Gregory Tombline

The accumulation of damage caused by oxidative stress has been linked to aging and to the etiology of numerous age-related diseases. The longevity gene, sirtuin 6 (SIRT6), promotes genome stability by facilitating DNA repair, especially under oxidative stress conditions. Here we uncover the mechanism by which SIRT6 is activated by oxidative stress to promote DNA double-strand break (DSB) repair. We show that the stress-activated protein kinase, c-Jun N-terminal kinase (JNK), phosphorylates SIRT6 on serine 10 in response to oxidative stress. This post-translational modification facilitates the mobilization of SIRT6 to DNA damage sites and is required for efficient recruitment of poly (ADP-ribose) polymerase 1 (PARP1) to DNA break sites and for efficient repair of DSBs. Our results demonstrate a post-translational mechanism regulating SIRT6, and they provide the link between oxidative stress signaling and DNA repair pathways that may be critical for hormetic response and longevity assurance.

Naked mole-rat (NMR), the longest-living rodent, produces very-high-molecular-mass hyaluronan (vHMM-HA), compared to other mammalian species. However, it is unclear if exceptional polymer length of vHMM-HA is important for longevity. Here, we show that vHMM-HA (>6.1 MDa) has superior cytoprotective properties compared to the shorter HMM-HA. It protects not only NMR cells, but also mouse and human cells from stress-induced cell-cycle arrest and cell death in a polymer length-dependent manner. The cytoprotective effect is dependent on the major HA-receptor, CD44. We find that vHMM-HA suppresses CD44 protein-protein interactions, whereas HMM-HA promotes them. As a result, vHMM-HA and HMM-HA induce opposing effects on the expression of CD44-dependent genes, which are associated with the p53 pathway. Concomitantly, vHMM-HA partially attenuates p53 and protects cells from stress in a p53-dependent manner. Our results implicate vHMM-HA in anti-aging mechanisms and suggest the potential applications of vHMM-HA for enhancing cellular stress resistance.

Kinetics of inhibition of ATPase activity of pure mouse Mdr3 P-glycoprotein upon incubation with MgADP and vanadate were studied along with the trapping of [14C]ADP in presence of vanadate. The presence of verapamil strongly magnified both effects. Inhibition of ATPase was also increased by several other drugs known to bind to drug-binding sites. Inhibition by ADP-vanadate was slow and depended cooperatively on nucleotide binding. Stoichiometry of [14C]ADP trapping by vanadate was 1 mol/mol P-glycoprotein at full inhibition. Catalytic site mutants prevented [14C]ADP trapping, whereas interdomain signal communication mutants reduced it in approximate correlation with their effects upon drug stimulation of ATPase. In explanation of the results, we propose that a "closed conformation" involving dimerization and interdigitation of the two nucleotide-binding domains is necessary to allow inhibition by ADP-vanadate. The results suggest that such a conformation occurs naturally during ATP hydrolysis. It is proposed that in order for the catalytic transition state to form, the two nucleotide-binding domains dimerize to form an integrated single entity containing two bound ATP with just one of the two ATP being hydrolyzed per dimerization event.