Super-resolution microscopy reveals majorly mono- and dimeric presenilin1/γ-secretase at the cell surface
Abril Escamilla‐Ayala, Ragna Sannerud, Magali Mondin, Karin Poersch, Wendy Vermeire, Laura Paparelli, Caroline Berlage, Marcelle Koenig, Lucía Chávez‐Gutiérrez, Maximilian H. Ulbrich, Sebastian Munck, Hideaki Mizuno, Wim Annaert
Abstract
γ-Secretase is a multi-subunit enzyme whose aberrant activity is associated with Alzheimer's disease and cancer. While its structure is atomically resolved, γ-secretase localization in the membrane in situ relies mostly on biochemical data. Here, we combined fluorescent tagging of γ-secretase subunits with super-resolution microscopy in fibroblasts. Structured illumination microscopy revealed single γ-secretase complexes with a monodisperse distribution and in a 1:1 stoichiometry of PSEN1 and nicastrin subunits. In living cells, sptPALM revealed PSEN1/γ-secretase mainly with directed motility and frequenting 'hotspots' or high track-density areas that are sensitive to γ-secretase inhibitors. We visualized γ-secretase association with substrates like amyloid precursor protein and N-cadherin, but not with its sheddases ADAM10 or BACE1 at the cell surface, arguing against pre-formed megadalton complexes. Nonetheless, in living cells PSEN1/γ-secretase transiently visits ADAM10 hotspots. Our results highlight the power of super-resolution microscopy for the study of γ-secretase distribution and dynamics in the membrane.