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Real-time PCR analysis of CS1 and CS 2 mRNA expression during the myogenic differentiation of C2C12 cells (day 0 to day 7). Values are expressed as ratios of the basal transcription levels on culture day 0 for CS2.
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The calsarcins comprise a novel family of muscle-specific calcineurin-interaction proteins that play an important role in modulating both the function and substrate specificity of calcineurin in muscle cells. The expression of calsarcin-1 (CS-1) is restricted to slow-twitch skeletal muscle fibres, whereas that of both calsarcin-2 (CS-2) and calsarc...
Citations
... levels of MYOZ1 and MYOZ2 were elevated during myogenic differentiation of mouse C2C12 cells, and NF-κB could bind to the promoter region of MYOZ2, resulting in the up-regulation of MYOZ2 transcription37 . Another study found that MEF2A also binds to the proximal promoter region of MYOZ2, which in turn regulates the expression of MYOZ2 to affect the differentiation of myoblasts 38 . ...
Objective To investigate the difference of typeⅠand typeⅡmyofibers of paraspinal muscles between the concave and convex side of main thoracic curve in adolescent idiopathic scoliosis (AIS) and its regulatory mechanism.
Methods The paraspinal muscle samples of 46 patients with AIS were collected and stained with ATPase. The myofiber parameters were measured and compared between the concave and convex side and among different vertebrae. The correlation between myofiber parameters and clinical data of the patients was analyzed. The paraspinal muscle samples of 5 AIS patients were selected for transcriptome RNA sequencing and RT-qPCR plus Western-blot were used to verify the results. Meanwhile, the correlation between the mRNA expression and myofiber parameters were analyzed. Finally, The expression of microRNAs between the concave and convex side were screened by microRNA sequencing and verified by RT-qPCR. In addition, the correlations between the microRNAs expression and myofiber parameters were analyzed to explore the possibility of microRNA regulating myofiber transformation.
Results There was transformation between typeⅠ and typeⅡ myofibers especially in the apical vertebrae region. Although the transformation may be a secondary change under the different tension load, it is closely related to the progress of curvature in AIS. The expression of MYOZ2 on the convex side was significantly different from that on the concave side, and the MYOZ2 expression was closely related to myofiber parameters.There was a significant difference in the expression of miR-499-5p and miR-133a-3p between the concave and convex side, and the differences of miR-499-5p and miR-133a-3p expression were closely related to to myofiber parameters.
Conclusion In AIS patients, the tension load was higher on the convex side and lower on the concave side, which caused an increased expression of miR-499-5p and miR-133a-3p in the paraspinal muscles on the convex side and a decrease expression on the concave side, which inhibited the expression of MYOZ2 on the convex side and promoted the expression of MYOZ on the concave side, and ultimately resulted in the transformation of type II myofibers to type I on the convex side and the transformation of type I to type II on the concave side.
... Curcumin downregulated ADAMTS18 gene though NF-κB Signaling Pathway [70]. NF-κB can regulate the activity of MYOZ2 gene [71]. In addition, NF-κB played critical roles in the regulation of IL-1β [72] and IL-1β in adipose tissues increased inhibition of FAM13A expression [73]. ...
N6-methyladenosine (m6A) widely participates in various life processes of animals, including disease, memory, growth and development, etc. However, there is no report on m6A regulating intramuscular fat deposition in rabbits. In this study, m6A modification of Hycole rabbit muscle and adipose tissues were detected by MeRIP-Seq. In this case, 3 methylases and 12 genes modified by m6A were found to be significantly different between muscle and adipose tissues. At the same time, we found 3 methylases can regulate the expression of 12 genes in different ways and the function of 12 genes is related to fat deposition base on existing studies. 12 genes were modified by m6A methylase in rabbit muscle and adipose tissues. These results suggest that 3 methylases may regulate the expression of 12 genes through different pathways. In addition, the analysis of results showed that 6 of the 12 genes regulated eight signaling pathways, which regulated intramuscular fat deposition. RT-qPCR was used to validate the sequencing results and found the expression results of RT-qPCR and sequencing results are consistent. In summary, METTL4, ZC3H13 and IGF2BP2 regulated intramuscular fat by m6A modified gene/signaling pathways. Our work provided a new molecular basis and a new way to produce rabbit meat with good taste.
... The expression pattern of Myoz3 is also related to mutations in other muscle-specific genes such as ACTN3; the upregulation of Myoz3 was identified in human carriers of the ACTN3 R577X polymorphism [30]. Differences in the expression pattern among the three members of the Myoz family may also be due to the activation of different promoters [31]. In the current study, we found that Myoz3 mRNA was predominantly expressed in leg and breast muscles. ...
Myozenin3
(Myoz3) has been reported to bind multiple Z-disc proteins and hence play a key role in signal transduction and muscle fiber type differentiation. The purpose of current study is to better understand the basic characteristics ofMyoz3. Firstly, we cloned the ORF (open reading frame) of theMyoz3gene. AA (amino acid) sequence analysis revealed that theMyoz3gene encodes a 26 kDa protein which have 97% identities with that of turkey. Expression profiling showed thatMyoz3mRNA is mainly expressed in leg muscle and breast muscle. Furthermore, we investigatedMyoz3gene polymorphisms in two broiler breeds, the Yellow Bantam (YB) and the Avian. Five SNPs (single nucleotide polymorphisms) were identified in the YB breed and 3 were identified in the Avian breed. Genotypes and haplotype were constructed and their associations with carcass traits were analyzed. In the YB breed, c.516 C>T had a strong effect on both shank bone length and the [Formula: see text] value of breast muscle, and the H1H3 diplotype had the highest FC compared to other diplotypes. The markers identified in this study may serve as useful targets for the marker-assisted selection (MAS) of growth and meat quality traits in chickens.
... G9a primarily represses its target genes through mediation of histone H3 lysine 9 di-methylation (H3K9me2) repression marks. We therefore performed chromatin immunoprecipitation (ChIP) assays for G9a occupancy and H3K9me2 enrichment at the Myom2 and Myoz2 promoters which have been previously characterized as MEF2 targets 20,36 . In control cells, G9a occupancy (Fig. 2c) and its signature H3K9me2 enrichment (Fig. 2d) were apparent at these promoters in undifferentiated myoblasts (D0), and the occupancy was reduced during differentiation (D2). ...
In this study, we demonstrate that the lysine methyltransferase G9a inhibits sarcomere organization through regulation of the MEF2C-HDAC5 regulatory axis. Sarcomeres are essential for muscle contractile function. Presently, skeletal muscle disease and dysfunction at the sarcomere level has been associated with mutations of sarcomere proteins. This study provides evidence that G9a represses expression of several sarcomere genes and its over-expression disrupts sarcomere integrity of skeletal muscle cells. G9a inhibits MEF2C transcriptional activity that is essential for expression of sarcomere genes. Through protein interaction assays, we demonstrate that G9a interacts with MEF2C and its co-repressor HDAC5. In the presence of G9a, calcium signaling-dependent phosphorylation and export of HDAC5 to the cytoplasm is blocked which likely results in enhanced MEF2C-HDAC5 association. Activation of calcium signaling or expression of constitutively active CaMK rescues G9a-mediated repression of HDAC5 shuttling as well as sarcomere gene expression. Our results demonstrate a novel epigenetic control of sarcomere assembly and identifies new therapeutic avenues to treat skeletal and cardiac myopathies arising from compromised muscle function.
... Genomic DNA template was prepared from a landrace pig liver tissue. An 2612 bp amplicon (-2,335 to ?277, with the first nucleotide ''G'' in DNA corresponding to the first base incorporated into RNA designated as ?1) was cloned using LA Taq DNA Polymerase and ligated into the pMD18-T vector (TaKaRa, Dalian, China) for subsequent manipulation as described previously [18]. Briefly, constructs containing variable length of truncated pig APOE promoter were individually amplified using different forward primers and a common reverse primer. ...
Apolipoprotein E (APOE), a component of lipoproteins plays an important role in the transport and metabolism of cholesterol, and is associated with hyperlipoproteinemia and Alzheimer's disease. In order to further understand the characterization of APOE gene, the promoter of APOE gene of Landrace pigs was analyzed in the present study. The genomic structure and amino acid sequence in pigs were analyzed and found to share high similarity in those of human but low similarity in promoter region. Real-time PCR revealed the APOE gene expression pattern of pigs in diverse tissues. The highest expression level was observed in liver, relatively low expression in other tissues, especially in stomach and muscle. Furthermore, the promoter expressing in Hepa 1-6 was significantly better at driving luciferase expression compared with C2C12 cell. After analysis of porcine APOE gene promoter regions, potential transcription factor binding sites were predicted and two GC signals, a TATA box were indicated. Results of promoter activity analysis indicated that one of potential regulatory elements was located in the region -669 to -259, which was essential for a high expression of the APOE gene. Promoter mutation and deletion analysis further suggested that the C/EBPA binding site within the APOE promoter was responsible for the regulation of APOE transcription. Electrophoretic mobility shift assays also showed the binding site of the transcription factor C/EBPA. This study advances our knowledge of the promoter of the porcine APOE gene.
... In order to shed light upon the transcriptional regulation of porcine LYN, we characterized the transcription start site (TSS) of the gene using RT-PCR (Her et al., 1998;Lee et al., 2005;Wang et al., 2007), a procedure that employed an antisense primer located in −56 position (position is relative to the translation start codon) and seven various overlapping sense primers ranging from nt −222 to nt −757 (see Additional File A). RT-PCRs were performed using both pooled DNA and pooled cDNA templates isolated from eleven tissues. ...
Resistance to disease and improvement of product quality are important goals in pig farming. Tyrosine Protein Kinase Lyn (LYN) is one of several Src-family tyrosine kinases in immune cells. This protein functions both as a positive and negative regulator of B cell activation, and regulates signaling pathways through phosphorylation of inhibitory receptors, enzymes and adaptors, which suggested that LYN could be correlated with immunity and can be considered as a candidate gene to study in disease resistance. Until now, the profiles of expression and transcriptional regulation of the LYN gene in pig breeds different in immune capacity remain unclear. Using real-time PCR, it indicated that porcine LYN mRNA expressed mainly in immune organs including the spleen, duodenum and liver. Furthermore, Dahuabai pigs (a kind of Chinese indigenous pig breeds with higher immune capacity) showed significant higher LYN mRNA expression levels than that in Landrace. Methylation analysis indicates that LYN expression levels were associated with the methylation status of the LYN promoter, and methylation of the novel CpG site at − 1268C/− 1267G generated by transposition at − 1267 (A → G) results in up-regulating transcriptional activity of this gene. Interestingly, the base A located in − 1267 mainly exhibited in landrace while the base G mainly in Dahuabai pigs. These results might contribute to study the function of this gene in pig breeding for disease resistance.
... However, the disconnection between Mef2c and Myoz2 gene expression suggests negative regulation by MEF2C in the presence or absence of possibly another regulatory molecule or an alternative, as yet unknown regulatory mechanism independent of MEF2C. For instance NF-κB has previously been shown to be a mediator of Myoz2 transcription but not Myoz1 in C2C12 myogenic cells [38]. NF-κB has been associated with skeletal muscle wasting which regulates protein degradation and the ubiquitin-proteasome pathway [39]. ...
Cancer cachexia is a highly debilitating paraneoplastic disease observed in more than 50% of patients with advanced cancers and directly contributes to 20% of cancer deaths. Loss of skeletal muscle is a defining characteristic of patients with cancer cachexia and is associated with poor survival. The present study reveals the involvement of a myogenic transcription factor Myocyte Enhancer Factor (MEF) 2C in cancer-induced skeletal muscle wasting. Increased skeletal muscle mRNA expression of Suppressor of Cytokine Signaling (Socs) 3 and the IL-6 receptor indicative of active IL-6 signaling was seen in skeletal muscle of mice bearing the Colon 26 (C26) carcinoma. Loss of skeletal muscle structural integrity and distorted mitochondria were also observed using electron microscopy. Gene and protein expression of MEF2C was significantly downregulated in skeletal muscle from C26-bearing mice. MEF2C gene targets myozenin and myoglobin as well as myokinase were also altered during cachexia, suggesting dysregulated oxygen transport capacity and ATP regeneration in addition to distorted structural integrity. In addition, reduced expression of calcineurin was observed which suggested a potential pathway of MEF2C dysregulation. Together, these effects may limit sarcomeric contractile ability and also predispose skeletal muscle to structural instability; associated with muscle wasting and fatigue in cachexia.
... Genomic DNA template was prepared from Bama mini-pig ear tissue. For deletion analyses, an 1840 bp amplicon (-1822 to ?17, with the first nucleotide of the ''ATG'' start codon designated as ?1) was cloned using LA Taq DNA Polymerase and ligated into the pMD18-T vector (TaKaRa, Dalian, China) for subsequent manipulation as described previously [21]. Briefly, constructs containing variable length of truncated pig SelS promoter were individually amplified using four different forward primers and a common reverse primer. ...
Selenoprotein S (SelS), a member of selenoprotein family, plays important regulatory function in inflammation and metabolic diseases. SelS expression is up-regulated response to the inflammatory stimulus in many mammal cells, animal models as well as patients. In order to further understand the function of SelS gene, molecular characterization and transcriptional regulation of SelS from a Bama mini-pig were analyzed in the present study. The results showed that pig SelS encoded a protein of 190 amino acid with estimated molecular weight of 21.23 kDa and pI of 9.526. The genomic structure, promoter and deduced amino acid sequence were analyzed and found to share high similarity with those of human SelS. Pig SelS fusion protein was demonstrated to localize in the cytoplasm by fluorescence microscopy. Real-time PCR revealed the ubiquitous expression pattern of pig SelS in diverse tissues, a high level expression was observed in the liver and lung, relatively low expression in other tissues, especially in muscle. Promoter deletion analysis further suggests that an NF-κB binding site within the SelS promoter is responsible for the up-regulation of SelS transcription.
... Several groups independently and simultaneously identified the calsarcin family and termed it calsarcin ( Frey et al., 2000;Frey and Olson, 2002), FATZ ( Faulkner et al., 2000), myozenin ( Takada et al., 2001) and c4 or f5 ( Ahmad et al., 2000). The calsarcin family consists of 3 members, with CS-1 being expressed in the adult heart and in slowtwitch fibers of skeletal muscle, while CS-2 and CS-3 are exclusively expressed in fast-twitch fibers of skeletal muscle tissue ( Frey et al., 2000;Frey and Olson, 2002; Wang et al., 2007;Frey et al., 2008). Calsarcins are hallmarked by a multitude of Z-disc interaction partners that, in addition t°calcineurin, include α-actinin, LIM domain-binding 3 (LDB3, also known as Cypher, ZASP and Oracle), Telethonin/T-cap, γ-filamin ( Frey et al., 2000;Frey and Olson, 2002) and myotilin (Gontier et al., 2005 In slow-twitch skeletal muscle, the lack of CS-1 led to an increase in calcineurin activity, as a consequence, mice with a null mutation in CS-1 show an excess of slow skeletal muscle fibers ( Frey et al., 2004). ...
This study aimed to investigate polymorphisms of Calsarcin-1 gene and evaluate its effect on carcass traits in 429 samples of 3 Chinese indigenous cattle breeds, namely, Luxi (LX), Nanyang (NY) and Jiaxianred (JXR) breeds, PCR products with a 320 bp fragment of the Calsarcin-1 gene spanning over a part of intron 2, complete exon 3 and a part of intron 4 were amplified and sequenced. A synonymous alteration (NW_001495138:g.16718C>A) in the exon 3 region of the calsarcin-1 gene was detected and PCR-SSCP method was then developed to genotype all of the individuals. The results showed that the allele frequencies of A/C in LX, NY and IXR breeds were 0.1116/0.8884, 0.1591/0.8409 and 0.0947/0.9053, respectively. Least squares analysis revealed a significant effect (P
Through the analysis and prediction of the atmospheric environment in China, in atmospheric pollution as the main research object of environmental pollution, application system dynamics research methods, according to the economic resource and environment of the structure of composite system figure to draw the environment, resource, population and gross domestic product (GDP) causality diagram, establish on atmospheric environment, economy, energy system dynamics simulation model, This model can reflect exhaust emissions, resource exploitation and air pollution and the relationship between GDP, have certain universality and expansion.
