Organization of the laminin polymer node

Laminin polymerization is a key step of basement membrane assembly that depends on the binding of alpha, beta and gamma N-terminal LN domains to form a polymer node. Nodal assembly can be divided into two steps consisting of beta- and gamma-LN dimerization followed by calcium-dependent addition of the alpha-LN domain. The assembly and structural organization of laminin-111 LN-LEa segments was examined by size-exclusion chromatography (SEC) and electron microscopy. Triskelion-like structures were observed in negatively-stained images of purified alpha 1/beta 1/gamma 1 LN-LEa trimers. Image averaging of these revealed a heel-to-toe organization of the LN domains with angled outward projections of the LEa stem-like domains. A series of single-amino acid substitutions was introduced into the polymerization faces of the alpha 1, beta 1 and gamma 1 LN domains followed by SEC analysis to distinguish between loss of beta-gamma mediated dimerization and loss of alpha-dependent trimerization (with intact beta-gamma dimers). Dimer-blocking mutations were confined to the gamma 1-toe and the beta 1-heel, whereas the trimer-only-blocking mutations mapped to the gamma 1-heel, beta 1-toe and the alpha 1-toe and heel. Thus, in the polymer node the gamma 1-toe pairs with the beta 1-heel, the beta 1-toe pairs with the alpha 1-heel, and the alpha 1-toe pairs with the gamma 1-heel. (C) 2021 The Authors. Published by Elsevier B.V.

Automated summary

Laminin polymerization is a crucial step in basement membrane assembly, involving the binding of alpha, beta, and gamma N-terminal LN domains to form a polymer node. The study examined the assembly and organization of laminin-111 LN-LEa segments, revealing triskelion-like structures and the pairing of LN and LEa domains. Single amino acid substitutions were introduced to analyze the impact of mutations on beta-gamma dimerization and alpha-dependent trimerization, unveiling specific regions critical for each process.