Skeletal muscle–specific knockout of DEP domain containing 5 protein increases mTORC1 signaling, muscle cell hypertrophy, and mitochondrial respiration

Theodore Graber, Christopher Fry, Camille R. Brightwell, Tatiana Moro, Rosario Maroto, Nisha Bhattarai, Craig Porter, Maki Wakamiya, Blake Rasmussen

Research output: Contribution to journalArticle

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Abstract

mTORC1 regulates protein synthesis and in turn is regulated by growth factors, energy status, and amino acid availability. In kidney cell (HEK293-T) culture, the GAP activity toward RAG (GATOR1) protein complex suppresses activation of the RAG A/B–RAG C/D heterodimer when amino acids are insufficient. During amino acid sufficiency, the RAG heterodimer recruits mTORC1 to the lysosomal membrane where its interaction with Ras homolog enriched in brain (Rheb) stimulates mTORC1’s kinase activity. The DEP domain containing 5 (DEPDC5) protein, a GATOR1 subunit, causes familial focal epilepsy when mutated, and global knockout of the Depdc5 gene is embryonically lethal. To study the function of DEPDC5 in skeletal muscle, we generated a muscle-specific inducible Depdc5 knockout mouse, hypothesizing that knocking out Depdc5 in muscle would make mTORC1 constitutively active, causing hypertrophy and improving muscle function. Examining mTORC1 signaling, morphology, mitochondrial respiratory capacity, contractile function, and applied physical function (e.g. rotarod, treadmill, grip test, and wheel running), we observed that mTORC1 activity was significantly higher in knockout (KO) mice, indicated by the increased phosphorylation of mTOR and its downstream effectors (by 118% for p-mTOR/mTOR, 114% for p-S6K1/S6K1, and 35% for p-4E-BP1/4E-BP1). The KO animals also exhibited soleus muscle cell hypertrophy and a 2.5-fold increase in mitochondrial respiratory capacity. However, contrary to our hypothesis, neither physical nor contractile function improved. In conclusion, DEPDC5 depletion in adult skeletal muscle removes GATOR1 inhibition of mTORC1, resulting in muscle hypertrophy and increased mitochondrial respiration, but does not improve overall muscle quality and function.

Original languageEnglish (US)
Pages (from-to)4091-4102
Number of pages12
JournalJournal of Biological Chemistry
Volume294
Issue number11
DOIs
StatePublished - Jan 1 2019

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Muscle Cells
Hypertrophy
Muscle
Respiration
Cells
Muscles
Proteins
Skeletal Muscle
Amino Acids
Knockout Mice
Gene Knockout Techniques
Partial Epilepsy
HEK293 Cells
Exercise equipment
Hand Strength
Phosphorylation
1-(2-(dodecyloxy)ethyl)pyrrolidine hydrochloride
mechanistic target of rapamycin complex 1
Exercise Test
Running

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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Skeletal muscle–specific knockout of DEP domain containing 5 protein increases mTORC1 signaling, muscle cell hypertrophy, and mitochondrial respiration. / Graber, Theodore; Fry, Christopher; Brightwell, Camille R.; Moro, Tatiana; Maroto, Rosario; Bhattarai, Nisha; Porter, Craig; Wakamiya, Maki; Rasmussen, Blake.

In: Journal of Biological Chemistry, Vol. 294, No. 11, 01.01.2019, p. 4091-4102.

Research output: Contribution to journalArticle

Graber, Theodore ; Fry, Christopher ; Brightwell, Camille R. ; Moro, Tatiana ; Maroto, Rosario ; Bhattarai, Nisha ; Porter, Craig ; Wakamiya, Maki ; Rasmussen, Blake. / Skeletal muscle–specific knockout of DEP domain containing 5 protein increases mTORC1 signaling, muscle cell hypertrophy, and mitochondrial respiration. In: Journal of Biological Chemistry. 2019 ; Vol. 294, No. 11. pp. 4091-4102.
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abstract = "mTORC1 regulates protein synthesis and in turn is regulated by growth factors, energy status, and amino acid availability. In kidney cell (HEK293-T) culture, the GAP activity toward RAG (GATOR1) protein complex suppresses activation of the RAG A/B–RAG C/D heterodimer when amino acids are insufficient. During amino acid sufficiency, the RAG heterodimer recruits mTORC1 to the lysosomal membrane where its interaction with Ras homolog enriched in brain (Rheb) stimulates mTORC1’s kinase activity. The DEP domain containing 5 (DEPDC5) protein, a GATOR1 subunit, causes familial focal epilepsy when mutated, and global knockout of the Depdc5 gene is embryonically lethal. To study the function of DEPDC5 in skeletal muscle, we generated a muscle-specific inducible Depdc5 knockout mouse, hypothesizing that knocking out Depdc5 in muscle would make mTORC1 constitutively active, causing hypertrophy and improving muscle function. Examining mTORC1 signaling, morphology, mitochondrial respiratory capacity, contractile function, and applied physical function (e.g. rotarod, treadmill, grip test, and wheel running), we observed that mTORC1 activity was significantly higher in knockout (KO) mice, indicated by the increased phosphorylation of mTOR and its downstream effectors (by 118{\%} for p-mTOR/mTOR, 114{\%} for p-S6K1/S6K1, and 35{\%} for p-4E-BP1/4E-BP1). The KO animals also exhibited soleus muscle cell hypertrophy and a 2.5-fold increase in mitochondrial respiratory capacity. However, contrary to our hypothesis, neither physical nor contractile function improved. In conclusion, DEPDC5 depletion in adult skeletal muscle removes GATOR1 inhibition of mTORC1, resulting in muscle hypertrophy and increased mitochondrial respiration, but does not improve overall muscle quality and function.",
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AU - Fry, Christopher

AU - Brightwell, Camille R.

AU - Moro, Tatiana

AU - Maroto, Rosario

AU - Bhattarai, Nisha

AU - Porter, Craig

AU - Wakamiya, Maki

AU - Rasmussen, Blake

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