Disordered Motifs and Domains in Cell Control
Poster Session I
15-POS
Board 15
The Non-structured Amino-terminal Domain of Intermediate Filaments Powers Assembly
Harald Herrmann
1
, Tatjana Wedig
1
, Norbert Mücke
1
, Ueli Aebi
2
.
1
German Cancer Research Center, Heidelberg, Germany,
2
University of Basel, Basel,
Switzerland.
Intermediate filament (IF) proteins are principal structures of the metazoan cytoskeleton. The IF
multi-gene family represents fibrous proteins consisting of a central alpha-helical "rod" domain
flanked by non-alpha-helical amino- (“head”) and carboxy-terminal (“tail) domains. IF-proteins
form robust but highly flexible fibers and networks, both in the cytoplasm and the nucleus, that
are not soluble under physiological conditions. Their role for the mechanical properties of cells is
being elucidated by recent work quite convincingly. However, the assembly mechanism is still
not completely understood, in particular the role of the non-structured “head” is by no means
clear. At the molecular level, IF-proteins form coiled coils that laterally associate in an anti-
parallel fashion to yield tetrameric complexes, which are stable under low ionic strength
conditions. Hence, the assembly module for IFs is non-polar. The principal reaction for the
elongation of IF proteins is a “head-to-tail” association of the “rods” with a 2 to 4 nm overlap.
The “head” domain is essential for assembly, both laterally and longitudinally, as headless IF-
proteins form only dimers under tetramer conditions, and tetramers under filament forming
conditions. In order to investigate the contribution of individual parts of the “head” domain in
these two assembly reactions, we have subjected the fibroblast-specific IF-protein vimentin to
systematic truncation by recombinant techniqiues within the “head” domain. The assembly
properties were followed by analytical ultracentrifugation, electron and atomic force microscopy
in vitro and by transfection of the corresponding cDNA clones into vimentin-free fibroblasts
generated from vimentin-(-/-) embryos. We demonstrate that the first 20 amino acids of the 85
amino acid-long "head" are dispensable for both tetramer and IF formation. Further truncation
abolishes filament-forming capacity completely. Truncation of more than 40 amino acids impairs
tetramer formation indicating the “head” is needed for orderly coiled-coil alignment.
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