Abstract
Cells require distinct adhesion complexes to form contacts with their neighbors or the extracellular matrix, and vinculin links these complexes to the actin cytoskeleton. Metavinculin, an isoform of vinculin that harbors a unique 68-residue insert in its tail domain, has distinct actin bundling and oligomerization properties and plays essential roles in muscle development and homeostasis. Moreover, patients with sporadic or familial mutations in the metavinculin-specific insert invariably develop fatal cardiomyopathies. Here we report the high resolution crystal structure of the metavinculin tail domain, as well as the crystal structures of full-length human native metavinculin (1,134 residues) and of the full-length cardiomyopathy-associated ΔLeu954 metavinculin deletion mutant. These structures reveal that an α-helix (H1′) and extended coil of the metavinculin insert replace α-helix H1 and its preceding extended coil found in the N-terminal region of the vinculin tail domain to form a new five-helix bundle tail domain. Further, biochemical analyses demonstrate that this helix replacement directs the distinct actin bundling and oligomerization properties of metavinculin. Finally, the cardiomyopathy associated ΔLeu954 and Arg975Trp metavinculin mutants reside on the replaced extended coil and the H1′ α-helix, respectively. Thus, a helix replacement mechanism directs metavinculin's unique functions.
Highlights
The morphology and functions of specialized cells within tissues such as muscle requires unique organization of the actin cytoskeleton, but how this is controlled is poorly understood
The H19 for H1 replacement in metavinculin results in a new fivehelix bundle that closely mimics the five-helix bundle found in vinculin tail (Vt), Figure 1
The helix replacement is directed by the unique intramolecular interactions of the H19 a-helix and its preceding extended coil of the metavinculin insert with the H2–H5 helical bundle present in the tail domain, which replaces the H1 a-helix and its preceding extended coil that are present in the five-helix bundle tail domain of vinculin
Summary
The morphology and functions of specialized cells within tissues such as muscle requires unique organization of the actin cytoskeleton, but how this is controlled is poorly understood At one level this relies on proper links of the actin network to cadherin receptor-mediated cell-cell adherens junctions, to integrin receptor-directed focal adhesions, and to intercalated discs that are required for muscle cell function and that orchestrate coordinated movement. The most severe mutant Arg975Trp is associated with both dilated (DCM) and hypertrophic (HCM) cardiomyopathies in man where it disrupts the organization of intercalated discs, results in a pI drop of about 1.4 pH units for residues 966–983, augments cross-linking of actin filaments [8], and may compromise the interactions of metavinculin with its partners, including vinculin [9]. The DLeu954 and Ala934Val mutations are associated with DCM and have more modest effects on the cross-linking of actin filaments, especially the Ala934Val mutation [7]
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