Journal Article Ppublish
PubMed: 40105919 DOI: 10.1091/mbc.E24-09-0419
Membrane composition and curvature in SNX9-mediated actin polymerization.
Mol Biol Cell 36(5):ar54
Authors: Pankti Vaishnav, Hanae Shimo Kondo, Jonathan R Gadsby, Thomas C A Blake, Ulrich Dobramysl, Julia Mason, Joseph Atherton, Jennifer L Gallop
Affiliations
- Gurdon Institute, University of Cambridge, Cambridge CB2 1QN, United Kingdom. Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, United Kingdom.
- Gurdon Institute, University of Cambridge, Cambridge CB2 1QN, United Kingdom.
- Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, United Kingdom.
- Randall Centre for Cell and Molecular Biophysics, King's College, London SE1 1YR, United Kingdom.
Abstract
Sorting nexin 9 (SNX9) is a membrane-binding scaffold protein that contributes to viral uptake and inflammation and is associated with worse outcomes in several cancers. It is involved in endocytosis of epidermal growth factor receptors, β1-integrin and membrane type 1 matrix metalloprotease, and formation of mitochondrial-derived vesicles. The SNX9 Bin-Amphiphysin-Rvs (BAR)-Phox homology (PX) domains bind phosphoinositide lipids and the Src homology 3 (SH3) domain interacts with dynamin and Neural-Wiskott Aldrich syndrome protein (N-WASP) to stimulate Arp2/3 complex-mediated actin polymerization. Here we use biolayer interferometry, cell-free reconstitution, and superresolution microscopy to analyze the specificity and activities of SNX9 at membranes. We find that more SNX9 can bind liposomes containing phosphatidylinositol (4,5)-bisphosphate (PI(4,5)P 2) and phosphatidylinositol (3)-phosphate (PI(3)P) compared with phosphatidylinositol (3,4)-bisphosphate (PI(3,4)P 2), despite similar affinities. Actin assembly requires the network of both PX-BAR and SH3 interactions. Three-dimensional direct stochastic optical reconstruction microscopy on filopodia-like reconstitutions shows that SNX9 and related protein transducer of Cdc42-dependent actin assembly-1 (TOCA-1) can form both flat and ∼0.5 µm curved assemblies at actin incorporation sites. Finally, using cryo-electron tomography, we show that SNX9 builds both branched and bundled actin networks demonstrating its potential for multifunctional roles in actin remodeling.
MeSH
Cell Physiological Phenomena, Actins, Molecular Biology, Phosphatidylinositols, Microscopy, Wiskott-Aldrich Syndrome, Membranes