Author:

Keywords:

Science & Technology, Life Sciences & Biomedicine, Biology, Life Sciences & Biomedicine - Other Topics, SINGLE-PARTICLE TRACKING, ACTIVE GAMMA-SECRETASE, AMYLOID PRECURSOR PROTEIN, ALZHEIMERS-DISEASE, PLASMA-MEMBRANE, LATERAL DIFFUSION, BETA-APP, PRESENILIN, NICASTRIN, COMPLEX, Alzheimer's disease, cell biology, gamma-secretase, human, mouse, presenilin1, single-particle tracking, super-resolution microscopy, Amyloid Precursor Protein Secretases, Animals, Cell Line, Cell Membrane, Fibroblasts, Humans, Membrane Proteins, Mice, Microscopy, Presenilin-1, 0601 Biochemistry and Cell Biology, 31 Biological sciences, 32 Biomedical and clinical sciences, 42 Health sciences

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.