (P<0.05). With replacement level increasing, the percentage of eicosapentaenoic acid (EPA) and Σn-3/Σn-6 poly-unsaturated fatty acids (PUFA) in muscle significantly decreased (P<0.05). Meanwhile, the skin redness (a*) and yellowness (b*) values in the ventral and bottom of ventral regions showed a decreasing and increasing trend, respectively (P<0.05). The TM100 group showed a higher myofibre diameter and lower myofibre density compared to the control group (P<0.05). Total replacement of fishmeal with TM meal significantly down-regulated and up-regulated the expression of myf6 and mstn, respectively (P<0.05). The contents of inosine-5′-monophosphate and total free amino acids were significantly decreased with the increase of TM meal inclusion (P<0.05). In conclusion, TM meal can replace at least 30% of dietary fishmeal protein without negative effects on the growth, feed utilisation and flesh quality of large yellow croaker.

Characterization of MRF genes and their tissue distributions and analysis of the effects of starvation and refeeding on the expression of these genes in Acipenser dabryanus muscle.

The purpose of the study was to clone the sequences of myogenic regulatory factors in Acipenser dabryanus and explore the changes in their expression during starvation and refeeding in A. dabryanus muscle. One hundred twenty fish (60.532±0.284g) were randomly assigned to four groups (fasted for 0, 3, 7 or 14 d and then refed for 14 d). Our predictions showed that the coding sequences of myod1, myf5, myog and myf6 in A. dabryanus encoded 275, 248, 248 and 243 amino acids, respectively, and the expression of the four genes was the highest in muscle. During fasting, the expression of myod1 in muscle was significantly decreased in the 14 d group. The expressions of myf5 and myf6 were increased significantly at first and then decreased with prolonged starvation time. The expression of myog in the 14 d group was significantly decreased compared with other groups (P<0.05). During refeeding, the highest values of myod1 and myf6 expression were found in the 3 d group (P<0.05).The expressions of myf5 and myog in 0 d and 3 d group were significantly higher than those in 7 d and 14 d group (P<0.05). These results indicate that myogenic regulatory factors (MRFs) play important roles in muscle growth and development in A. dabryanus. The inhibition of long-term starvation (14 d) on the expression of myogenic regulatory factors is probably one of the reasons why it can not achieve full compensation for growth.

Possible application of muscle specific conditional mouse-derived induced pluripotent stem cells for muscle research

The Cre-driver mouse line, which allows for in vivo regulation of target gene(s) in specific cells, is an indispensable tool for recent muscle research. In this study, I aimed to explore new applications of muscle specific Cre-driver mouse line in muscle research. For this purpose, I generated an iPS cells from a myofiber specific conditional mouse with tamoxifen inducible GFP expression, and then I checked whether homologous recombination was induced in the iPS-derived myogenic cells by tamoxifen administration. Fibroblasts were isolated from the tails of Myf6CE/wt::CAG-EGFP mice, which expressed GFP specifically in Myf6 lineages by tamoxifen injection, and then iPS cells was generated by transfection with a vector based on sendai-virus and containing OSKM genes. Muscle specific conditional mouse-derived iPS cells (mCM-iPSCs) were successfully differentiated to myogenic cells, such as Pax7+ muscle progenitors, MyoD+ myoblasts, and MHC+ myotubes, under myogenic differentiation conditions. Using this model, I examined whether homologous recombination was induced in mCM-iPSC-derived myotubes by 4-hydroxytamoxifen (4OH-TAM) administration. As a result, multinucleated myotubes showed GFP expression, while no GFP signals were detected in both Pax7+ muscle progenitor and non-myogenic cells. These results indicated that homologous recombination could be induced in mCM-iPSC–derived myotubes by tamoxifen administration, and that this system operated normally even in reprogrammed cells. Also, I evidenced that GFP reporter was expressed in myoblasts in addition to multinucleated myotubes when tamoxifen-pulse was applied at an early phase of myogenesis. Taken together, Myf6CE/wt::CAG-EGFP mouse-derived iPS cells reproduced at least in part Myf6 expression during mouse myogenesis. This study demonstrated a novel application of muscle specific conditional mouse in addition to in vivo application, and mCM-iPSCs could also be used in in vitro investigations with muscle specific conditional knock-out mouse.

Effects of polymorphism in the bovine PTPRQ gene on the expression of MYF6 and MYF5 genes in skeletal muscle and on meat production traits in beef bulls

Effects of polymorphism in the bovine PTPRQ gene on the expression of MYF6 and MYF5 genes in skeletal muscle and on meat production traits in beef bulls

Myostatin knockdown and its effect on myogenic gene expression program in stably transfected goat myoblasts

Myostatin, a negative regulator of skeletal muscle mass, is a proven candidate to modulate skeletal muscle mass through targeted gene knockdown approach. Here, we report myostatin (MSTN) knockdown in goat myoblasts stably expressing small hairpin RNA (shRNAs) against MSTN gene through lentivirus vector-mediated integration. We observed 72% (p = 0.003) and 54% (p = 0.022) downregulation of MSTN expression with sh2 shRNA compared to empty vector control and untransduced myoblasts, respectively. The knockdown of MSTN expression was accompanied with concomitant downregulation of myogenic regulatory factor MYOD (77%, p = 0.001), MYOG (94%, p = 0.000), and MYF5 (36%, p = 0.000), cell cycle regulator p21 (62%, p = 0.000), MSTN receptor ACVR2B (23%, p = 0.061), MSTN antagonist follistatin (81%, p = 0.000), and downstream signaling mediators SMAD2 (20%, p = 0.060) and SMAD3 (49%, p = 0.006). However, the expression of MYF6 was upregulated by 14% compared to control lentivirus-transduced myoblasts (p = 0.354) and 79% compared to untransduced myoblasts (p = 0.018) in sh2 shRNA-transduced goat myoblasts cells. Although, MSTN knockdown led to sustained cell proliferation of myoblasts, the myoblasts fusion was suppressed in both MSTN knocked down and control lentivirus-transduced myoblasts. The expression of interferon response gene OAS1 was significantly upregulated in control lentivirus (10.86-fold; p = 0.000)- and sh2 (1.71-fold; p = 0.002)-integrated myoblasts compared to untransduced myoblasts. Our study demonstrates stable knockdown of MSTN in goat myoblasts cells and its potential for use in generation of transgenic goat by somatic cell nuclear transfer.

Expression of androgen receptor target genes in skeletal muscle

We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor (AR)-regulated genes in in vitro and in vivo models. The expression of the myogenic regulatory factor myogenin was significantly decreased in skeletal muscle from testosterone-treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity (ARΔZF2) versus wildtype mice, demonstrating that myogenin is repressed by the androgen/AR pathway. The ubiquitin ligase Fbxo32 was repressed by 12 h dihydrotestosterone treatment in human skeletal muscle cell myoblasts, and c-Myc expression was decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle, and increased in ARΔZF2 muscle. The expression of a group of genes that regulate the transition from myoblast proliferation to differentiation, Tceal7, p57Kip2, Igf2 and calcineurin Aa, was increased in ARΔZF2 muscle, and the expression of all but p57Kip2 was also decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle. We conclude that in males, androgens act via the AR in part to promote peak muscle mass by maintaining myoblasts in the proliferative state and delaying the transition to differentiation during muscle growth and development, and by suppressing ubiquitin ligase-mediated atrophy pathways to preserve muscle mass in adult muscle.

Myogenic, matrix, and growth factor mRNA expression in human skeletal muscle: Effect of contraction intensity and feeding

We examined short-term (3-hour) and long-term (12-week) training effects after heavy load [HL; 70% 1RM] and light load (LL; 16% 1RM) exercise. mRNA expression of genes involved in skeletal muscle remodeling were analyzed and muscle activity (EMG measurements) was measured. Relative muscle activity differed between HL and LL resistance exercise, whereas median power frequency was even, suggesting an equal muscle-fiber-type recruitment distribution. mRNA expression of Myf6, myogenin, and p21 was mostly increased, and myostatin was mostly depressed by HL resistance exercise. No major differences were seen in atrophy-related genes between HL and LL resistance exercise. No changes were seen over 12-week training for any of the targets. Resistance exercise at LL and HL elevated the expression of genes involved in skeletal muscle hypertrophy, although the greatest response was from HL. However, no long-term effect from either LL or HL resistance exercise was seen on basal levels of the mRNA targets.

Effects of restriction of fetal jaw movement on prenatal development of the temporalis muscle

Jaw movement affects masticatory muscles during the postnatal period. Prenatal jaw movement has also been implicated in the development of the temporomandibular joint; however, its effect on prenatal development of the masticatory muscles has not been extensively analysed. In the present study, we examined the effects of the restriction of fetal jaw movement on the temporalis muscle, a major masticatory muscle, in mice by suturing the maxilla and mandible (sutured group) using an exo utero development system. We compared the morphology of the temporalis muscle between sutured, sham-operated and normal in utero groups. At embryonic day (E) 18.5, the volume of muscle fibres, but not that of connective tissue, in the temporalis muscle was decreased in the sutured group. The E18.5 temporalis muscle in the sutured group appeared morphologically similar to that of the E17.5 in utero group, except for frequent muscle fibre irregularities. By transmission electron microscopy, in the sutured group, the myofibrils were immature and scattered, the nuclei appeared comparatively immature, the mitochondria were expanded in volume with fewer cristae, and cytoplasmic inclusion bodies were frequently observed. Expression of Myf-6, a late myogenic transcription factor, by real-time RT-PCR was not significantly different between the sutured and sham-operated groups. These findings demonstrated approximately 1-day delay in the morphological development of the temporalis muscle in the sutured group, and some abnormalities were observed, although Myf-6 level was not affected in the sutured group. The present study revealed that the prenatal jaw movement influences the development of the temporalis muscle.

Evidence for a Myotomal Hox/Myf Cascade Governing Nonautonomous Control of Rib Specification within Global Vertebral Domains

Hox genes are essential for the patterning of the axial skeleton. Hox group 10 has been shown to specify the lumbar domain by setting a rib-inhibiting program in the presomitic mesoderm (PSM). We have now produced mice with ribs in every vertebra by ectopically expressing Hox group 6 in the PSM, indicating that Hox genes are also able to specify the thoracic domain. We show that the information provided by Hox genes to specify rib-containing and rib-less areas is first interpreted in the myotome through the regional-specific control of Myf5 and Myf6 expression. This information is then transmitted to the sclerotome by a system that includes FGF and PDGF signaling to produce vertebrae with or without ribs at different axial levels. Our findings offer a new perspective of how Hox genes produce global patterns in the axial skeleton and support a redundant nonmyogenic role of Myf5 and Myf6 in rib formation.

EQTL Analysis and Association of MYF6 mRNA Expression with Meat Quality Traits in Pigs

Summary The aim of this research was to measure mRNA expression of porcine MYF6 (myogenic factor 6, also known in the medical literature as herculin and myogenic regulatory factor 4, MRF4) and to perform association study with meat quality traits as well as to unravel the transcriptional regulation of this gene by expression QTL (eQTL) study. For this purpose, Duroc × Pietrain F2 resource population (DuPi; n = 313) were used for association and eQTL study. The mRNA levels in Longissimus dorsi muscle tissue of MYF6 gene were evaluated by using qRT-PCR to identify association between gene expression and meat quality traits as well as to analyse eQTL. The mRNA expression of MYF6 associated with conductivity24L (P<0.01) and pH24L (P<0.1). Expression of MYF6 gene was higher in animals with high pH and conductivity of muscle. Linkage analysis using GridQTL revealed 4 trans-regulated eQTL on four porcine autosomes. Significant eQTL [P<0.01, CW (chromosome-wide)] were found for MYF6 on SSC2. A suggestive eQTL (P<0.05, CW) was identified on SSC8. These results revealed that gene expression of MYF6 associated with the meat quality traits and this gene could be potential functional candidate gene for meat quality traits in pigs. However, the analysis of eQTL also suggested that additional genes encoding for transcription factors (TF) could be considered, via fine-mapping underlying the eQTL peaks, in order to understand interaction among these genes.

Identification and analysis of putative regulatory sequences for the MYF5/ MYF6 locus in different vertebrate species

The myogenic factors ( MYF) 5 and 6 are integral to the initiation and development of skeletal muscle and to the maintenance of its phenotype. Thus, they are candidate genes for growth- and meat quality-related traits. We performed a comparative sequence analysis of the MYF5/ MYF6 locus in swine, cattle, dog, chicken and zebrafish on the basis of structural and functional information from human and mouse. Beside the characterization of upstream regulatory elements recently identified in mice, we demonstrate the existence of further highly conserved elements (E1 to E4) which may play a role in the regulation of MYF5 and MYF6 expression. Comparative sequence analysis of putative regulatory sequences in swine revealed a total of 21 single nucleotide polymorphisms (SNP) including 1 and 6 SNPs new for the promoters of MYF5 and MYF6, respectively. The conserved organization of the locus in vertebrates indicates a common basic mechanism of muscle development. However, the existence of numerous regulatory elements at large distances to MYF5 and MYF6 points to a very complex pattern of the gene regulation with significant differences between species.

Cloning of Human Acetyl-CoA Carboxylase β Promoter and Its Regulation by Muscle Regulatory Factors

The 280-kDa beta-isoform of acetyl-CoA carboxylase (ACCbeta) is predominantly expressed in heart and skeletal muscle, whereas the 265-kDa alpha-isoform (ACCalpha) is the major ACC in lipogenic tissues. The ACCbeta promoter showed myoblast-specific promoter activity and was strongly induced by MyoD in NIH3T3 cells. Serial deletions of the promoter revealed that MyoD acts on the E-boxes located at positions -498 to -403 and on the proximal region including the 5'-untranslated region. Destruction of the E-boxes at positions -498 to -403 by site-directed mutagenesis resulted in a significant decrease of MyoD responsiveness. The "TGAAA" at -32 to -28 and the region around the transcription start site play important roles in basal transcription, probably as a TATA box and an Inr element, respectively. Mutations of another E-box at -14 to -9 and a "GCCTGTCA" sequence at +17 to +24 drastically decreased the MyoD responsiveness. The novel cis-element GCCTGTCA was preferentially bound by MyoD homodimer in EMSA and conferred MyoD responsiveness to a luciferase reporter, which was repressed by the overexpression of E12. This finding is unique since activation via E-boxes is mediated by heterodimers of MyoD and E-proteins. We screened a human skeletal muscle cDNA library to isolate clones expressing proteins that bind to the region around the GCCTGTCA (+8 to +27) sequence, and isolated Myf4 and Myf6 cDNAs. Electrophoretic mobility shift assay showed that recombinant Myf4 and Myf6 bind to this novel cis-element. Moreover, transient expression of Myf6 induced significant activation on the ACCbeta promoter or an artificial promoter harboring this novel cis-element. These findings suggest that muscle regulatory factors, such as MyoD, Myf4, and Myf6, contribute to the muscle-specific expression of ACCbeta via E-boxes and the novel cis-element GCCTGTCA.

Messenger ribonucleic acid expression of the MyoD gene family in muscle tissue at slaughter in relation to selection for porcine growth rate.

Livestock meat production capacity is related to muscle fiber numbers and growth. Muscle fibers develop during early embryonic development from proliferating and differentiating myoblasts. Post-natal muscle growth requires satellite cell proliferation and differentiation. Myoblast and satellite cell proliferation and differentiation is regulated by the genes of the MyoD gene family (myogenin, myf-5, myf-6, and MyoD1). Our aim was to study the mRNA expression of these genes in postnatal muscle tissue in relation to porcine selection for growth rate or leanness. Five boars from a line selected for fast growth (F-line) and five boars from a line selected against backfat thickness (L-line) were slaughtered, and biopsies were taken from 12 muscles. Between-line effects, within-line effects in relation to the performance of the pigs, and muscle-specific effects were studied. Comparing the F-line with the L-line revealed significantly greater myogenin, myf-5, and MyoD1 mRNA expression in some muscles of the F-line. The expression of myf-6 showed a tendency for the opposite effect in some muscles. Muscles were ordered by their muscle-specific growth rate (b-value). Within-line evaluation of the data revealed a systematic muscle effect for the myf-6 expression level in the F-line because higher b-values correlated with increased myf-6 expression level. Backfat thickness was negatively related to myogenin expression in the F-line. A relationship was found between myogenin:MyoD1 mRNA expression ratio and meat color/muscle fiber type composition in the L-line. Furthermore, the myogenin:MyoD1 ratio was greater in muscles from F-line boars than in muscles from L-line boars, which relates to the difference between the lines in muscle fiber type. We conclude that the mRNA levels of the MyoD genes in porcine muscle tissue at slaughter showed different relationships to selection for growth rate when evaluated between selection lines and within selection lines.

Expression of myogenic regulatory factors (MRFs) in human neuromuscular disorders.

Immunohistochemical studies using antibodies to myogenic regulatory factors (MRFs) Myo D, myogenin, myf-5, and myf-6, and transcription factors c-Fos and c-Jun, were performed on muscle biopsies from patients suffering from Duchenne and Becker muscular dystrophies, polymyositis, and denervation atrophy, to investigate whether expression of these factors occurs during degeneration and regeneration of adult muscle fibres. Strong Myo D, myogenin, myf-5 and myf-6 immunoreactivity was observed in the nuclei of small regenerating fibres and satellite cells, as revealed by double-labelling immunohistochemistry with N-CAM antibodies, in Duchenne and Becker muscular dystrophies and in polymyositis. This suggests that the myogenic programme is activated during regeneration of adult human muscle fibres. In addition, strong myf-6 and c-Jun immunoreactivity was found in the cytoplasm of some necrotic muscle fibres in patients with Duchenne and Becker muscular dystrophies and in patients with polymyositis. The latter findings suggest that strong cytoplasmic expression of myf-6 and c-Jun is related to the process of muscle fibre degeneration that occurs in these conditions. Increased Myo D, myogenin, myf-5 and myf-6 immunoreactivity was not observed in the nuclei of denervated muscle fibres, although strong c-Fos and c-Jun immunoreactivity was seen in the nuclei of denervated muscle fibres; this suggests that denervation triggers the expression of these transcription factors. Taken together, these observations demonstrate that MRFs and c-Fos and c-Jun are selectively expressed in different human muscular disorders.

The muscle regulatory gene, Myf-6, has a biphasic pattern of expression during early mouse development.

The spatial and temporal expression pattern of the muscle regulatory gene Myf-6 (MRF4/herculin) has been investigated by in situ hybridization during embryonic and fetal mouse development. Here, we report that the Myf-6 gene shows a biphasic pattern of expression. Myf-6 transcripts are first detected in the most rostral somites of the mouse embryo at 9 d of gestation and accumulate progressively in myotomal cells along the rostro-caudal axis. This expression is transient and Myf-6 mRNA can no longer be detected in myotomal cells after day 12 post coitum (p.c.). In contrast to other muscle determination genes (MyoD1, myogenin, Myf-5), Myf-6 mRNA is not detected in limb buds or visceral arches and skeletal muscle of the mouse embryo (day 8-15 p.c.). In fetal mice, Myf-6 transcripts appear at day 16 p.c. in all skeletal muscles, and the gene continues to be expressed at a high level after birth. These results suggest that early Myf-6 expression may be restricted to a population of myogenic cells that does not contribute to the embryonic muscle masses in limb buds and visceral arches. The reappearance of Myf-6 mRNA in fetal skeletal muscle coincides approximately with secondary muscle fiber formation and the onset of innervation.

Myf-6, a new member of the human gene family of myogenic determination factors: evidence for a gene cluster on chromosome 12.

The Myf-6 gene, a novel member of the human gene family of muscle determination factors has been detected by its highly conserved sequence coding for a putative helix-loop-helix domain. This sequence motif is a common feature of all Myf factors and other regulatory proteins. The new Myf gene is located on human chromosome 12, approximately 6.5 Kb upstream of the Myf-5 locus in a closely linked cluster of myogenic determination genes. Myf-6 cDNAs were isolated from human and mouse skeletal muscle, the only tissue in which expression of the corresponding mRNA was observed. In contrast to human primary muscle cell cultures which express moderate levels of Myf-6 mRNA, most established rodent muscle cell lines completely lack this mRNA. Myogenic 10T1/2 cells, however, induced by the expression of either pEMSV-Myf-4 or pEMSV-Myf-5 activate their endogenous mouse Myf-6 gene. Constitutive expression of Myf-6 cDNA in C3H 10T1/2 fibroblasts establishes the muscle phenotype at a similar frequency to the previously characterized myogenic factors. Moreover, muscle-specific CAT reporter constructs containing either the human myosin light chain (MLC) enhancer or the promoter of the embryonic myosin light chain gene are activated in NIH 3T3 fibroblasts or in CV1 kidney cells by cotransfection of Myf-6 expression vehicles. This transcriptional activation occurs in the absence of any apparent conversion of the cellular phenotype of the recipient cells. Glutathione-S-transferase fusion proteins with Myf-3, Myf-4 or Myf-5 specifically bind to a MEF-like consensus sequence present in the human MLC enhancer and the MLC1 emb promoter. In contrast, the Myf-6 hybrid protein interacts weakly with the same sequences showing lower affinity and reduced specificity. Since co-expressed pEMSV-Myf-6, nevertheless, is able to activate transcription of the MLC-CAT reporter constructs in non-muscle tissue culture cells, the different DNA binding properties in vitro might suggest that transactivation of gene expression by Myf-6 involves distinct binding sites and/or additional protein factors.