Role In Myogenesis Research Articles

The Pattern of SOX11 During Chicken Limb Development Suggests a Role in Myogenesis

SOX11 has been implicated in several aspects of development including epithelial to mesenchymal interactions, neuronal migration, skeletogenesis, joint maintenance, and hedgehog signaling. As a first step in evaluating its role in limb development, we determined the expression pattern for SOX11 between Hamilton‐Hamburger (HH) stages 19–35 chicken embryos. Initally (HH19), SOX11 expression is detected in the the emerging limb bud, with strong ventral expression centrally. At HH22, dorsal anterior and posterior proximal expression is evident. As the limb develops (HH24), SOX11 expression shifts distally and wanes proximally. At HH25, expression surrounds condensing cartilage, and by HH29, SOX11 expression surrounds and accentuates the elbow and distal phalanges. By HH35, SOX11 has clearly shifted to selected muscles and tendons but is poorly expressed in joint spaces. This is the first report to implicate SOX11 in myogenesis. These findings suggest that SOX11 may play multiple roles during skeletogenesis, including the regulation of selected muscle tracts. Further work is needed to determine whether there is a common role for SOX11 during these various components of musculoskeletal development or whether SOX11 adapts to multiple pleiotropic roles that occur during limb development.

Immunological mechanisms of exosome mediated therapeutic bioactivity in Duchenne muscular dystrophy

Duchenne muscular dystrophy (DMD) is a lethal myodegenerative disease associated with chronic inflammation. Disruption in the balance between pro‐inflammatory and pro‐regenerative macrophage immunophenotypes contributes to disease progression. Standard‐of‐care is currently limited to anti‐inflammatory steroids, which marginally delay disease progression. We have recently characterized the therapeutic benefits of cardiosphere‐derived cells (CDCs), which are cardiac progenitor/stromal cells with immunomodulatory, pro‐myogenic, and anti‐fibrotic properties. CDCs partially reverse the key abnormalities of heart and skeletal muscle in mdx mice, findings which motivated two clinical trials (HOPE‐Duchenne and HOPE‐2) of CDCs in DMD patients. The bioactivity of CDCs is mediated by the secretion of lipid bilayer nanovesicles called exosomes, which transfer genetic material between cells to modify recipient cell behavior. We previously discovered macrophages are a primary feature in skeletal muscle of CDC‐exosome treated mdx mice, suggesting their presence is not pathological. Here, we sought to elucidate the role macrophages play in the repair of dystrophic skeletal muscle. Circulating monocytes, at least in part, traffic fluorescently‐labeled CDC‐exosomes to damaged muscle after intravenous delivery. Labeled CDC‐exosomes are detected in the interstitium adjacent to non‐muscle cells, macrophages, and surviving muscle fibers. Moreover, CDC‐exosomes confer a unique immunophenotype to mdx macrophages (Mexo). Next generation sequencing and pathway/network analysis indicate Mexo share features of both M1 (pro‐inflammatory) and M2 (pro‐regenerative) macrophages. Indeed, media conditioned by Mexo supports both myoblast proliferation and differentiation in vitro. Antibody chip array indicated Mexo secrete many chemokines and cytokines known to promote inflammatory cell recruitment and modulation. In contrast to M1 and M2 macrophages, Mexo secrete tissue inhibitor of matrix metalloproteinase‐2 (TIMP‐2). Interestingly, TIMP‐2 has been shown to promote myogenic differentiation of C2C12 myoblasts and may function in both ECM remodeling and myogenesis in vivo. Sequencing of small RNAs revealed that several miRNAs with known roles in myogenesis are uniquely enriched in Mexo exosomes. Together, these data suggest CDC‐exosomes are trafficked to areas of muscle damage, in part, by circulating monocytes and modulate the mdx macrophage phenotype to support tissue regeneration. We propose that Mexo macrophages mediate their pleiotropic effects via the secretion of cytokines, chemokines, and possibly exosomes. Modulation of mdx macrophage functions by CDC‐exosomes, to support muscle health, may represent a novel approach to treating DMD.Support or Funding InformationMuscular Dystrophy Association, Coalition Duchenne, NIH R01 to EM

The Circular RNA circHUWE1 Sponges the miR-29b-AKT3 Axis to Regulate Myoblast Development

Myogenesis is controlled by a well-established transcriptional hierarchy that coordinates the activities of a set of muscle genes. Recently, roles in myogenesis have been described for non-coding RNAs, including a role of circular RNA (circRNA) to regulate muscle gene expression. However, the functions of circRNA and the underlying mechanism by which circRNAs affect myogenesis remain poorly understood. In this study, we analyzed circRNA high-throughput sequencing results of bovine skeletal muscle samples and constructed a circRNA-miRNA-mRNA network according to the competitive endogenous RNA (ceRNA) theory. The putative circHUWE1-miR-29b-AKT3 network was analyzed and its involvement in myogenesis was confirmed through a series of assays. To assess the potential function of this regulation, bovine myoblasts were infected with overexpression plasmids and small interfering RNAs (siRNAs) that target circHUWE1. Next, cell proliferation, apoptosis, and differentiation were analyzed using Cell Counting Kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU), flow cytometry, western blotting, and qRT-PCR assays. The results suggest that circHUWE1 facilitates bovine myoblast proliferation and inhibits cell apoptosis and differentiation. Next, bioinformatics, dual-luciferase reporter assay, and AGO2 RNA immunoprecipitation (RIP) approaches were used to verify the interaction between circHUWE1, miR-29b, and AKT3. Subsequently, we identified that circHUWE1 could directly interfere with the ability of miR-29b to relieve AKT3 suppression, which ultimately activates the AKT signaling pathway. These findings suggested a new regulatory pathway for bovine skeletal muscle development, and they also expand our understanding of circRNA functions in mammals.

Zfp422 promotes skeletal muscle differentiation by regulating EphA7 to induce appropriate myoblast apoptosis

Zinc finger protein 422 (Zfp422) is a widely expressed zinc finger protein that serves as a transcriptional factor to regulate downstream gene expression, but until now, little is known about its roles in myogenesis. We found here that Zfp422 plays a critical role in skeletal muscle development and regeneration. It highly expresses in mouse skeletal muscle during embryonic development. Specific knockout of Zfp422 in skeletal muscle impaired embryonic muscle formation. Satellite cell-specific Zfp422 deletion severely inhibited muscle regeneration. Myoblast differentiation and myotube formation were suppressed in Zfp422-deleted C2C12 cells, isolated primary myoblasts, and satellite cells. Chromatin Immunoprecipitation Sequencing (ChIP-Seq) revealed that Zfp422 regulated ephrin type-A receptor 7 (EphA7) expression by binding an upstream 169-bp DNA sequence, which was proved to be an enhancer of EphA7. Knocking EphA7 down in C2C12 cells or deleting Zfp422 in myoblasts will inhibit cell apoptosis which is required for myoblast differentiation. These results indicate that Zfp422 is essential for skeletal muscle differentiation and fusion, through regulating EphA7 expression to maintain proper apoptosis.

MicroRNA-206 regulates cell proliferation by targeting G6PD in skeletal muscle.

Skeletal muscle is a major component of body mass and plays a central role in the control of whole-body metabolism in humans and animals. Therefore, elucidation of the underlying mechanisms of skeletal growth and development are expected to lead to the discovery of novel genes and pathways related to muscle disease. miR-206, a skeletal muscle-specific microRNA, plays a crucial role in myogenesis; however, miR-206 is known to function in myogenic differentiation, whether or not it affects muscle cells' proliferation, and the underlying mechanisms are unknown. In this study, we investigated the effect of miR-206 on muscle cell proliferation and differentiation, as well as its effect on myofiber type conversion using mouse C2C12 myoblasts. The results showed that overexpression of miR-206 inhibited cell proliferation and promoted muscle cell differentiation, but it did not affect myofiber type conversion. Intriguingly, we found that overexpression of miR-206 suppressed muscle cell proliferation and induced cell cycle arrest in G0/G1 phase by inhibiting the glucose-6-phosphate dehydrogenase (G6PD) gene. Taken together, we demonstrated that the miR-206-G6PD pathway suppresses muscle cell proliferation, and these findings may facilitate the treatment of muscle diseases.-Jiang, A., Dong, C., Li, B., Zhang, Z., Chen, Y., Ning, C., Wu, W., Liu, H. MicroRNA-206 regulates cell proliferation by targeting G6PD in skeletal muscle.

MicroRNA-323-3p promotes myogenesis by targeting Smad2.

Skeletal muscle is an important and complex organ with multiple biological functions in humans and animals. Proliferation and differentiation of myoblasts are the key steps during the development of skeletal muscle. MicroRNA (miRNA) is a class of 21-nucleotide noncoding RNAs regulating gene expression by combining with the 3'-untranslated region of target messenger RNA. Many studies in recent years have suggested that miRNAs play a critical role in myogenesis. Through high-throughput sequencing, we found that miR-323-3p showed significant changes in the longissimus dorsi muscle of Rongchang pigs in different age groups. In this study, we discovered that overexpression of miR-323-3p repressed myoblast proliferation and promoted differentiation, whereas the inhibitor of miR-323-3p displayed the opposite results. Furthermore, we predicted Smad2 as the target gene of miR-323-3p and found that miR-323-3p directly modulated the expression level of Smad2. Then luciferase reporter assays verified that Smad2 was a target gene of miR-323-3p during the differentiation of myoblasts. These findings reveal that miR-323-3p is a positive regulator of myogenesis by targeting Smad2. This provides a novel mechanism of miRNAs in myogenesis.

Differential expression of myosin heavy chain isoforms in cardiac segments of gnathostome vertebrates\xa0and its evolutionary implications

BackgroundImmunohistochemical studies of hearts from the lesser spotted dogfish, Scyliorhinus canicula (Chondrichthyes) revealed that the pan-myosin heavy chain (pan-MyHC) antibody MF20 homogeneously labels all the myocardium, while the pan-MyHC antibody A4.1025 labels the myocardium of the inflow (sinus venosus and atrium) but not the outflow (ventricle and conus arteriosus) cardiac segments, as opposed to other vertebrates. We hypothesized that the conventional pattern of cardiac MyHC isoform distribution present in most vertebrates, i.e. MYH6 in the inflow and MYH7 in the outflow segments, has evolved from a primitive pattern that persists in Chondrichthyes. In order to test this hypothesis, we conducted protein detection techniques to identify the MyHC isoforms expressed in adult dogfish cardiac segments and to assess the pan-MyHC antibodies reactivity against the cardiac segments of representative species from different vertebrate groups.ResultsWestern and slot blot results confirmed the specificity of MF20 and A4.1025 for MyHC in dogfish and their differential reactivity against distinct myocardial segments. HPLC-ESI-MS/MS and ESI-Quadrupole-Orbitrap revealed abundance of MYH6 and MYH2 in the inflow and of MYH7 and MYH7B in the outflow segments. Immunoprecipitation showed higher affinity of A4.1025 for MYH2 and MYH6 than for MYH7 and almost no affinity for MYH7B. Immunohistochemistry showed that A4.1025 signals are restricted to the inflow myocardial segments of elasmobranchs, homogeneous in all myocardial segments of teleosts and acipenseriforms, and low in the ventricle of polypteriforms.ConclusionsThe cardiac inflow and outflow segments of the dogfish show predominance of fast- and slow-twitch MyHC isoforms respectively, what can be considered a synapomorphy of gnathostomes. The myocardium of the dogfish contains two isomyosins (MYH2 and MYH7B) not expressed in the adult heart of other vertebrates. We propose that these isomyosins lost their function in cardiac contraction during the evolution of gnathostomes, the later acquiring a regulatory role in myogenesis through its intronic miRNA. Loss of MYH2 and MYH7B expression in the heart possibly occurred before the origin of Osteichthyes, being the latter reacquired in polypteriforms. We raise the hypothesis that the slow tonic MYH7B facilitates the peristaltic contraction of the conus arteriosus of fish with a primitive cardiac anatomical design and of the vertebrate embryo.

Guanine-rich RNA binding protein GRSF1 inhibits myoblast differentiation through repressing mitochondrial ROS production

Guanine-rich RNA sequence binding factor 1 (GRSF1) is a member of the RNA-binding protein (RBP) family. GRSF1 regulates RNA metabolism through RNA processing, transport and translation in the cytoplasm and mitochondria. However, its role in myogenesis has not been investigated. Here, we demonstrated that the expression of mitochondrial GRSF1 was negatively related to the differentiation of mouse skeletal myoblasts. Interference with GRSF1 promotes myogenesis both in vitro and in vivo without affecting MyoD expression or cell proliferation. Further studies illustrated that GRSF1 regulated myogenic differentiation through direct targeting of mitochondrial GPX4, a key regulator of the cellular redox status, leading to the modulation of ROS levels, which is important for myogenesis. Our findings underscore a critical function for GRSF1 during skeletal myogenesis linked to its regulation of muscle redox homeostasis.

H3K27me3 Depletion during Differentiation Promotes Myogenic Transcription in Porcine Satellite Cells.

Background: Porcine skeletal muscle satellite cells play important roles in myogenesis and muscle regeneration. Integrated analysis of transcriptome and histone modifications would reveal epigenomic roles in promoting myogenic differentiation in swine. Methods: Porcine satellite cells (PSCs) were isolated and in-vitro cultured from newborn piglets. RNA Sequencing (RNA-Seq) and Chromatin Immunoprecipitation Sequencing (ChIP-Seq) experiments were performed using proliferating cells and terminal myotubes in order to interrogate the transcriptomic profiles, as well as the distribution of histone markers—H3K4me3, H3K27me3, and H3K27ac—and RNA polymerase II. Results: The study identified 917 differentially expressed genes during cell differentiation. The landscape of epigenetic marks was displayed on a genome-wide scale, which had globally shrunken. H3K27me3 reinforcement participated in obstructing the transcription of proliferation-related genes, while its depletion was closely related to the up-regulation of myogenic genes. Furthermore, the degree of H3K27me3 modification was dramatically reduced by 50%, and 139 myogenic genes were upregulated to promote cell differentiation. Conclusions: The depletion of H3K27me3 was shown to promote porcine satellite cell differentiation through upregulating the transcription level of myogenic genes. Our findings in this study provide new insights of the epigenomic mechanisms occurring during myogenic differentiation, and shed light on chromatin states and the dynamics underlying myogenesis.

Circular RNA circHIPK3 Promotes the Proliferation and Differentiation of Chicken Myoblast Cells by Sponging miR-30a-3p.

Circular RNAs and microRNAs widely exist in various species and play crucial roles in multiple biological processes. It is essential to study their roles in myogenesis. In our previous sequencing data, both miR-30a-3p and circular HIPK3 (circHIPK3) RNA, which are produced by the third exon of the HIPK3 gene, were differentially expressed among chicken skeletal muscles at 11 embryo age (E11), 16 embryo age (E16), and 1-day post-hatch (P1). Here, we investigated their potential roles in myogenesis. Proliferation experiment showed that miR-30a-3p could inhibit the proliferation of myoblast. Through dual-luciferase assay and Myosin heavy chain (MYHC) immunofluorescence, we found that miR-30a-3p could inhibit the differentiation of myoblast by binding to Myocyte Enhancer Factor 2 C (MEF2C), which could promote the differentiation of myoblast. Then, we found that circHIPK3 could act as a sponge of miR-30a-3p and exerted a counteractive effect of miR-30a-3p by promoting the proliferation and differentiation of myoblasts. Taking together, our data suggested that circHIPK3 could promote the chicken embryonic skeletal muscle development by sponging miR-30a-3p.

Identification of a novel miR-206-Notch3 pathway regulating mouse myoblasts proliferation

MicroRNAs (miRNAs) are endogenous short non-coding RNAs and exert their function by targeting mRNAs of genes. miRNA-206 (miR-206) is exclusively expressed in adult skeletal muscles and plays an important role in myogenesis. However, the regulatory mechanisms of miR-206 in myoblasts proliferation and differentiation are still limited. In this study, we validated that Notch3 is a direct target gene of mouse miR-206 using dual-luciferase reporter assay and miR-206-overexpressed experiments. Furthermore, we demonstrated that the negative effect of overexpression of miR-206 and knockdown of Notch3 on C2C12 cells proliferation, and found that the effect was produced by controlling the transition of G0/G1 and S phase in C2C12 cell cycle. In summary, our results provide direct evidences that miR-206 regulates skeletal muscle cells proliferation and cell cycle arrest partly by targeting the Notch3 gene, which strengthens our understanding of regulatory mechanisms on miR-206 in skeletal muscle growth and development.

Podocan affects C2C12 myogenic differentiation by enhancing Wnt/β-catenin signaling.

Podocan, a small leucine-rich repeat protein, is a negative regulator of cell proliferation. In this study, we demonstrated that podocan is involved in the differentiation of C2C12 murine myoblasts. Podocan expression increased with the progression of C2C12 differentiation. As expect, siRNA-mediated podocan knockdown inhibited C2C12 differentiation, as indicated by inhibition of MYOG, MYH2, and desmin expression, as well as reductions in the differentiation and fusion indices. Overexpression of podocan using dCas9 technology promoted C2C12 cell differentiation. In addition, supplementation of culture medium with podocan influenced C2C12 differentiation. Podocan knockdown reduced Wnt/β-catenin signaling, characterized by a reduction in the nuclear translocation of β-catenin, whereas podocan overexpression had the opposite effect. Furthermore, treatment with XAV939, an inhibitor of Wnt/β-catenin, reduced the podocan-mediated promotion of C2C12 differentiation. Induction of muscle injury in mice by bupivacaine administration suggested that podocan may play a role in muscle regeneration. In summary, our results suggest that podocan is required for normal C2C12 differentiation and that its role in myogenesis is mediated by the Wnt/β-catenin pathway.

PAX3-FOXO1 escapes miR-495 regulation during muscle differentiation

ABSTRACTPax3 plays an essential role in myogenesis. Previously, we found a tumor-signature chimeric fusion RNA, PAX3-FOXO1 also present during muscle differentiation, raising the possibility of its physiological role. Here we demonstrated that the fusion is needed transiently for muscle lineage commitment. Interestingly, the fusion ortholog was not found in seven mouse muscle differentiation/regeneration systems, nor in other stem cell differentiation systems of another three mammal species. We noticed that Pax3 is expressed at a much lower level in human stem cells, and during muscle differentiation than in other mammals. Given the fact that the fusion and the parental Pax3 share common downstream targets, we reasoned that forming the fusion may be a mechanism for human cells to escape certain microRNA regulation on Pax3. By sequence comparison, we identified 16 candidate microRNAs that may specifically target the human PAX3 3ʹUTR. We used a luciferase reporter assay, examined the microRNAs expression, and conducted mutagenesis on the reporters, as well as a CRISPR/Cas9 mediated editing on the endogenous allele. Finally, we identified miR-495 as a microRNA that specifically targets human PAX3. Examining several other fusion RNAs revealed that the human-specificity is not limited to PAX3-FOXO1. Based on these observations, we conclude that PAX3-FOXO1 fusion RNA is absent in mouse, or other mammals we tested, the fusion RNA is a mechanism to escape microRNA, miR-495 regulation in humans, and that it is not the only human-specific fusion RNA.

Association of MyoD1 Gene Polymorphisms with Meat Quality Traits in Domestic Pigeons (Columba livia).

Myogenic differentiation 1 (MyoD1) belongs to the MyoD family and plays a key role in myogenesis and consequently, in determining muscle fiber characteristics. In this study, single nucleotide polymorphisms (SNPs) in the exons of MyoD1 were identified in 200 domestic pigeons (Columba livia) by direct DNA sequencing, and the association between MyoD1 polymorphisms and meat quality traits was analyzed. We found four novel variations (A2967G, G3044A, A3164C, and C3311G) in exon 3. The SNP A2967G is a synonymous mutation, while the other 3 SNPs are located in the 3′ untranslated region. The analysis revealed 3 genotypes, in which allele A was the predominant allele in the SNP A2967G, while allele B was the predominant allele in the SNPs G3044A and A3164C. The mutations A2967G and G3044A were significantly associated with meat quality traits in pigeon. Pigeons with AA or AB genotypes had higher breast muscle concentrations of inosinic acid and intramuscular fat than those of BB genotype. Moreover, these 2 SNPs had significant effects on MyoD1 mRNA expression. The SNPs A2967G and G3044A were organized into 4 haplotypes, which formed 7 diplotypes. Association analysis showed that the diplotypes were not significantly associated with meat quality traits. Our results implied that associations do exist between MyoD1 gene polymorphisms and meat quality traits in domestic pigeons, and the AA and AB genotypes could be applied as genetic markers in marker-aid pigeon breeding.

Abstract 16995: Inhibition of lncRNA MAAT Contributes to Multiple Types of Muscle Atrophy

Background: Skeletal muscle atrophy is the consequence of many diseases, such as denervation, long-term inactivity, excessive fasting, ageing, and heart failure. Recently, increasing studies have shown that long noncoding RNAs (lncRNAs) play an important role in myogenesis and adult skeletal muscle regeneration. However, if lncRNAs are responsible for muscle atrophy remains largely unclear. Hypothesis: We aimed to select and investigate the role of lncRNAs in muscle atrophy. Methods and Results: LncRNA arrays indicated that several lncRNA transcripts were changed during denervation-induced muscle atrophy in mice. After detecting the expression of lncRNAs in multiple types of muscle atrophy in vivo (denervation, Angiotensin II and fasting induced muscle atrophy) and in vitro (Angiotensin II, H 2 O 2 and TNFα induced muscle atrophy) by qRT-PCRs, we identified that MAAT (Muscle Atrophy-Associated Transcript) was down-regulated in multiple types of muscle atrophy. Then lentivirus of shRNA and overexpression of MAAT were further used to study the role of MAAT in muscle atrophy in vitro and in vivo . We found that knockdown of MAAT promoted muscle atrophy, whereas upregulating MAAT prevented multiple types of muscle atrophy. Mechanistically, we found that MAAT negatively regulated the transcription of microRNA-29b through SOX6, and that MAAT influenced the expression of neighbor gene-MBNL by a cis-regulatory module. Conclusions: Inhibition of LncRNA MAAT contributes to muscle atrophy by increasing miR-29b and repressing MBNL. Overexpressing MAAT might be a novel approach to counteract muscle atrophy.

Promotion of Myoblast Differentiation by Fkbp5 via Cdk4 Isomerization

SUMMARYFkbp5 is a widely expressed peptidyl prolyl isomerase that serves as a molecular chaperone through conformational changes of binding partners. Although it regulates diverse protein functions, little is known about its roles in myogenesis. We found here that Fkbp5 plays critical roles in myoblast differentiation through two mechanisms. First, it sequesters Cdk4 within the Hsp90 storage complex and prevents the formation of the cyclin D1-Cdk4 complex, which is a major inhibitor of differentiation. Second, Fkbp5 promotes cis-trans isomerization of the Thr172-Pro173 peptide bond in Cdk4 and inhibits phosphorylation of Thr172, an essential step for Cdk4 activation. Consistent with these in vitro findings, muscle regeneration is delayed in Fkbp5−/− mice. The related protein Fkbp4 also sequesters Cdk4 within the Hsp90 complex but does not isomerize Cdk4 or induce Thr173 phosphorylation despite its highly similar sequence. This study demonstrates protein isomerization as a critical regulatory mechanism of myogenesis by targeting Cdk4.

Long Noncoding RNA lncMUMA Reverses Established Skeletal Muscle Atrophy following Mechanical Unloading.

Reversing established muscle atrophy following mechanical unloading is of great clinical challenge. Long noncoding RNAs (lncRNAs) have been demonstrated to play important roles in myogenesis. Here we identified a lncRNA (mechanical unloading-induced muscle atrophy-related lncRNA [lncMUMA]) enriched in muscle, which was the most downregulated lncRNA during muscle atrophy development in hindlimb suspension (HLS) mice. The in vitro and in vivo data demonstrated that the decreased expression levels of lncMUMA closely associated with a reduction of myogenesis during mechanical unloading. Mechanistically, lncMUMA promoted myogenic differentiation by functioning as a miR-762 sponge to regulate the core myogenic regulator MyoD in vitro. The enforced expression of lncMUMA relieved the decreases in MyoD protein and muscle mass in miR-762 knockin mice. Therapeutically, the enforced expression of lncMUMA improved the in vitro myogenic differentiation of myoblasts under microgravity simulation, prevented the muscle atrophy development, and reversed the established muscle atrophy in HLS mice. These findings identify lncMUMA as an anabolic regulator to reverse established muscle atrophy following mechanical unloading.

Multifaceted Interweaving Between Extracellular Matrix, Insulin Resistance, and Skeletal Muscle.

The skeletal muscle provides movement and support to the skeleton, controls body temperature, and regulates the glucose level within the body. This is the core tissue of insulin-mediated glucose uptake via glucose transporter type 4 (GLUT4). The extracellular matrix (ECM) provides integrity and biochemical signals and plays an important role in myogenesis. In addition, it undergoes remodeling upon injury and/or repair, which is also related to insulin resistance (IR), a major cause of type 2 diabetes (T2DM). Altered signaling of integrin and ECM remodeling in diet-induced obesity is associated with IR. This review highlights the interweaving relationship between the ECM, IR, and skeletal muscle. In addition, the importance of the ECM in muscle integrity as well as cellular functions is explored. IR and skeletal muscle ECM remodeling has been discussed in clinical and nonclinical aspects. Furthermore, this review considers the role of ECM glycation and its effects on skeletal muscle homeostasis, concentrating on advanced glycation end products (AGEs) as an important risk factor for the development of IR. Understanding this complex interplay between the ECM, muscle, and IR may improve knowledge and help develop new ideas for novel therapeutics for several IR-associated myopathies and diabetes.

Deubiquitinase UCHL1 Regulates Myogenesis in Skeletal Muscles

Myogenesis is crucial for skeletal muscle regeneration in injury and disease. This multi‐step process is closely regulated by several myogenic factors. Timely activation and inactivation of these factors require ubiquitin‐proteasome system (UPS)‐mediated protein degradation. In this aspect, deubiquitinases (DUBs) could also play a role in myogenesis. Ubiquitin C‐terminal hydrolase L1 (UCHL1) is a DUB that is highly expressed in the brain. Though undetectable in adult skeletal muscles, UCHL1 protein level is significantly increased in injured muscles. However, the functional significance of UCHL1 in skeletal muscles is unknown. Using in vitro and in vivo approaches, this study tested a hypothesis that UCHL1 can play a role in skeletal muscle myogenesis and regeneration.Our data show that UCHL1 protein level was gradually downregulated during myoblast C2C12 differentiation. Knockdown of UCHL1 expression significantly reduced C2C12 cell proliferation. In contrast, UCHL1 knockdown markedly accelerated cell differentiation, evidenced by more myotubes in UCHL1 deficient cells, which was accompanied by significant upregulation of myogenic factors.In mouse muscles, UCHL1 was expressed by type I and IIa muscle fibers. Denervation increased UCHL1 protein levels in muscle fibers, which was completely abolished in muscles from the mice with UCHL1 skeletal muscle specific knockout (smKO). smKO mice showed reduced satellite cells in soleus and declined type I fiber number. Moreover, smKO muscles exhibited right shift of fiber distribution and a remarkable loss of type I muscle fiber upon denervation as compared with wildtype ones. These observations further suggest the potential involvement of UCHL1 in myogenesis for the regeneration of type I fibers.Support or Funding InformationNIH grants No. 1R15HL118696 and 1R03AG051926 and USD SSOM BBS bridge grant, CBBRe pilot grant, PQCD pilot grant.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Spindle associated membrane protein 1 (Samp1) is required for the differentiation of muscle cells

Muscles are developed and regenerated in a differentiation process called myogenesis, which involves components of the nuclear envelope. We have investigated Samp1 (Spindle Associated Membrane Protein 1), a transmembrane nuclear envelope protein, which interacts with emerin and lamin A, both of which are linked to Emery-Dreifuss muscular dystrophy (EDMD). We found that the levels of Samp1 increased seven-fold during differentiation of mouse C2C12 muscle progenitor cells. To test if Samp1 could have a role in myogenesis we developed stable C2C12 knockdown cell lines expressing short hairpin RNA targeting Samp1 expression. The Samp1 depleted C2C12 cells displayed normal mobility and normal distribution of emerin and lamin A. However, Samp1 depletion increased ERK signaling and completely blocked differentiation of C2C12 cells, which failed to express myogenic marker proteins and failed to form myotubes. The block in myogenesis in Samp1 depleted cells was completely rescued by ectopic expression of RNAi resistant human Samp1, showing that Samp1 is required for muscle differentiation.