Developmental regulation of MURF E3 ubiquitin ligases in skeletal muscle

The striated muscle-specific tripartite motif (TRIM) proteins TRIM63/MURF1, TRIM55/MURF2 and TRIM54/MURF3 can function as E3 ubiquitin ligases in ubiquitin-mediated muscle protein turnover. Despite the well-characterised role of MURF1 in skeletal muscle atrophy, the dynamics of MURF isogene expression in the development and early postnatal adaptation of skeletal muscle is unknown. Here, we show that MURF2 is the isogene most highly expressed in embryonic skeletal muscle at E15.5, with the 50 kDa A isoform predominantly expressed. MURF1 and MURF3 are upregulated only postnatally. Knockdown of MURF2 p50A by isoform-specific siRNA results in delayed myogenic differentiation and myotube formation in vitro, with perturbation of the stable, glutamylated microtubule population. This underscores that MURF2 plays an important role in the earliest stages of skeletal muscle differentiation and myofibrillogenesis. During further development, there is a shift towards the 60 kDa A isoform, which dominates postnatally. Analysis of the fibre-type expression shows that MURF2 A isoforms are predominantly slow-fibre associated, whilst MURF1 is largely excluded from these fibres, and MURF3 is ubiquitously distributed in both type I and II fibres

Expression of MuRF1 or MuRF2 is essential for the induction of skeletal muscle atrophy and dysfunction in a murine pulmonary hypertension model

Background Pulmonary hypertension leads to right ventricular heart failure and ultimately to cardiac cachexia. Cardiac cachexia induces skeletal muscles atrophy and contractile dysfunction. MAFbx and MuRF1 are two key proteins that have been implicated in chronic muscle atrophy of several wasting states. Methods Monocrotaline (MCT) was injected over eight weeks into mice to establish pulmonary hypertension as a murine model for cardiac cachexia. The effects on skeletal muscle atrophy, myofiber force, and selected muscle proteins were evaluated in wild-type (WT), MuRF1, and MuRF2-KO mice by determining muscle weights, in vitro muscle force and enzyme activities in soleus and tibialis anterior (TA) muscle. Results In WT, MCT treatment induced wasting of soleus and TA mass, loss of myofiber force, and depletion of citrate synthase (CS), creatine kinase (CK), and malate dehydrogenase (MDH) (all key metabolic enzymes). This suggests that the murine MCT model is useful to mimic peripheral myopathies as found in human cardiac cachexia. In MuRF1 and MuRF2-KO mice, soleus and TA muscles were protected from atrophy, contractile dysfunction, while metabolic enzymes were not lowered in MuRF1 or MuRF2-KO mice. Furthermore, MuRF2 expression was lower in MuRF1KO mice when compared to C57BL/6 mice. Conclusions In addition to MuRF1, inactivation of MuRF2 also provides a potent protection from peripheral myopathy in cardiac cachexia. The protection of metabolic enzymes in both MuRF1KO and MuRF2KO mice as well as the dependence of MuRF2 expression on MuRF1 suggests intimate relationships between MuRF1 and MuRF2 during muscle atrophy signaling

Microgenomic Analysis in Skeletal Muscle: Expression Signatures of Individual Fast and Slow Myofibers

BACKGROUND: Skeletal muscle is a complex, versatile tissue composed of a variety of functionally diverse fiber types. Although the biochemical, structural and functional properties of myofibers have been the subject of intense investigation for the last decades, understanding molecular processes regulating fiber type diversity is still complicated by the heterogeneity of cell types present in the whole muscle organ. METHODOLOGY/PRINCIPAL FINDINGS: We have produced a first catalogue of genes expressed in mouse slow-oxidative (type 1) and fast-glycolytic (type 2B) fibers through transcriptome analysis at the single fiber level (microgenomics). Individual fibers were obtained from murine soleus and EDL muscles and initially classified by myosin heavy chain isoform content. Gene expression profiling on high density DNA oligonucleotide microarrays showed that both qualitative and quantitative improvements were achieved, compared to results with standard muscle homogenate. First, myofiber profiles were virtually free from non-muscle transcriptional activity. Second, thousands of muscle-specific genes were identified, leading to a better definition of gene signatures in the two fiber types as well as the detection of metabolic and signaling pathways that are differentially activated in specific fiber types. Several regulatory proteins showed preferential expression in slow myofibers. Discriminant analysis revealed novel genes that could be useful for fiber type functional classification. CONCLUSIONS/SIGNIFICANCE: As gene expression analyses at the single fiber level significantly increased the resolution power, this innovative approach would allow a better understanding of the adaptive transcriptomic transitions occurring in myofibers under physiological and pathological condition

Different configurations of specific thyroid hormone response elements mediate opposite effects of thyroid hormone and GC-1 on gene expression

T3 regulates transcription of the rat sarcoendoplasmic reticulum calcium ATPase in the heart. The T3 effect is mediated by three differently configured T3 response elements (TREs). Here we report the mutation of each individual TRE in the promoter and the contribution of each TRE on gene expression. Mutation of TRE1, a direct repeat element, exerted the strongest T3 response, compared with TRE2 and TRE3, which are inverted palindromes. The isolated TRE2 and TRE3, which showed no response (TRE2) or were weakly positive with T3 (TRE3), became strong negative regulatory elements with the T3 analog GC-1. We found that TRE1 recruits corepressor complexes containing nuclear receptor corepressor and histone deacetylase 3 in the absence of ligand, and steroid receptor coactivator-1-containing coactivator complexes with both T3 and GC-1. TRE3 bound the same corepressor complexes without ligand but showed only a weak association with steroid receptor coactivator-1 with T3 and a strong association with corepressor complexes with GC-1. Thus, GC-1 appears to control cofactor association differentially on these two sarcoendoplasmic reticulum calcium ATPase TREs, which could be the mechanism of ligand-dependent transcriptional activation and repression observed with the isolated TRE1 and TRE3 elements. Because the x-ray crystal structures of GC-1 and T3 complexed with the TR ligand binding domain are superimposable, the results imply that GC-1 and T3 induce differential effects on the receptor that are not evident in the static structures but must occur in the dynamic setting of receptor function. These results have implications for selective modulation of receptor function by agonist ligands

Exercise Training Prevents Oxidative Stress and Ubiquitin-Proteasome System Overactivity and Reverse Skeletal Muscle Atrophy in Heart Failure

Background: Heart failure (HF) is known to lead to skeletal muscle atrophy and dysfunction. However, intracellular mechanisms underlying HF-induced myopathy are not fully understood. We hypothesized that HF would increase oxidative stress and ubiquitin-proteasome system (UPS) activation in skeletal muscle of sympathetic hyperactivity mouse model. We also tested the hypothesis that aerobic exercise training (AET) would reestablish UPS activation in mice and human HF. Methods/Principal Findings: Time-course evaluation of plantaris muscle cross-sectional area, lipid hydroperoxidation, protein carbonylation and chymotrypsin-like proteasome activity was performed in a mouse model of sympathetic hyperactivity-induced HF. At the 7th month of age, HF mice displayed skeletal muscle atrophy, increased oxidative stress and UPS overactivation. Moderate-intensity AET restored lipid hydroperoxides and carbonylated protein levels paralleled by reduced E3 ligases mRNA levels, and reestablished chymotrypsin-like proteasome activity and plantaris trophicity. In human HF (patients randomized to sedentary or moderate-intensity AET protocol), skeletal muscle chymotrypsin-like proteasome activity was also increased and AET restored it to healthy control subjects’ levels. Conclusions: Collectively, our data provide evidence that AET effectively counteracts redox imbalance and UPS overactivation, preventing skeletal myopathy and exercise intolerance in sympathetic hyperactivity-induced HF in mice. Of particular interest, AET attenuates skeletal muscle proteasome activity paralleled by improved aerobic capacity in HF patients, which is not achieved by drug treatment itself. Altogether these findings strengthen the clinical relevance of AET in the treatment of HF

Histological and functional renal alterations caused by Bothrops alternatus snake venom: Expression and activity of Na+/K+-ATPase

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Background: Acute renal failure is a serious complication of human envenoming by Bothrops snakes. The ion pump Na+/K+-ATPase has an important role in renal tubule function, where it modulates sodium reabsorption and homeostasis of the extracellular compartment. Here, we investigated the morphological and functional renal alterations and changes in Na+/K+-ATPase expression and activity in rats injected with Bothrops alternatus snake venom. Methods: Male Wistar rats were injected with venom (0.8 mg/kg, iv.) and renal function was assessed 6.24, 48 and 72 h and 7 days post-venom. The rats were then killed and renal Na+/K+-ATPase activity was assayed based on phosphate release from ATP; gene and protein expressions were assessed by real time PCR and immunofluorescence microscopy, respectively. Results: Venom caused lobulation of the capillary tufts, dilation of Bowman's capsular space. F-actin disruption in Bowman's capsule and renal tubule brush border, and deposition of collagen around glomeruli and proximal tubules that persisted seven days after envenoming. Enhanced sodium and potassium excretion, reduced proximal sodium reabsorption, and proteinuria were observed 6 h post-venom, followed by a transient decrease in the glomerular filtration rate. Gene and protein expressions of the Na+/K+-ATPase alpha(1) subunit were increased 6 h post-venom, whereas Na+/K+-ATPase activity increased 6 h and 24 h post-venom. Conclusions: Bothrops alternatus venom caused marked morphological and functional renal alterations with enhanced Na+/K+-ATPase expression and activity in the early phase of renal damage. General significance: Enhanced Na+/K+-ATPase activity in the early hours after envenoming may attenuate the renal dysfunction associated with venom-induced damage. (C) 2011 Elsevier B.V. All rights reserved.18109895906Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Recent developments in the field of cachexia, sarcopenia, and muscle wasting: highlights from the 12th Cachexia Conference

Abstract This article highlights preclinical and clinical studies in the field of wasting disorders that were presented at the 12th Cachexia Conference held in Berlin, Germany, in December 2019. Herein, we summarize the biological and clinical significance of different strategies including antibodies that target Fn14, Spsb 1, SAA1 treatment, ZIP14, a MuRF1 inhibitor, and new diagnostic tools like T‐cell communication targets and cut‐offs for the detection of skeletal muscle wasting. Of particular interest were the transplantation of mesenchymal stromal cells and muscle stem cell communication. Importantly, one presentation discussed the effect of metal ion transporter ZIP14 loss that reduces cancer‐induced cachexia. The potential of anti‐ZIP14 antibodies and zinc chelation as anti‐cachexia therapy may require testing in patients with cancer cachexia. Large clinical studies were presented such as RePOWER (observational study of patients with primary mitochondrial myopathy), MMPOWER (treatment with elamipretide in patients with primary mitochondrial myopathy), and ACT‐ONE as well as new mouse models like the KPP mouse. Promising treatments include rapamycin analogue treatment, anamorelin, elanapril, glucocorticoids, SAA1, antibodies that target Fn14, and a MuRF1 inhibitor. Clinical studies investigated novel approaches, including the role of exercise. It remains a fact, however, that effective treatments for cachexia and wasting disorders are urgently needed in order to improve patients' quality of life and their survival