Visualization of membrane-stabilized SorCS2 cis interactions

Area of Science:

• Structural biology
• Neuroscience
• Biochemistry

Background:

• The Vps10p receptor family plays a crucial role in protein trafficking and cellular differentiation within the nervous system.
• Previous structural studies of SorCS2, a Vps10p family member, primarily examined isolated ectodomains, neglecting the impact of membrane association on its organization.

Purpose of the Study:

• To develop and utilize novel tools for reconstituting the SorCS2 ectodomain on proteoliposomes in its native orientation.
• To visualize and characterize the membrane-associated organization of SorCS2 using advanced imaging techniques.

Main Methods:

• Established two complementary methods for SorCS2 ectodomain reconstitution: non-covalent His-tag/nickel affinity coupling and covalent azide-alkyne cycloaddition.
• Employed electron cryo-tomography to visualize SorCS2 organization on proteoliposomes.
• Generated nanometer-resolution subtomogram averages of SorCS2 ectodomain dimers.

Main Results:

• Identified four previously unreported arrangements of SorCS2 dimers, including "head-to-side" and "head-to-head" interactions.
• Observed less frequent "side-by-side" and "face-to-face" interactions between SorCS2 domains.
• Demonstrated that SorCS2 organizes into distinct helical arrangements and small clusters on liposome surfaces.
• Highlighted the promiscuity of membrane-stabilized SorCS2 interactions.

Conclusions:

• Membrane association significantly influences the organization and interactions of SorCS2 receptors.
• The observed SorCS2 arrangements suggest a general mechanism where membrane association dictates receptor system organization.
• The developed tools offer a versatile platform for studying ectodomain-mediated receptor assemblies in a membrane context.