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American Journal of Physiology.... Jul 2000Myogenin is a muscle-specific transcription factor participating in denervation-induced increases in nicotinic ACh receptor (nAChR) gene expression. Although myogenin... (Comparative Study)
Comparative Study
Myogenin is a muscle-specific transcription factor participating in denervation-induced increases in nicotinic ACh receptor (nAChR) gene expression. Although myogenin RNA expression in denervated muscle is well documented, surprisingly little is known about myogenin protein expression. Therefore, we assayed myogenin protein and RNA in innervated and denervated muscles from young (4 mo) and old (24-32 mo) rats and compared this expression to that of the nAChR alpha-subunit RNA. These assays revealed increased myogenin protein expression within 1 day of denervation, preceding detectable increases in nAChR RNA. By 3 days of denervation, myogenin and nAChR alpha-subunit RNA were increased 500- and 130-fold, respectively, whereas myogenin protein increased 14-fold. Interestingly, old rats (32 mo) had 6-fold higher myogenin protein and approximately 80-fold higher mRNA levels than young rats. However, after denervation, expression levels were similar for young and old animals. The increased myogenin expression during aging, which tends to localize to small fibers, likely reflects spontaneous denervation and/or regeneration. Our results show that increased myogenin protein in denervated muscles correlates with the upregulation of its mRNA.
Topics: Aging; Animals; Blotting, Western; Cell Nucleus; Immunohistochemistry; Muscle Denervation; Muscle Fibers, Skeletal; Muscle, Skeletal; Myogenin; RNA, Messenger; Rats; Rats, Wistar; Receptors, Cholinergic
PubMed: 10896880
DOI: 10.1152/ajpregu.2000.279.1.R179 -
Skeletal Muscle Feb 2018Tyrosine kinase inhibitors (TKIs) are effective therapies with demonstrated antineoplastic activity. Nilotinib is a second-generation FDA-approved TKI designed to...
BACKGROUND
Tyrosine kinase inhibitors (TKIs) are effective therapies with demonstrated antineoplastic activity. Nilotinib is a second-generation FDA-approved TKI designed to overcome Imatinib resistance and intolerance in patients with chronic myelogenous leukemia (CML). Interestingly, TKIs have also been shown to be an efficient treatment for several non-malignant disorders such fibrotic diseases, including those affecting skeletal muscles.
METHODS
We investigated the role of Nilotinib on skeletal myogenesis using the well-established C2C12 myoblast cell line. We evaluated the impact of Nilotinib during the time course of skeletal myogenesis. We compared the effect of Nilotinib with the well-known p38 MAPK inhibitor SB203580. MEK1/2 UO126 and PI3K/AKT LY294002 inhibitors were used to identify the signaling pathways involved in Nilotinib-related effects on myoblast. Adult primary myoblasts were also used to corroborate the inhibition of myoblasts fusion and myotube-nuclei positioning by Nilotinib.
RESULTS
We found that Nilotinib inhibited myogenic differentiation, reducing the number of myogenin-positive myoblasts and decreasing myogenin and MyoD expression. Furthermore, Nilotinib-mediated anti-myogenic effects impair myotube formation, myosin heavy chain expression, and compromise myotube-nuclei positioning. In addition, we found that p38 MAPK is a new off-target protein of Nilotinib, which causes inhibition of p38 phosphorylation in a similar manner as the well-characterized p38 inhibitor SB203580. Nilotinib induces the activation of ERK1/2 and AKT on myoblasts but not in myotubes. We also found that Nilotinib stimulates myoblast proliferation, a process dependent on ERK1/2 and AKT activation.
CONCLUSIONS
Our findings suggest that Nilotinib may have important negative effects on muscle homeostasis, inhibiting myogenic differentiation but stimulating myoblasts proliferation. Additionally, we found that Nilotinib stimulates the activation of ERK1/2 and AKT. On the other hand, we suggest that p38 MAPK is a new off-target of Nilotinib. Thus, there is a necessity for future studies to investigate the long-term effects of TKIs on skeletal muscle homeostasis, along with potential detrimental effects in cell differentiation and proliferation in patients receiving TKI therapies.
Topics: Animals; Apoptosis; Cell Differentiation; Cell Proliferation; Cells, Cultured; Gene Expression Regulation; MAP Kinase Signaling System; Mice, Inbred C57BL; Muscle Development; Muscle Fibers, Skeletal; Myoblasts, Skeletal; Myogenin; Phosphorylation; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Proteostasis; Proto-Oncogene Proteins c-akt; Pyrimidines; RNA, Messenger; p38 Mitogen-Activated Protein Kinases
PubMed: 29463296
DOI: 10.1186/s13395-018-0150-5 -
Genes To Cells : Devoted To Molecular &... May 2010We established cardiac pluripotent stem-like cells from the left atrium (LA-PCs) of adult rat hearts. These cells could differentiate not only into beating myocytes but...
We established cardiac pluripotent stem-like cells from the left atrium (LA-PCs) of adult rat hearts. These cells could differentiate not only into beating myocytes but also into cells of other lineages, including adipocytes and endothelial cells in the methylcellulose-based medium containing interleukin-3 (IL-3), interleukin-6 (IL-6), and stem cell factor (SCF). In particular, IL-3 and SCF contributed to the differentiation into cardiac troponin I-positive cells. Notably, small population of LA-PCs coexpressed GATA4 and myogenin, which are markers specific to cardiomyocytes and skeletal myocytes, respectively, and could differentiate into both cardiac and skeletal myocytes. Therefore, we investigated the involvement of these two tissue-specific transcription factors in the cardiac transcriptional activity. Coexpression of GATA4 and myogenin synergistically activated GATA4-specific promoter of the atrial natriuretic peptide gene. This combinatorial function was shown to be dependant on the GATA site, but independent of the E-box. The results of chromatin immunoprecipitation and electrophoretic mobility shift assays suggested that myogenin bound to GATA4 on the GATA elements and the C-terminal Zn-finger domain of GATA4 and the N-terminal region of myogenin were required for this synergistic activation of transcription. Taken together, these two transcription factors could be involved in the myogenesis of LA-PCs.
Topics: Adult Stem Cells; Animals; Atrial Natriuretic Factor; Cells, Cultured; Cytokines; GATA4 Transcription Factor; Gene Expression Regulation; Heart Atria; Interleukin-3; Male; Muscle Development; Myogenin; Pluripotent Stem Cells; Promoter Regions, Genetic; Proto-Oncogene Proteins c-kit; Rats; Stem Cell Factor
PubMed: 20384792
DOI: 10.1111/j.1365-2443.2010.01394.x -
Developmental Biology Jun 1995Tissue culture studies using muscle cell lines suggest that in addition to mitogenic effects, fibroblast growth factors (FGF) inhibit skeletal muscle differentiation and...
Tissue culture studies using muscle cell lines suggest that in addition to mitogenic effects, fibroblast growth factors (FGF) inhibit skeletal muscle differentiation and the expression of members of a family of muscle-specific regulatory genes including MyoD and myogenin. We examined the possible coexpression of bFGF and myogenin by tandem in situ hybridization (detecting mRNA) and immunocytochemistry studies (detecting protein) to determine whether myogenic cells in vivo endogenously produce bFGF. Mdx mouse muscle, which shows characteristic dystrophic damage and regeneration, demonstrated mononuclear cells containing myogenin and bFGF transcripts in similar regions of adjacent sections of focal degeneration and repair, particularly near recent segmental fiber damage. Using immunocytochemistry and in situ hybridization concurrently on the same sections, bFGF protein and myogenin mRNA were colocalized in both muscle precursors and new myotubes. The in vivo results were confirmed in vitro using primary explant cultures of mdx muscle. Approximately one-half of mononuclear cells in vivo were myogenic by the criterion of myogenin mRNA expression. Both myogenin and bFGF mRNAs were also colocalized with bFGF protein, indicating endogenous expression of bFGF in a subpopulation of myogenic cells. Small numbers of myogenic mononuclear cells were differentiated, as determined by the presence of developmental myosin heavy chain protein (DevMHC). These cells and new myotubes also colocalized myogenin, DevMHC, and bFGF. Since bFGF and myogenin are colocalized in mpec and myotubes in vivo and in vitro, endogenous expression of bFGF is not mutually exclusive of myogenic regulatory gene expression, either before or after differentiation of the skeletal muscle phenotype. Such features of coexpression suggest an important and complex role for bFGF in muscle regeneration in vivo.
Topics: Animals; Cell Differentiation; Culture Techniques; Fibroblast Growth Factor 2; In Situ Hybridization; Mice; Mice, Inbred C57BL; Muscle Development; Muscles; Muscular Dystrophy, Animal; Myogenin; RNA, Messenger
PubMed: 7781901
DOI: 10.1006/dbio.1995.1172 -
Animal : An International Journal of... Jul 2016Poor maternal nutrition inhibits muscle development and postnatal muscle growth. Satellite cells are myogenic precursor cells that contribute to postnatal muscle growth,...
Poor maternal nutrition inhibits muscle development and postnatal muscle growth. Satellite cells are myogenic precursor cells that contribute to postnatal muscle growth, and their activity can be evaluated by the expression of several transcription factors. Paired-box (Pax)7 is expressed in quiescent and active satellite cells. MyoD is expressed in activated and proliferating satellite cells and myogenin is expressed in terminally differentiating cells. Disruption in the expression pattern or timing of expression of myogenic regulatory factors negatively affects muscle development and growth. We hypothesized that poor maternal nutrition during gestation would alter the in vitro temporal expression of MyoD and myogenin in satellite cells from offspring at birth and 3 months of age. Ewes were fed 100% or 60% of NRC requirements from day 31±1.3 of gestation. Lambs from control-fed (CON) or restricted-fed (RES) ewes were euthanized within 24 h of birth (birth; n=5) or were fed a control diet until 3 months of age (n=5). Satellite cells isolated from the semitendinosus muscle were used for gene expression analysis or cultured for 24, 48 or 72 h and immunostained for Pax7, MyoD or myogenin. Fusion index was calculated from a subset of cells allowed to differentiate. Compared with CON, temporal expression of MyoD and myogenin was altered in cultured satellite cells isolated from RES lambs at birth. The percent of cells expressing MyoD was greater in RES than CON (P=0.03) after 24 h in culture. After 48 h of culture, there was a greater percent of cells expressing myogenin in RES compared with CON (P0.05). In satellite cells from RES lambs at 3 months of age, the percent of cells expressing MyoD and myogenin were greater than CON after 72 h in culture (P<0.05). Fusion index was reduced in RES lambs at 3 months of age compared with CON (P<0.001). Restricted nutrition during gestation alters the temporal expression of myogenic regulatory factors in satellite cells of the offspring, which may reduce the pool of myoblasts, decrease myoblast fusion and contribute to the poor postnatal muscle growth previously observed in these animals.
Topics: Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Cell Differentiation; Diet; Female; Gene Expression Regulation; Maternal Nutritional Physiological Phenomena; Muscle Development; Myoblasts; Myogenic Regulatory Factors; Myogenin; Pregnancy; Prenatal Exposure Delayed Effects; Satellite Cells, Skeletal Muscle; Sheep
PubMed: 26856892
DOI: 10.1017/S1751731116000070 -
Targeted inactivation of myogenic factor genes reveals their role during mouse myogenesis: a review.The International Journal of... Feb 1996The role of the four myogenic regulating genes Myf-5, myogenin, MyoD, and MRF4 (herculin, Myf-6) during mouse embryogenesis has been investigated by targeted gene... (Review)
Review
The role of the four myogenic regulating genes Myf-5, myogenin, MyoD, and MRF4 (herculin, Myf-6) during mouse embryogenesis has been investigated by targeted gene inactivation. Null mutations for the MyoD gene generate no skeletal muscle phenotype due to a compensatory activation of the Myf-5 gene. Mice carrying a homozygous Myf-5 mutation exert considerably delayed myotome formation with unexpected consequences. While skeletal myogenesis in these mutant mice resumes normally at the onset of MyoD expression, a skeletal defect of the ribs persists. Apparently, Myf-5 and MyoD individually are not absolutely essential for skeletal muscle development, most likely because they have overlapping or redundant functions. In fact, double mutants lacking both, MyoD and Myf-5, fail to develop skeletal musculature and the muscle forming regions seem to be devoid of myoblasts. Homozygous inactivation of the myogenin gene leads to drastically reduced myofiber formation. These mice accumulate apparently normal numbers of myoblasts which are arrested in their terminal differentiation program. Myf-6 null mutant mice exhibit drastically reduced expression of Myf-5 for reasons presently unknown. The phenotype is very similar to Myf-5 mutants with an additional reduction of deep back muscles and minor alterations in sarcomeric protein isoforms. Based on the phenotypes obtained from these various gene "knock-out" mice, we now begin to understand the regulatory network and the homostatic relationship of genes which are critically involved in myogenesis of vertebrates.
Topics: Animals; DNA-Binding Proteins; Gene Expression Regulation, Developmental; Gene Targeting; Homozygote; Mice; Mice, Knockout; Muscle Proteins; Muscle, Skeletal; MyoD Protein; Myogenic Regulatory Factor 5; Myogenic Regulatory Factors; Myogenin; Phenotype; Trans-Activators
PubMed: 8735947
DOI: No ID Found -
BMC Complementary and Alternative... Oct 2019Sarcopenia, the decline of skeletal muscle tissue attributed to primary aging is a major concern in older adults. Flavonoids might have potential benefits by modulating...
BACKGROUND
Sarcopenia, the decline of skeletal muscle tissue attributed to primary aging is a major concern in older adults. Flavonoids might have potential benefits by modulating the regulation of satellite cells, thus preventing muscle loss. Sinensetin (SIN), a citrus methylated flavone with anti-inflammatory and anti-proliferative activity, can enhance lipolysis. The objective of the present study was to investigate whether SIN might have sarcopenia-suppressing effect on satellite cells from thigh and calf muscle tissues of young and old rats.
METHODS
Primary muscle cells were obtained from thigh and calf tissues of young and old group rats by dissection. Obtained satellite cells were incubated with indicated concentrations of SIN (50 and 100 μM) treated and untreated condition in differentiation medium. Morphological changes of cells were examined using a phase-contrast microscope. Protein expression levels of myoD and myogenin were analyzed by Western blot. Cells treated with or without SIN under differentiation condition were also immunocytochemically stained for myogenin and 4',6-diamidino-2-phenylindole (DAPI).
RESULTS
Morphologically, the differentiation extracted satellite cells was found to be more evident in SIN treated group of aged rat's cells than that in SIN untreated group. Expression levels of myoD and myogenin proteins involved in myogenesis were increased upon treatment with SIN.
CONCLUSIONS
Collectively, our results indicate that SIN can alleviate age-related sarcopenia by increasing differentiation rate and protein levels of myoD and myogenin.
Topics: Aging; Animals; Cells, Cultured; Flavonoids; Humans; Male; Muscle Cells; Muscle Development; Muscle, Skeletal; MyoD Protein; Myogenin; Rats; Rats, Sprague-Dawley; Sarcopenia
PubMed: 31660942
DOI: 10.1186/s12906-019-2714-2 -
Molecules and Cells Sep 2017Abnormal differentiation of muscle is closely associated with aging (sarcopenia) and diseases such as cancer and type II diabetes. Thus, understanding the mechanisms...
Abnormal differentiation of muscle is closely associated with aging (sarcopenia) and diseases such as cancer and type II diabetes. Thus, understanding the mechanisms that regulate muscle differentiation will be useful in the treatment and prevention of these conditions. Protein lysine acetylation and methylation are major post-translational modification mechanisms that regulate key cellular processes. In this study, to elucidate the relationship between myogenic differentiation and protein lysine acetylation/methylation, we performed a PCR array of enzymes related to protein lysine acetylation/methylation during C2C12 myoblast differentiation. Our results indicated that the expression pattern of HDAC11 was substantially increased during myoblast differentiation. Furthermore, ectopic expression of HDAC11 completely inhibited myoblast differentiation, concomitant with reduced expression of key myogenic transcription factors. However, the catalytically inactive mutant of HDAC11 (H142/143A) did not impede myoblast differentiation. In addition, wild-type HDAC11, but not the inactive HDAC11 mutant, suppressed MyoD-induced promoter activities of MEF2C and MYOG (Myogenin), and reduced histone acetylation near the E-boxes, the MyoD binding site, of the MEF2C and MYOG promoters. Collectively, our results indicate that HDAC11 would suppress myoblast differentiation via regulation of MyoD-dependent transcription. These findings suggest that HDAC11 is a novel critical target for controlling myoblast differentiation.
Topics: Acetylation; Animals; Binding Sites; Cell Differentiation; Gene Expression Regulation; Histone Deacetylases; Humans; MEF2 Transcription Factors; Mice; Muscle Development; Mutation; MyoD Protein; Myoblasts; Myogenin; Transcription, Genetic
PubMed: 28927261
DOI: 10.14348/molcells.2017.0116 -
The Journal of Biological Chemistry Feb 2014The complex process of skeletal muscle differentiation is organized by the myogenic regulatory factors (MRFs), Myf5, MyoD, Myf6, and myogenin, where myogenin plays a...
The complex process of skeletal muscle differentiation is organized by the myogenic regulatory factors (MRFs), Myf5, MyoD, Myf6, and myogenin, where myogenin plays a critical role in the regulation of the final stage of muscle differentiation. In an effort to investigate the role microRNAs (miRNAs) play in regulating myogenin, a bioinformatics approach was used and six miRNAs (miR-182, miR-186, miR-135, miR-491, miR-329, and miR-96) were predicted to bind the myogenin 3'-untranslated region (UTR). However, luciferase assays showed only miR-186 inhibited translation and 3'-UTR mutagenesis analysis confirmed this interaction was specific. Interestingly, the expression of miR-186 mirrored that of its host gene, ZRANB2, during development. Functional studies demonstrated that miR-186 overexpression inhibited the differentiation of C2C12 and primary muscle cells. Our findings therefore identify miR-186 as a novel regulator of myogenic differentiation.
Topics: 3' Untranslated Regions; Animals; Cell Differentiation; Cell Line; Mice; MicroRNAs; Muscle, Skeletal; Myogenin; RNA-Binding Proteins
PubMed: 24385428
DOI: 10.1074/jbc.M113.507343 -
Toxicology and Applied Pharmacology May 2012Arsenic is a toxicant commonly found in water systems and chronic exposure can result in adverse developmental effects including increased neonatal death, stillbirths,...
Arsenic is a toxicant commonly found in water systems and chronic exposure can result in adverse developmental effects including increased neonatal death, stillbirths, and miscarriages, low birth weight, and altered locomotor activity. Previous studies indicate that 20 nM sodium arsenite exposure to C2C12 mouse myocyte cells delayed myoblast differentiation due to reduced myogenin expression, the transcription factor that differentiates myoblasts into myotubes. In this study, several mechanisms by which arsenic could alter myogenin expression were examined. Exposing differentiating C2C12 cells to 20 nM arsenic increased H3K9 dimethylation (H3K9me2) and H3K9 trimethylation (H3K9me3) by 3-fold near the transcription start site of myogenin, which is indicative of increased repressive marks, and reduced H3K9 acetylation (H3K9Ac) by 0.5-fold, indicative of reduced permissive marks. Protein expression of Glp or Ehmt1, a H3-K9 methyltransferase, was also increased by 1.6-fold in arsenic-exposed cells. In addition to the altered histone remodeling status on the myogenin promoter, protein and mRNA levels of Igf-1, a myogenic growth factor, were significantly repressed by arsenic exposure. Moreover, a 2-fold induction of Ezh2 expression, and an increased recruitment of Ezh2 (3.3-fold) and Dnmt3a (~2-fold) to the myogenin promoter at the transcription start site (-40 to +42), were detected in the arsenic-treated cells. Together, we conclude that the repressed myogenin expression in arsenic-exposed C2C12 cells was likely due to a combination of reduced expression of Igf-1, enhanced nuclear expression and promoter recruitment of Ezh2, and altered histone remodeling status on myogenin promoter (-40 to +42).
Topics: Animals; Arsenites; Cell Line; Enhancer of Zeste Homolog 2 Protein; Gene Expression Regulation; Histone-Lysine N-Methyltransferase; Histones; Insulin-Like Growth Factor I; Mice; Myoblasts; Myogenin; Polycomb Repressive Complex 2; Promoter Regions, Genetic; Sodium Compounds
PubMed: 22426358
DOI: 10.1016/j.taap.2012.03.002