ROLE OF SIRT6 AND PARP-1 IN MUSCLE HOMEOSTASIS

Abstract

Sirtuins and PARPs are stress responsive family of proteins regulating genome instability, inflammation, aging-related diseases and energy metabolism. Both sirtuins and PARPs are NAD+ enzymes with deacetylase and ADP-ribose polymerase activity respectively. Sirtuin family is comprised of 7 members (SIRT1-SIRT7) with distributed intracellular localisation that catalyse target protein deacetylation in NAD+ dependent manner. Members of PARP family transfer ADP-ribose residue from donor NAD+ to target proteins. Since, both skeletal muscle and heart muscle are highly energy demanding and metabolically active organs, sirtuins and PARPs, which are known to affect cellular energy metabolism may regulate muscle homeostasis. We have studied the role of SIRT6 and PARP-1 in maintenance of skeletal and cardiac muscle homeostasis respectively. PART 1: Understanding the role of SIRT6 in stress-induced skeletal muscle degeneration We have explored the role of a SIRT6, a nuclear localised sirtuin in the maintenance of muscle mass at basal as well as under stress conditions. It has been shown previously that SIRT6 deficient mice show aging like phenotype by 2-3 weeks of age and die within 1 month after birth. SIRT6 deficient mice suffer from spontaneous muscle atrophy. To decipher the role of SIRT6 in maintenance of skeletal muscle homeostasis, we studied stress induced muscle atrophy in muscle-specific SIRT6 deficient mice (msSIRT6-KO) muscle and primary myotubes. Dexamethasone (Dex) administration increases SIRT6 level in mice muscle as well as in primary myotubes. SIRT6 depletion increases myotubes diameter and overexpression results in myotubes atrophy in vitro. msSIRT6 deficiency does not affect muscle mass at basal level, but it ameliorates skeletal muscle atrophy induced by Dex administration. SIRT6 deficiency results in reduced FoxOs levels, nuclear localisation, activity and occupancy to promoter of atrogenes. Mechanistically, SIRT6 deletion hyperactivates c-Jun transcription factor leading to enhanced PI3K/AKT/mTOR signalling. The results suggest that activation of FoxO activity or inhibition of IGF-AKT signalling using PI3K specific inhibitor or c-Jun activity by using c-Jun dominant negative adenovirus abrogated hypertrophic effect of SIRT6 depletion in primary myotubes. These findings suggest that muscle specific SIRT6 deletion protected mice muscle and primary myotubes against Dex-mediated muscle atrophy. Further, pharmacological inhibition of SIRT6 protects against glucocorticoid induced muscle atrophy in mice. Overall, these results provide novel insights into the regulation of glucocorticoid-induced skeletal muscle atrophy and suggest SIRT6 inhibition as a potential strategy for treating muscle wasting associated with stress and disease conditions. PART 2: Understanding the role of PARP1 in aging-associated heart failure in mice Poly(ADP-ribose) polymerases (PARPs) are a family of stress sensors known to play role in genomic instability, cell death, oxidative stress, inflammation and aging-related diseases. PARP-1 mediated PARylation accounts for majority of intracellular PARylation. PARP-1 is known to transcriptionally regulate genes involved in oxidative stress, inflammation and cell death. However, role of PARP-1 in aging-associated heart failure is not studied. Our study suggests that PARP-1 deficient mice show aging related cardiac hypertrophy as assessed by heart weight to tibia length ratio. PARP-1 deficiency results in cardiac systolic dysfunction in old mice with increased expression of fetal genes and fibrosis markers, suggesting cardiac remodelling in old PARP-1 deficient mice. PARP-1 deficiency results in hyperactivation of AKT signalling pathway in heart. We found that PARP-1 poly(ADP-ribosy)lates AKT, inhibits its phosphorylation and further activation. Our results suggest that E40 and E49 residues in the Pleckstrin homology (PH) domain of AKT are poly(ADP-ribosy)lated. Mechanistically, poly(ADP-ribosyl)ation prevents membrane recruitment of AKT, which without altering PIP3 levels. Interestingly, AKT specific chemical inhibitor abrogates cardiomyocyte hypertrophy in PARP-1 depleted cells. Our findings suggest that PARP-1 regulates AKT signalling and protects against aging-associated detrimental changes in heart.