scholarly journals Knockdown of the E3 Ubiquitin ligase UBR5 and its role in skeletal muscle anabolism

2020 ◽  
Author(s):  
Daniel C. Turner ◽  
David C. Hughes ◽  
Leslie M. Baehr ◽  
Robert A. Seaborne ◽  
Mark Viggars ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Muscle Mass ◽  
Ubiquitin Ligase ◽  
Muscle Protein ◽  
E3 Ubiquitin Ligase ◽  
Skeletal Muscle Atrophy ◽  
Human Myotubes ◽  
The Impact

AbstractUBR5 is an E3-ubiquitin-ligase positively associated with anabolism, hypertrophy and recovery from atrophy in skeletal muscle. The precise mechanisms underpinning UBR5’s role in the regulation of skeletal muscle mass remain unknown. The present study aimed to elucidate these mechanisms by silencing the UBR5 gene in-vitro and in-vivo. The siRNA-induced reduction (−77%) in UBR5 gene expression in human myotubes was prevented by mechanical loading, suggesting that UBR5 gene expression may be regulated via mechano-transduction signalling. Therefore, we electroporated a UBR5-RNAi plasmid into mouse tibialis anterior muscle in-vivo to investigate the impact of reduced UBR5 on mechano-transduction signalling MEK/ERK/p90RSK and Akt/p70S6K/4E-BP1/rpS6 pathways. Seven days post UBR5 RNAi electroporation, while reductions in overall muscle mass were not detected, mean CSA of GFP-positive fibers was reduced (−9.5%) and the number of large fibers was lower versus the control. Importantly, UBR5-RNAi significantly reduced total RNA, muscle protein synthesis, ERK1/2 and Akt phosphorylation. Whilst p90RSK phosphorylation significantly increased, total p90RSK protein levels demonstrated a 45% reduction with UBR5-RNAi. Finally, these early signalling events after 7 days of UBR5 knockdown culminated in significant reductions in muscle mass (−4.6%) and larger reductions in fiber CSA (−18.5%) after 30 days. This was associated with increased levels of the phosphatase, PP2Ac, and inappropriate chronic elevation of p70S6K and rpS6 between 7 and 30 days, and corresponding reductions in eIF4e. This study demonstrates UBR5 plays an important role in anabolism/hypertrophy, whereby knockdown of UBR5 culminates in skeletal muscle atrophy.

scholarly journals Knockdown of the E3 ubiquitin ligase UBR5 and its role in skeletal muscle anabolism

2021 ◽  
Vol 320 (1) ◽  
pp. C45-C56
Author(s):  
David C. Hughes ◽  
Daniel C. Turner ◽  
Leslie M. Baehr ◽  
Robert A. Seaborne ◽  
Mark Viggars ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Mass ◽  
Ubiquitin Ligase ◽  
Fluorescent Protein ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Tibialis Anterior Muscle ◽  
E3 Ubiquitin Ligase ◽  
Skeletal Muscle Atrophy ◽  
The Impact

UBR5 is an E3 ubiquitin ligase positively associated with anabolism, hypertrophy, and recovery from atrophy in skeletal muscle. The precise mechanisms underpinning UBR5’s role in the regulation of skeletal muscle mass remain unknown. The present study aimed to elucidate these mechanisms by silencing the UBR5 gene in vivo. To achieve this aim, we electroporated a UBR5-RNAi plasmid into mouse tibialis anterior muscle to investigate the impact of reduced UBR5 on anabolic signaling MEK/ERK/p90RSK and Akt/GSK3β/p70S6K/4E-BP1/rpS6 pathways. Seven days after UBR5 RNAi electroporation, although reductions in overall muscle mass were not detected, the mean cross-sectional area (CSA) of green fluorescent protein (GFP)-positive fibers were reduced (−9.5%) and the number of large fibers were lower versus the control. Importantly, UBR5-RNAi significantly reduced total RNA, muscle protein synthesis, ERK1/2, Akt, and GSK3β activity. Although p90RSK phosphorylation significantly increased, total p90RSK protein levels demonstrated a 45% reduction with UBR5-RNAi. Finally, these early events after 7 days of UBR5 knockdown culminated in significant reductions in muscle mass (−4.6%) and larger reductions in fiber CSA (−18.5%) after 30 days. This was associated with increased levels of phosphatase PP2Ac and inappropriate chronic elevation of p70S6K and rpS6 between 7 and 30 days, as well as corresponding reductions in eIF4e. This study demonstrates that UBR5 plays an important role in anabolism/hypertrophy, whereby knockdown of UBR5 culminates in skeletal muscle atrophy.

scholarly journals UBR5 is a Novel E3 Ubiquitin Ligase involved in Skeletal Muscle Hypertrophy and Recovery from Atrophy

2019 ◽  
Author(s):  
RA Seaborne ◽  
DC Hughes ◽  
DC Turner ◽  
DJ Owens ◽  
LM Baehr ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Muscle Atrophy ◽  
Ubiquitin Ligase ◽  
Protein Level ◽  
E3 Ubiquitin Ligase ◽  
Skeletal Muscle Atrophy ◽  
Muscle Fibres ◽  
E3 Ligases ◽  
Atrophy And Hypertrophy

AbstractWe aimed to investigate a novel and uncharacterised E3 ubiquitin ligase in skeletal muscle atrophy, recovery from atrophy/injury, anabolism and hypertrophy. We demonstrated an alternate gene expression profile for UBR5 versus well characterised E3-ligases, MuRF1/MAFbx, where after atrophy evoked by continuous-low-frequency electrical-stimulation in rats, MuRF1/MAFbx were both elevated yet UBR5 was unchanged. Furthermore, after recovery of muscle mass post tetrodotoxin (TTX) induced-atrophy in rats, UBR5 was hypomethylated and increased at the gene expression level, while a suppression of MuRF1/MAFbx was observed. At the protein level, we also demonstrated a significant increase in UBR5 after recovery of muscle mass from hindlimb unloading in both adult and aged rats, and after recovery from atrophy evoked by nerve crush injury in mice. During anabolism and hypertrophy, UBR5 gene expression increased following acute loading in three-dimensional bioengineered mouse muscle in-vitro, and after chronic electrical-stimulation-induced hypertrophy in rats in-vivo, without increases in MuRF1/MAFbx. Additionally, UBR5 protein abundance increased following functional overload-induced hypertrophy of the plantaris muscle in mice and during differentiation of primary human muscle cells. Finally, in humans, genetic association studies (>700,000 SNPs) demonstrated that the A alleles of rs10505025 and rs4734621 SNPs in the UBR5 gene were strongly associated with larger cross-sectional area of fast-twitch muscle fibres and favoured strength/power versus endurance/untrained phenotypes. Overall, we suggest that UBR5 is a novel E3 ubiquitin ligase that is inversely regulated to MuRF1/MAFbx, is epigenetically regulated, and is elevated at both the gene expression and protein level during recovery from skeletal muscle atrophy and hypertrophy.Key PointsWe have recently identified that a HECT domain E3 ubiquitin ligase, named UBR5, is altered epigenetically (via DNA methylation) after human skeletal muscle hypertrophy, where its gene expression is positively correlated with increasing lean leg mass after training and retraining.In the present study we extensively investigate this novel and uncharacterised E3 ubiquitin ligase (UBR5) in skeletal muscle atrophy, recovery from atrophy and injury, anabolism and hypertrophy.We demonstrated that UBR5 was epigenetically via altered DNA methylation during recovery from atrophy.We also determined that UBR5 was alternatively regulated versus well characterised E3 ligases, MuRF1/MAFbx, at the gene expression level during atrophy, recovery from atrophy and hypertrophy.UBR5 also increased at the protein level during recovery from atrophy and injury, hypertrophy and during human muscle cell differentiation.Finally, in humans, genetic variations of the UBR5 gene were strongly associated with larger fast-twitch muscle fibres and strength/power performance versus endurance/untrained phenotypes.

Functional rice with tandemly repeated Cbl-b ubiquitin ligase inhibitory pentapeptide prevents denervation-induced muscle atrophy in vivo

2021 ◽  
Author(s):  
Kazuhito Akama ◽  
Yasuka Shimajiri ◽  
Kumiko Kainou ◽  
Ryota Iwasaki ◽  
Reiko Nakao ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Mass ◽  
Transgenic Rice ◽  
Muscle Atrophy ◽  
Ubiquitin Ligase ◽  
Critical Role ◽  
Phosphorylation Site ◽  
Skeletal Muscle Atrophy ◽  
Rice Seeds

Abstract Ubiquitin ligase Cbl-b play a critical role in non-loading-mediated skeletal muscle atrophy: Cbl-b ubiquitinates insulin receptor substrate-1 (IRS-1), an important insulin-like growth factor-1 signaling intermediate molecule, leading to its degradation and a resulting loss in muscle mass. We reported that intramuscular injection of a pentapeptide, DGpYMP, which acts as a mimic of the phosphorylation site in IRS-1, significantly inhibited denervation-induced skeletal muscle loss. In order to explore the possibility of the prevention of muscle atrophy by diet therapy, we examined the effects of oral administration of transgenic rice containing Cblin (Cbl-b inhibitor) peptide (DGYMP) on denervation-induced muscle mass loss in frogs. We generated transgenic rice seeds in which 15 repeats of Cblin peptides with a WQ spacer were inserted into the rice storage protein glutelin for expression. A diet of the transgenic rice seeds had significant inhibitory effects on denervation-induced atrophy of the leg skeletal muscles in frogs, compared with those receiving a diet of wild-type rice.

Iron-induced skeletal muscle atrophy involves an Akt-forkhead box O3–E3 ubiquitin ligase-dependent pathway

2016 ◽  
Vol 35 ◽  
pp. 66-76 ◽  
Author(s):  
Yasumasa Ikeda ◽  
Mizuki Imao ◽  
Akiho Satoh ◽  
Hiroaki Watanabe ◽  
Hirofumi Hamano ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Atrophy ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Skeletal Muscle Atrophy ◽  
Forkhead Box ◽  
Dependent Pathway

scholarly journals The E3 Ubiquitin Ligase TRAF6 Intercedes in Starvation-Induced Skeletal Muscle Atrophy through Multiple Mechanisms

2012 ◽  
Vol 32 (7) ◽  
pp. 1248-1259 ◽  
Author(s):  
P. K. Paul ◽  
S. Bhatnagar ◽  
V. Mishra ◽  
S. Srivastava ◽  
B. G. Darnay ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Atrophy ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Skeletal Muscle Atrophy

scholarly journals A constitutively monomeric UVR8 photoreceptor allele confers enhanced UV-B photomorphogenesis

2020 ◽  
Author(s):  
Roman Podolec ◽  
Kelvin Lau ◽  
Timothée B. Wagnon ◽  
Michael Hothorn ◽  
Roman Ulm
Keyword(s):  
Gene Expression ◽  
Crystal Structure ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Genetic Tool ◽  
Loop Region ◽  
Extreme Uv ◽  
Insight Into

AbstractThe plant UV-B photoreceptor UVR8 plays an important role in UV-B acclimation and survival. UV-B absorption by homodimeric UVR8 induces its monomerization and interaction with the E3 ubiquitin ligase COP1, leading ultimately to gene expression changes. UVR8 is inactivated through redimerization, facilitated by RUP1 and RUP2. Here, we describe a novel semi-dominant, hyperactive allele, namely uvr8-17D, that harbors a glycine-101 to serine mutation. UVR8G101S-overexpression led to weak constitutive photomorphogenesis and extreme UV-B responsiveness. UVR8G101S was observed to be predominantly monomeric in vivo and, once activated by UV-B, was not efficiently inactivated. Analysis of a UVR8G101S crystal structure revealed the distortion of a loop region normally involved in stabilization of the UVR8 homodimer. Plants expressing a UVR8 variant combining G101S with the previously described W285A mutation exhibited robust constitutive photomorphogenesis. This work provides further insight into UVR8 activation and inactivation mechanisms, and describes a genetic tool for the manipulation of photomorphogenic responses.

scholarly journals Mechanistic Links Underlying the Impact of C-Reactive Protein on Muscle Mass in Elderly

2017 ◽  
Vol 44 (1) ◽  
pp. 267-278 ◽  
Author(s):  
Britta Wåhlin-Larsson ◽  
Daniel J. Wilkinson ◽  
Emelie Strandberg ◽  
Adrian Hosford-Donovan ◽  
Philip J. Atherton ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Muscle Mass ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Inflammatory Marker ◽  
Immunofluorescent Staining ◽  
Age Related ◽  
Human Myotubes ◽  
The Impact ◽  
The Relationship

Background/Aims: Mechanisms underlying the relationship between systemic inflammation and age-related decline in muscle mass are poorly defined. The purpose of this work was to investigate the relationship between the systemic inflammatory marker CRP and muscle mass in elderly and to identify mechanisms by which CRP mediates its effects on skeletal muscle, in-vitro. Methods: Muscle mass and serum CRP level were determined in a cohort of 118 older women (67±1.7 years). Human muscle cells were differentiated into myotubes and were exposed to CRP. The size of myotubes was determined after immunofluorescent staining using troponin. Muscle protein synthesis was assessed using stable isotope tracers and key signalling pathways controlling protein synthesis were determined using western-blotting. Results: We observed an inverse relationship between circulating CRP level and muscle mass (β= -0.646 (95% CI: -0.888, -0.405) p<0.05) and demonstrated a reduction (p < 0.05) in the size of human myotubes exposed to CRP for 72 h. We next showed that this morphological change was accompanied by a CRP-mediated reduction (p < 0.05) in muscle protein fractional synthetic rate of human myotubes exposed to CRP for 24 h. We also identified a CRP-mediated increased phosphorylation (p<0.05) of regulators of cellular energy stress including AMPK and downstream targets, raptor and ACC-β, together with decreased phosphorylation of Akt and rpS6, which are important factors controlling protein synthesis. Conclusion: This work established for the first time mechanistic links by which chronic elevation of CRP can contribute to age-related decline in muscle function.

scholarly journals Aerobic Exercise Training and In Vivo Akt Activation Counteract Cancer Cachexia by Inducing a Hypertrophic Profile through eIF-2α Modulation

Cancers ◽  
2021 ◽  
Vol 14 (1) ◽  
pp. 28
Author(s):  
Marcelo G. Pereira ◽  
Vanessa A. Voltarelli ◽  
Gabriel C. Tobias ◽  
Lara de Souza ◽  
Gabriela S. Borges ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Exercise Training ◽  
Aerobic Exercise ◽  
Muscle Mass ◽  
Skeletal Muscle Mass ◽  
Cancer Cachexia ◽  
Aerobic Exercise Training ◽  
Akt Activation ◽  
The Impact

Cancer cachexia is a multifactorial and devastating syndrome characterized by severe skeletal muscle mass loss and dysfunction. As cachexia still has neither a cure nor an effective treatment, better understanding of skeletal muscle plasticity in the context of cancer is of great importance. Although aerobic exercise training (AET) has been shown as an important complementary therapy for chronic diseases and associated comorbidities, the impact of AET on skeletal muscle mass maintenance during cancer progression has not been well documented yet. Here, we show that previous AET induced a protective mechanism against tumor-induced muscle wasting by modulating the Akt/mTORC1 signaling and eukaryotic initiation factors, specifically eIF2-α. Thereafter, it was determined whether the in vivo Akt activation would induce a hypertrophic profile in cachectic muscles. As observed for the first time, Akt-induced hypertrophy was able and sufficient to either prevent or revert cancer cachexia by modulating both Akt/mTORC1 pathway and the eIF-2α activation, and induced a better muscle functionality. These findings provide evidence that skeletal muscle tissue still preserves hypertrophic potential to be stimulated by either AET or gene therapy to counteract cancer cachexia.

scholarly journals Gene expression of calpains and their specific endogenous inhibitor, calpastatin, in skeletal muscle of fed and fasted rabbits

1992 ◽  
Vol 287 (1) ◽  
pp. 163-171 ◽  
Author(s):  
M A Ilian ◽  
N E Forsberg
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Protein Degradation ◽  
Muscle Protein ◽  
Fold Increase ◽  
Myofibrillar Protein ◽  
Mrna Levels ◽  
Genes Encoding ◽  
Calpain Ii

To investigate the role of calpains in myofibrillar protein degradation in skeletal muscle and the regulation of their activity in vivo, we studied the effects of fasting on gene expression of calpains and calpastatin in the skeletal muscle of rabbits. In response to fasting, myofibrillar protein degradation increased 2-fold and mRNA levels of calpain I, calpain II and calpastatin were also increased. However, calpain and calpastatin activities remained unchanged. To investigate this discrepancy, we analysed polysomal calpain mRNA. Results indicated that fasting caused a 2-fold increase in the loading of calpain I and II mRNAs on ribosomes. Thus transcription of genes encoding calpain may be increased during fasting to ensure adequate synthesis of the proteinases needed to mobilize muscle protein reserves. The effect of fasting on calpain and calpastatin mRNA expression is shared by cathepsin D and proteasome C2 but not by beta-actin, implying that fasting invokes control of several proteolytic systems in skeletal muscle and underscores the possibility that each proteolytic system plays a role in the adaptation of skeletal muscle to the fasted state.

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