Identification of Ubiquitin Ligases Required for Skeletal Muscle Atrophy

113

BackgroundMuscle ring finger 1 (MuRF1) is a muscle‐specific ubiquitin E3 ligase activated during clinical conditions associated with skeletal muscle wasting. Yet, there remains a paucity of therapeutic interventions that directly inhibit MuRF1 function, particularly in vivo. The current study, therefore, developed a novel compound targeting the central coiled coil domain of MuRF1 to inhibit muscle wasting in cardiac cachexia.MethodsWe identified small molecules that interfere with the MuRF1–titin interaction from a 130 000 compound screen based on Alpha Technology. A subset of nine prioritized compounds were synthesized and administrated during conditions of muscle wasting, that is, to C2C12 muscle cells treated with dexamethasone and to mice treated with monocrotaline to induce cardiac cachexia.ResultsThe nine selected compounds inhibited MuRF1–titin complexation with IC50 values <25 μM, of which three were found to also inhibit MuRF1 E3 ligase activity, with one further showing low toxicity on cultured myotubes. This last compound, EMBL chemical core ID#704946, also prevented atrophy in myotubes induced by dexamethasone and attenuated fibre atrophy and contractile dysfunction in mice during cardiac cachexia. Proteomic and western blot analyses showed that stress pathways were attenuated by ID#704946 treatment, including down‐regulation of MuRF1 and normalization of proteins associated with apoptosis (BAX) and protein synthesis (elF2B‐delta). Furthermore, actin ubiquitinylation and proteasome activity was attenuated.ConclusionsWe identified a novel compound directed to MuRF1's central myofibrillar protein recognition domain. This compound attenuated in vivo muscle wasting and contractile dysfunction in cardiac cachexia by protecting de novo protein synthesis and by down‐regulating apoptosis and ubiquitin‐proteasome‐dependent proteolysis.

Section: Introductionmentioning

confidence: 64%

Small‐molecule inhibition of MuRF1 attenuates skeletal muscle atrophy and dysfunction in cardiac cachexia

Bowen¹,

Adams²,

Werner

et al. 2017

113

“…Atrophy in myotubes was induced by expressing ubiquitin ligase muscle atrophy F‐box (MAFbx) in a rodent model 40. Consistent with this observation, mice deficient in either ubiquitin ligase MAFbx or Muscle RING Finger 1 (MuRF1) were resistant to atrophy 40. Besides degradation of proteins via the ubiquitin proteasome degradation pathway, the autophagy lysosomal degradation pathway is similarly important in skeletal muscles 41.…”

Section: Discussionmentioning

confidence: 83%

“…In muscle atrophy, the balance between synthesis of new proteins and degradation of existing proteins is tipped towards the latter 39. Atrophy in myotubes was induced by expressing ubiquitin ligase muscle atrophy F‐box (MAFbx) in a rodent model 40. Consistent with this observation, mice deficient in either ubiquitin ligase MAFbx or Muscle RING Finger 1 (MuRF1) were resistant to atrophy 40.…”

Section: Discussionmentioning

confidence: 91%

Comprehensive proteome analysis of human skeletal muscle in cachexia and sarcopenia: a pilot study

Ebhardt

Degen

Tadini

et al. 2017

BackgroundCancer cachexia (cancer‐induced muscle wasting) is found in a subgroup of cancer patients leaving the patients with a poor prognosis for survival due to a lower tolerance of the chemotherapeutic drug. The cause of the muscle wasting in these patients is not fully understood, and no predictive biomarker exists to identify these patients early on. Skeletal muscle loss is an inevitable consequence of advancing age. As cancer frequently occurs in old age, identifying and differentiating the molecular mechanisms mediating muscle wasting in cancer cachexia vs. age‐related sarcopenia are a challenge. However, the ability to distinguish between them is critical for early intervention, and simple measures of body weight may not be sufficiently sensitive to detect cachexia early.MethodsWe used a range of omics approaches: (i) undepleted proteome was quantified using advanced high mass accuracy mass spectrometers in SWATH‐MS acquisition mode; (ii) phospho epitopes were quantified using protein arrays; and (iii) morphology was assessed using fluorescent microscopy.ResultsWe quantified the soluble proteome of muscle biopsies from cancer cachexia patients and compared them with cohorts of cancer patients and healthy individuals with and without age‐related muscle loss (aka age‐related sarcopenia). Comparing the proteomes of these cohorts, we quantified changes in muscle contractile myosins and energy metabolism allowing for a clear identification of cachexia patients. In an in vitro time lapse experiment, we mimicked cancer cachexia and identified signal transduction pathways governing cell fusion to play a pivotal role in preventing muscle regeneration.ConclusionsThe work presented here lays the foundation for further understanding of muscle wasting diseases and holds the promise of overcoming ambiguous weight loss as a measure for defining cachexia to be replaced by a precise protein signature.

“…Akt prevents the nuclear translocation of the forkhead box factors 1 and 3a (FoxO1 and 3a) by phosphorylating multiple sites 20, 21. FoxOs are among the major transcription factors regulating the expression of the muscle‐specific ligases MAFbx and MuRF1, which are induced during several situations of muscle atrophy 20, 22. The phosphorylated form of FoxO1 and 3a were not modified by urolithin B ( Figure 2A and 2B).…”

Section: Resultsmentioning

confidence: 99%

Urolithin B, a newly identified regulator of skeletal muscle mass

Rodriguez

Pierre

Naslain

et al. 2017

BackgroundThe control of muscle size is an essential feature of health. Indeed, skeletal muscle atrophy leads to reduced strength, poor quality of life, and metabolic disturbances. Consequently, strategies aiming to attenuate muscle wasting and to promote muscle growth during various (pathological) physiological states like sarcopenia, immobilization, malnutrition, or cachexia are needed to address this extensive health issue. In this study, we tested the effects of urolithin B, an ellagitannin‐derived metabolite, on skeletal muscle growth.MethodsC2C12 myotubes were treated with 15 μM of urolithin B for 24 h. For in vivo experiments, mice were implanted with mini‐osmotic pumps delivering continuously 10 μg/day of urolithin B during 28 days. Muscle atrophy was studied in mice with a sciatic nerve denervation receiving urolithin B by the same way.ResultsOur experiments reveal that urolithin B enhances the growth and differentiation of C2C12 myotubes by increasing protein synthesis and repressing the ubiquitin–proteasome pathway. Genetic and pharmacological arguments support an implication of the androgen receptor. Signalling analyses suggest a crosstalk between the androgen receptor and the mTORC1 pathway, possibly via AMPK. In vivo experiments confirm that urolithin B induces muscle hypertrophy in mice and reduces muscle atrophy after the sciatic nerve section.ConclusionsThis study highlights the potential usefulness of urolithin B for the treatment of muscle mass loss associated with various (pathological) physiological states.