Alzheimer mutations stabilize synaptotoxic γ-secretase-substrate complexes

Abstract

AbstractAlzheimer’s disease is characterized pathologically by cerebral deposition of 42-residue amyloid β-peptide (Aβ42), proteolytically produced from amyloid precursor protein (APP) by β- and γ-secretases.1Although mutations in APP and presenilin, the catalytic component of γ-secretase, cause familial Alzheimer’s disease (FAD), a role for Aβ42 as the primary disease driver has not been clearly established and remains controversial.2,3Here we show through comprehensive analysis of the multi-step proteolysis of APP substrate C99 by γ-secretase that FAD mutations are consistently deficient in early proteolytic events, not later events that produce secreted Aβ peptides. Cryo-electron microscopy revealed that a substrate mimetic traps γ-secretase at the transition state for intramembrane proteolysis, and this structure closely aligns with activated enzyme-substrate complex captured by molecular dynamics simulations.In silicosimulations and fluorescence lifetime imaging microscopy in cultured cells support stabilization by FAD mutations of enzyme-substrate and/or enzyme-intermediate complexes. Neuronal expression of C99 and/or presenilin-1 inCaenorabditis elegansled to age-dependent synaptic loss only when one of the transgenes carried an FAD mutation. Designed mutations that stabilize the enzyme-substrate complex and block proteolysis likewise led to synaptic loss. Collectively, these findings implicate the stalled process—not the released products—of γ-secretase cleavage of substrates in FAD pathogenesis.