Abstract
The molecular architecture and assembly mechanism of intermediate filaments have been enigmatic for decades. Among those, lamin filaments are of particular interest due to their universal role in cell nucleus and numerous disease-related mutations. Filament assembly is driven by specific interactions of the elementary dimers, which consist of the central coiled-coil rod domain flanked by non-helical head and tail domains. We aimed to investigate the longitudinal ‘head-to-tail’ interaction of lamin dimers (the so-called ACN interaction), which is crucial for filament assembly. To this end, we prepared a series of recombinant fragments of human lamin A centred around the N- and C-termini of the rod. The fragments were stabilized by fusions to heterologous capping motifs which provide for a correct formation of parallel, in-register coiled-coil dimers. As a result, we established crystal structures of two N-terminal fragments one of which highlights the propensity of the coiled-coil to open up, and one C-terminal rod fragment. Additional studies highlighted the capacity of such N- and C-terminal fragments to form specific complexes in solution, which were further characterized using chemical cross-linking. These data yielded a molecular model of the ACN complex which features a 6.5 nm overlap of the rod ends.
Highlights
Lamins represent a distinct class within the intermediate filament (IF) protein family
We showed that carefully chosen short terminal fragments of lamin rod domain can be conveniently used for studying the longitudinal assembly in molecular detail
A key element of our strategy was the use of relatively short (~50 residues) N- and C-terminal rod fragments that are stabilized by capping motifs (Figure 1)
Summary
Lamins represent a distinct class within the intermediate filament (IF) protein family. These nuclear proteins are expressed in all human cell types. Lamins provide mechanical stability, which is the core function of IF family. Lamins are involved in a broad variety of cellular processes including chromatin organization and transcription, DNA replication and repair, cell differentiation, mitosis and gene expression [2,3,4,5,6,7]. Mutations in the lamin genes cause a wide range of diseases called laminopathies, including Emery-Dreifuss muscular dystrophy and Hutchinson-Gilford progeria syndrome [8]. Malfunctioning of lamins plays a role in diabetes, heat shock and cancer [9,10,11]
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