Induction and Myofibrillar Targeting of CARP, and Suppression of the Nkx2.5 Pathway in the MDM Mouse with Impaired Titin-based Signaling

Witt, Christian; Ono, Yasuko; Puschmann, Eva; McNabb, Mark; Wu, Yingliang; Gotthardt, Michael; Witt, Stephanie H.; Haak, Markus; Labeit, D.; Gregorio, Carol C.; Sorimachi, Hiroyuki; Granzier, Henk; Labeit, Siegfried

doi:10.1016/j.jmb.2003.12.021

Cited by 84 publications

(101 citation statements)

References 40 publications

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“…However, it still remains unclear whether the main effects of the compound in vivo are exerted by the inhibition of MuRF1's enzyme activity or rather its down‐regulation in protein expression, although our in vitro and in vivo data suggest that a combination of both is likely playing a role. MuRF1 is also known to interact with numerous substrates, with one in particular being CARP (a member of the muscle ankyrin repeat protein family20, 37), which is a purported nuclear‐based and sarcomere (titin)‐based protein with transcriptional functions 20, 37. CARP is known to be up‐regulated in stress‐related conditions and is associated with contractile dysfunction and muscle atrophy 20, 38, 39.…”

Section: Discussionmentioning

confidence: 99%

“…MuRF1 is also known to interact with numerous substrates, with one in particular being CARP (a member of the muscle ankyrin repeat protein family20, 37), which is a purported nuclear‐based and sarcomere (titin)‐based protein with transcriptional functions 20, 37. CARP is known to be up‐regulated in stress‐related conditions and is associated with contractile dysfunction and muscle atrophy 20, 38, 39. In line with such evidence, we also observed an increase in CARP expression in MCT‐stressed mice, but this effect was abolished in the ID#704946‐fed mice.…”

Section: Discussionmentioning

confidence: 99%

“…Our earlier studies assigned MuRF1's interaction with muscle proteins to a central coiled coil domain (‘MuRF1 central’) 19, 20. Therefore, we hypothesized that it might be beneficial to inhibit the recognition of muscle proteins by this MuRF1 central domain, thereby maintaining its basal expression level and E3 ligase activity that resides in MuRF1's N‐terminal ring finger domain.…”

Section: Introductionmentioning

confidence: 99%

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

Bowen¹,

Adams²,

Werner

et al. 2017

J cachexia sarcopenia muscle

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119

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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.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

Bowen¹,

Adams²,

Werner

et al. 2017

J cachexia sarcopenia muscle

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119

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“…2A) (31,32). Thus, expression of these regions was also investigated in cultured skeletal muscle cells.…”

Section: Transcription Of the P94 Gene During Myofibrillogenesis-ex-mentioning

confidence: 99%

“…Our previously proposed "signal complex" hypothesizes that connectin-binding proteins in Z, N2A, or M function as a complex at each position to maintain cooperativity among the many different levels of muscle components, e.g. between the sarcomere and the nucleus (32), where the extension/contraction states in N2A would be communicated to other levels by the action of p94.…”

Section: Regulated Expression Of P94⌬ex During Muscle Development Andmentioning

confidence: 99%

Myogenic Stage, Sarcomere Length, and Protease Activity Modulate Localization of Muscle-specific Calpain

Ojima

Ono

Doi

et al. 2007

Journal of Biological Chemistry

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p94/calpain 3 is a Ca2؉ -binding intracellular protease predominantly expressed in skeletal muscles. p94 binds to the N2A and M-line regions of connectin/titin and localizes in the Z-bands. Genetic evidence showing that compromised p94 proteolytic activity leads to muscular dystrophy (limb-girdle muscular dystrophy type 2A) indicates the importance of p94 function in myofibrils. Here we show that a series of p94 splice variants is expressed immediately after muscle differentiation and differentially change localization during myofibrillogenesis. We found that the endogenous N-terminal (but not C-terminal) domain of p94 was not only localized in the Z-bands but also directly bound to sarcomeric ␣-actinin. These data suggest the incorporation of proteolytic N-terminal fragments of p94 into the Z-bands. In myofibrils localization of exogenously expressed p94 shifted from the M-line to N2A as the sarcomere lengthens beyond ϳ2.6 and 2.8 m for wild-type and proteaseinactive p94, respectively. These data demonstrate for the first time that p94 proteolytic activity is involved in responses to muscle conditions, which may explain why p94 inactivation causes limb-girdle muscular dystrophy. p94/calpain 3 is a member of the calpain family of intracellular Ca 2ϩ -requiring Cys proteases, which cleave substrates at specific and limited sites to modulate their structure and function. In contrast to the conventional -and m-calpains, which are ubiquitously expressed in almost all cells (1, 2), p94 is predominantly expressed in skeletal muscle with lesser amounts of several differentially spliced variants in skeletal muscle and nonmuscle cells (3). The human p94 gene, CAPN3, was identified as responsible for limb-girdle muscular dystrophy type 2A (or "calpainopathy") (4). In mice transgenic overexpression of a protease-inactive form of p94, p94:C129S, and p94 gene knockout caused a mild myopathy and muscular dystrophy, respectively (5-7). Furthermore, a defect in the proper proteolytic activity of p94 is a common feature of limb-girdle muscular dystrophy type 2A pathogenic mutations (8). These findings indicate that p94 protease activity is essential to maintain healthy condition of skeletal muscle. However, the specific physiological functions of p94 as a protease in skeletal muscle remain largely unknown.Skeletal muscles contain highly organized sarcomere structures that consist of systematically expressed myofibrillar proteins. One of the earliest myofibrillar proteins expressed in vertebrate myogenic cells is connectin/titin (9), which is the largest protein molecule known and is abundantly expressed in striated muscles where it constitutes an intrasarcomeric filament (10, 11). During sarcomere assembly, connectin is considered a key molecule for integration of thin filament/Z-band precursors (I-Z-I bodies) and the thick filaments, which are independently assembled in growth tips of elongating myotubes. The N terminus of connectin is located in both I-Z-I body and mature Z, and the C terminus is in the M-line of the thick fil...

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CARP interacts with titin at a unique helical N2A sequence and at the domain Ig81 to form a structured complex

et al. 2016

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The cardiac ankyrin repeat protein (CARP) is up-regulated in the myocardium during cardiovascular disease and in response to mechanical or toxic stress. Stress-induced CARP interacts with the N2A spring region of the titin filament to modulate muscle compliance. We characterize the interaction between CARP and titin-N2A and show that the binding site in titin spans the dual domain UN2A-Ig81. We find that the unique sequence UN2A is not structurally disordered, but that it has a stable, elongated a-helical fold that possibly acts as a constant force spring. Our findings portray CARP/titin-N2A as a structured node and help to rationalize the molecular basis of CARP mechanosensing in the sarcomeric I-band.

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Induction and Myofibrillar Targeting of CARP, and Suppression of the Nkx2.5 Pathway in the MDM Mouse with Impaired Titin-based Signaling

Cited by 84 publications

References 40 publications

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

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

Myogenic Stage, Sarcomere Length, and Protease Activity Modulate Localization of Muscle-specific Calpain

CARP interacts with titin at a unique helical N2A sequence and at the domain Ig81 to form a structured complex

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