Amyloid-β induced membrane damage instigates tunneling nanotubes by exploiting p21-activated kinase dependent actin remodulation

Abstract Alzheimer’s disease (AD) pathology progresses gradually via anatomically connected brain regions. Earlier studies have shown that amyloid-β1-42 oligomers (oAβ) can be directly transferred between connected neurons. However, the mechanism of transfer is not fully revealed. We observed formation of oAβ induced tunneling nanotubes (TNTs), nanoscaled f-actin containing membrane conduit, in differentially differentiated SH-SY5Y neuronal models. Time-lapse images showed that TNTs propagate oligomers from one cell to another. Preceding the TNT-formation, we detected oAβ induced plasma membrane (PM) damage and calcium-dependent repair through lysosomal-exocytosis and significant membrane surface expansion, followed by massive endocytosis to re-establish the PM. Massive endocytosis was monitored by an influx of the membrane-impermeable dye TMA-DPH and PM damage was quantified by propidium iodide influx in the absence of calcium. The massive endocytosis eventually caused accumulation of internalized oAβ in Lamp1 positive multi vesicular bodies/lysosomes via the actin cytoskeleton remodulating p21-activated kinase1 (PAK1) dependent endocytic pathway. Three dimensional quantitative and qualitative confocal imaging, structured illumination superresolution microscopy (SIM) and flowcytometry data revealed that oAβ induces activated phospho-PAK1, which modulates the formation of long stretched f-actin extensions between cells. Moreover, formation of TNTs can be inhibited by preventing PAK1 dependent internalization of oAβ using small-molecule inhibitor IPA-3, a highly selective cell permeable auto-regulatory inhibitor of PAK1. The present study gives insight that the TNTs are probably instigated as a consequence of oAβ induced PM damage and repair process, followed by PAK1 dependent endocytosis and actin remodeling, probably to maintain cell surface expansion and/or membrane tension in equilibrium.