Molecular basis for N-terminal acetylation by human NatE and its modulation by HYPK.

The human N-terminal acetyltransferase E (NatE) contains NAA10 and NAA50 catalytic, and NAA15 auxiliary subunits and associates with HYPK, a protein with intrinsic NAA10 inhibitory activity. NatE co-translationally acetylates the N-terminus of half the proteome to mediate diverse biological processes, including protein half-life, localization, and interaction. The molecular basis for how NatE and HYPK cooperate is unknown. Here, we report the cryo-EM structures of human NatE and NatE/HYPK complexes and associated biochemistry. We reveal that NAA50 and HYPK exhibit negative cooperative binding to NAA15 in vitro and in human cells by inducing NAA15 shifts in opposing directions. NAA50 and HYPK each contribute to NAA10 activity inhibition through structural alteration of the NAA10 substrate-binding site. NAA50 activity is increased through NAA15 tethering, but is inhibited by HYPK through structural alteration of the NatE substrate-binding site. These studies reveal the molecular basis for coordinated N-terminal acetylation by NatE and HYPK.

Inhibition of the acetyltransferase NAT10 normalizes progeric and aging cells by rebalancing the Transportin-1 nuclear import pathway.

Hutchinson-Gilford progeria syndrome (HGPS) is an incurable premature aging disease. Identifying deregulated biological processes in HGPS might thus help define novel therapeutic strategies. Fibroblasts from HGPS patients display defects in nucleocytoplasmic shuttling of the GTP-bound form of the small GTPase Ran (RanGTP), which leads to abnormal transport of proteins into the nucleus. We report that microtubule stabilization in HGPS cells sequestered the nonclassical nuclear import protein Transportin-1 (TNPO1) in the cytoplasm, thus affecting the nuclear localization of its cargo, including the nuclear pore protein NUP153. Consequently, nuclear Ran, nuclear anchorage of the nucleoporin TPR, and chromatin organization were disrupted, deregulating gene expression and inducing senescence. Inhibiting N-acetyltransferase 10 (NAT10) ameliorated HGPS phenotypes by rebalancing the nuclear to cytoplasmic ratio of TNPO1. This restored nuclear pore complex integrity and nuclear Ran localization, thereby correcting HGPS cellular phenotypes. We observed a similar mechanism in cells from healthy aged individuals. This study identifies a nuclear import pathway affected in aging and underscores the potential for NAT10 inhibition as a possible therapeutic strategy for HGPS and perhaps also for pathologies associated with normal aging.

Chemical inhibition of NAT10 corrects defects of laminopathic cells.

Down-regulation and mutations of the nuclear-architecture proteins lamin A and C cause misshapen nuclei and altered chromatin organization associated with cancer and laminopathies, including the premature-aging disease Hutchinson-Gilford progeria syndrome (HGPS). Here, we identified the small molecule "Remodelin" that improved nuclear architecture, chromatin organization, and fitness of both human lamin A/C-depleted cells and HGPS-derived patient cells and decreased markers of DNA damage in these cells. Using a combination of chemical, cellular, and genetic approaches, we identified the acetyl-transferase protein NAT10 as the target of Remodelin that mediated nuclear shape rescue in laminopathic cells via microtubule reorganization. These findings provide insights into how NAT10 affects nuclear architecture and suggest alternative strategies for treating laminopathies and aging.