The AP2 adaptor enhances clathrin coat stiffness

• Published in: The FEBS journal Vol. 286; no. 20; pp. 4074 - 4085
• Main Authors: Lherbette, Michael, Redlingshöfer, Lisa, Brodsky, Frances M., Schaap, Iwan A. T., Dannhauser, Philip N.
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01.10.2019; John Wiley and Sons Inc
• Subjects: Adapters; Adaptor Protein Complex 2 - metabolism; Adaptor proteins; AFM; Animals; Atomic force microscopy; Bending; Brain - metabolism; Cell Membrane - metabolism; Chains; Clathrin; Clathrin - metabolism; Clathrin-Coated Vesicles - chemistry; Clathrin-Coated Vesicles - metabolism; Coated vesicles; Coating; Curvature; Endocytosis; Light chains; membrane biophysics; Membrane proteins; Membrane vesicles; Membranes; Original; Protein Binding; Proteins; Stiffness; Sus scrofa; clathrin; AFM; membrane biophysics
• ISSN: 1742-464X; 1742-4658
• DOI: 10.1111/febs.14961

Deformation of the plasma membrane into clathrin‐coated vesicles is a critical step in clathrin‐mediated endocytosis and requires the orchestrated assembly of clathrin and endocytic adaptors into a membrane‐associated protein coat. The individual role of these membrane‐bending and curvature‐stabilizing factors is subject to current debate. As such, it is unclear whether the clathrin coat itself is stiff enough to impose curvature and if so, whether this could be effectively transferred to the membrane by the linking adaptor proteins. We have recently demonstrated that clathrin alone is sufficient to form membrane buds in vitro. Here, we use atomic force microscopy to assess the contributions of clathrin and its membrane adaptor protein 2 (AP2) to clathrin coat stiffness, which determines the mechanics of vesicle formation. We found that clathrin coats are less than 10‐fold stiffer than the membrane they enclose, suggesting a delicate balance between the forces harnessed from clathrin coat formation and those required for membrane bending. We observed that clathrin adaptor protein AP2 increased the stiffness of coats formed from native clathrin, but did not affect less‐flexible coats formed from clathrin lacking the light chain subunits. We thus propose that clathrin light chains are important for clathrin coat flexibility and that AP2 facilitates efficient cargo sequestration during coated vesicle formation by modulating clathrin coat stiffness. Formation of clathrin‐coated vesicles facilitates endocytosis of membrane‐embedded cargo proteins. We used atomic force microscopy to establish how specific clathrin coat components contribute to membrane deformation during cargo uptake. We found that the cargo‐binding adaptor protein 2 (AP2) increases coat stiffness, generating a coat better able to deform cargo‐containing membrane. We further found that clathrin light chain subunits provide the clathrin coat with properties that enable AP2 to enhance coat stiffness.