Abstract
Cytoplasmic dynein is a multisubunit microtubule-based motor protein that is involved in several eukaryotic cell motilities. Two dynein heavy chains each form a motor domain that connects to a common cargo-binding tail. Although this tail domain is composed of multiple polypeptides, subunit organization within this region is poorly understood. Here we present an in vitro dissection of the tail-forming region of the dynein heavy chain from Dictyostelium. Our work identifies a sequence important for dimerization and for binding the dynein intermediate chain. The core of this motif localizes within an approximately 150-amino acid region that is strongly conserved among other cytoplasmic dyneins. This level of conservation does not extend to the axonemal dynein heavy chains, suggesting functional differences between the two. Dimerization appears to occur through a different mechanism than the heavy chain-intermediate chain interaction. We corroborate the in vitro interactions with in vivo expression of heavy chain fragments in Dictyostelium. Fragments lacking the interaction domain express well, without an obvious phenotype. On the other hand, the region crucial for both interactions appears to be lethal when overexpressed.
Highlights
Cytoplasmic dynein is a multisubunit microtubulebased motor protein that is involved in several eukaryotic cell motilities
The core of this motif localizes within an ϳ150-amino acid region that is strongly conserved among other cytoplasmic dyneins
This is the first identification of a subunit-interaction domain in a cytoplasmic dynein heavy chain, and it provides a target for investigations into the structure and assembly dynamics of the dynein tail
Summary
Cytoplasmic dynein is a multisubunit microtubulebased motor protein that is involved in several eukaryotic cell motilities. Our work identifies a sequence important for dimerization and for binding the dynein intermediate chain The core of this motif localizes within an ϳ150-amino acid region that is strongly conserved among other cytoplasmic dyneins. A fragment containing the amino-terminal region can only be expressed at low levels; its overexpression appears to be lethal [21] Because this fragment can associate with the native DHC in vivo [21], its overexpression may cause the formation of one-headed or headless dyneins that can bind cargo but cannot function as motors (see Ref. 22). We further correlate these results with expression of similar regions of the DHC in vivo This is the first identification of a subunit-interaction domain in a cytoplasmic dynein heavy chain, and it provides a target for investigations into the structure and assembly dynamics of the dynein tail
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