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Hypertrophic signalling

Hypertrophic signalling. Identify contraction-induced growth signals Describe the composition and regulation of mTORC1 Describe the effectors of mTOR Explain the role of mTOR in muscle hypertrophy Muscle contraction Diet Growth factors. Consequences of contraction.

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Hypertrophic signalling

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  1. Hypertrophic signalling • Identify contraction-induced growth signals • Describe the composition and regulation of mTORC1 • Describe the effectors of mTOR • Explain the role of mTOR in muscle hypertrophy • Muscle contraction • Diet • Growth factors

  2. Consequences of contraction • Intracellular calcium increase • ATP (energy) turnover • Muscle: Oxygen depletion, AMP accumulation • Systemic: nutrient mobilization • Membrane permeability • Growth factor release • Peptides: IGF-1, FGF, HGF • Lipids: PGF2a, PGE2 • Systemic hormones • Insulin, GH, adrenaline

  3. Exercise induces mTOR activity • Rats trained to lift 60%BW vest • Phosphorylation by WB • Protein synthesis over 16 h • Rapamycin blocks Akt phosphorylation mTOR phosphorylation Bolster & al., 2003 Kubica & al., 2005

  4. Rapamycin blocks hypertrophy • Synergist ablation • Cyclosporin to block Cn • Rapamycin to block mTOR • CsA muscles hypertrophy • Rap muscles don’t Bodine & al., 2001

  5. Why mTOR? • Powerful, multiplex regulator of protein synthesis and growth • Translation efficiency • Translational regulation/selection • Protein degradation • Activated by diverse growth and function relevant stimuli • Contraction/exercise • Nutrients • Hormones (insulin, IGF, HGH)

  6. Mammalian Target of Rapamycin Pro-growth stimuli mTORC mTOR Contraction p38 Protein synthesis (hypertrophy) Deldicque & al., 2005

  7. Two mTOR Complexes Rapamycin sensitive Rapamycin insensitive mTORC2 Composition mTOR GbL (mLST8) PRR5, mSin1 RICTOR Regulation Growth factors (PI3K/akt) mTORC1 (RICTOR) Targets Cytoskeleton (esp yeast) Proteasome (AktFOXO) Glycogen synthesis (GSK3) PKC • mTORC1 Composition • mTOR • GbL (mLST8) dispensible • PRAS40 • RAPTOR • Regulation • Growth factors (PI3K/akt) • Nutrients (TSC1/2, Rag) • Redox • Targets • Ribosomal biogenesis (p70S6k) • Translation (4EBP1) • Autophagy

  8. Core mTORC1 control • Active complex requires Rheb-GTP • Rheb GTPase • GTPase-Activating Protein (GAP) • Guanine Exchange Factor (GEF) • mTOR autophos S2481 • TSC 1/2 • Tuberous Sclerosis Complex • Major site of GF/energy reg. • GEF unknown/unnecessary • Translationally Controlled Tumor Protein TSC1 Rheb-GDP TSC2 TCTP(?) Rheb-GTP mTOR GbL Substrate RAPTOR

  9. Growth Factors and “Energy” • Phosphatidylinositol 3’ kinase (PI3K) • PIP2PIP3 • PDK1 • Akt • Extracellular-signal Regulated Kinase (ERK) • P38MK2 • AMPK (activates TSC2) • GSK3 (activates TSC2) • Hypoxia • HIFREDD ERK2 MK2 Akt GSK3 TSC1 TSC2 AMPK Rheb-GTP Rheb-GDP REDD

  10. Amino Acids • Branched-chain AA • Leucine, isoleucine, valine • Rag-GTPase • Ragulator AA-sensitive GEF • Translocation to Rheb-richlysosomes Rag-GDP TSC2 Ragulator RagB-GTP RAPTOR Rab7/ lysosome GbL mTOR Rheb-GTP AA-starved mTOR is distributed through the cytoplasm, and becomes localized to lysosomes rapidly on AA feeding Sanack & al., 2008

  11. Growth factors and overload • Insulin • Suppressed at low (<60% VO2max) intensity • Neutral at high (>80% VO2max) • Insulin-like growth factor-1 • Elevated after resistance exercise (up to 2 days) • Powerful growth stimulator • Insulin and IGF-1 Receptors • Insulin receptor substrate 1(IRS1) • PI3KAkt • ERK, p38, PLC IGF-1 expression after synergist ablation (Adams & al 2002)

  12. IGF-1 promotes muscle growth • Infused into muscle (notsystemic) • Activation of Akt, mTOR • p70S6k, 4EBP1 Adams & McCue 1998

  13. Overload seems independent of IGF-1 • Muscle hypertrophy by synergist ablation in IGF-1R knockout • Cardiac hypertrophy by swim-training in p70S6k knockout WT MKR-/- 35 d 7 d 0 d Plantaris mass after synergist ablation Spangenburg & al 2008 Heart weights after 8 weeks swimming (McMullen & al., 2004)

  14. Amino acid feeding • AA feeding alone increases mTOR &PS • Protein feeding with exercise gives much better/faster mTOR activation • No difference in hypertrophy (22 weeks) mTOR phosphorylation post-exercise with or without protein feeding (Hulmi & al 2009)

  15. Metabolic effects • Elevated AMP • AMP Kinase  TSC2 --| mTOR • Permissive? • GSK3 • InsulinAkt--|GSK3 • Oxygen • Hypoxia Inducible Factor REDDTSC2 • ROS directly oxidize cysteines AICAR-induced activation of AMPK blocks AA-induced protein synthesis (Pruznak & al., 2008)

  16. Intermediate summary • Exercise-related stresses tend to block mTOR during exercise and activate mTOR after exercise • Energetic stresses during exercise: Low O2, high AMP • Recovery processes/hormones after exercise • Nutrient mobilization • Insulin • IGF-1 • Acute mTOR signaling correlates with hypertrophy under normal conditions • Not in Insulin/IGF-1 receptor defective models • Not in p70 S6k defective models

  17. Correlation and causation 8000 6000 4000 Placebo Type II fiber area Protein 2000 0 5 10 15 20 Fold phosphorylation of p70S6k Muscle mass gain after 6 weeks HFES correlates with p70S6k phosphorylation at 6 hours. (Baar & Esser 1999) Fiber size after 3 weeks training vs p70S6k phosphorylation. (Hulmi & al 2009)

  18. mTOR effectors • Ribosome assembly • p70S6kRPS6 • 5’-TOP mRNAs (ribosome components) • Translational efficiency • 4EBP--|eIF4E • Cap dependent translation • Transcription factors • Akt/SGK--|FOXO • NFAT3, STAT3 • IRS-1 (negative feedback)

  19. Protein translation • Initiation • eIF4 recognition and melting of 7’mG cap • eIF4E cap-binding subunit • 4EBP competition with eIF4F scaffold • Recruit 40S ribosome • met-tRNA • eIF2 GTP-dependent met-tRNA loader • Recruit 60S ribosome • Start codon

  20. Initiation Transition to elongation Pre-initiation complex Fig 17-9

  21. Protein translation • Elongation • tRNA recruitment • eEF1 GTP-dependent tRNA carrier • GTP hydrolysis with peptide bond formation • Ribosome advance • eEF2 GTP-dependent procession • GTP hydrolysis with advance

  22. eEF1 Cycle Elongation Cycle Elongation eEF2 cycle Fig 17-10

  23. 3’ untranslated region structure • Post-transcriptional control • 2° and 3° structure of mRNA • Analogous to DNA promoter • 5’ Tract of Oligopyrimidines • Ribosomal proteins • eEF1, eEF2 • “Highly structured” 5’ cap • Ribosome scanning • Growth factors, cell cycle control • Internal Ribosome Entry Site (IRES) • Inflammation, angiogenesis Phosphorylated RPS6 favors these Active eIF4 complex favors these

  24. Species differences • Most proteins conserved yeast-human • Regulatory processes differ • S cerevisiae have 2 TORs • Drosophila akt doesn’t directly regulate TSC2 • C Elegans has no TSC1/2; transcriptional repression of RAPTOR via FOXO • S cerevisiae mTOR independent of Rheb

  25. Summary • High force contractions induce multiple signaling modes • Metabolites, growth factors, mechanical • Hypertrophy closely linked with mTOR • GF signaling • Metabolite signaling • mTOR is a powerful control of protein accretion • Makes more ribosomes via p70S6k • General translation efficiency via 4EBP • Reduced degradation via FOXO, NFAT3

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