Abstract
The ability of laminins to self-polymerize is crucial for the formation of basement membranes. Development of this polymerized network has profound effects upon tissue architecture as well as on the intracellular organization and differentiation of neighboring cells. The laminin N-terminal (LN) domains have been shown to mediate this interaction and studies using proteolytic fragments derived from laminin-1 led to the theory that network assembly depends on the formation of a heterotrimeric complex between LN domains derived from alpha, beta, and gamma chains in different laminin molecules with homologous interactions being insignificant. The laminin family consists of 15 known isoforms formed from five alpha, three beta, and three gamma chains, of which some are truncated and lack the N-terminal LN domain. To address whether the model of heterotrimeric complex formation is applicable to laminin isoforms other than laminin-1, eight LN domains found in the laminin protein family were recombinantly expressed and tested in three different assays for homologous and heterologous interactions. The results showed that the lack of homologous interactions is an exception, with such interactions being seen for LN domains derived from all alpha chains and from the beta2 and beta3 subunits. The gamma chain-derived LN domains showed a far more limited binding repertoire, particularly in the case of the gamma3 chain, which is found present in a range of non-basement membrane locations. Further, whereas the interactions depended upon Ca2+ ions, with EDTA reversibly abrogating binding, EDTA-induced conformational changes were not reversible. Together these results demonstrate that the assembly model proposed on the basis of laminin-1 may be a simplification, with the assembly of naturally occurring laminin networks being far more complex and highly dependent upon which laminin isoforms are present.
Highlights
Basement membranes are specialized extracellular matrices found underlying all epithelia and endothelia as well as surrounding many types of mesenchymal cells
The results showed that the lack of homologous interactions is an exception, with such interactions being seen for laminin N-terminal (LN) domains derived from all ␣ chains and from the 2 and 3 subunits
The expression level was dependent upon the number of LE modules following the LN domain and varied between 2 and 15 mg/liter of conditioned medium, with the highest yields obtained from constructs containing four LE domains (Fig. 1)
Summary
Basement membranes are specialized extracellular matrices found underlying all epithelia and endothelia as well as surrounding many types of mesenchymal cells. It has a cross structure with one long and three short arms, the latter being formed from the three free N-terminal ends of the ␣, , and ␥ chains [8] These parts of the  and ␥ chains each contain two globular domains, designated IV and VI, whereas there are three, IVa, IVb, and VI, in the short arm contributed by the ␣ chain. The laminin trimer is formed by assembly of the coiled-coil ␣-helical regions of the three chains, which results in the formation of the long arm structure [12]. The self-aggregation of laminin-1 is shared by many “full-sized” laminin isoforms but not by laminin-5 and -6, which contain chains with N-terminal truncations and thereby lack one or more LN domains [18]
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