Abstract
The cross-talk between dynamic microtubules and the cell cortex plays important roles in cell division, polarity, and migration. A critical adaptor that links the plus ends of microtubules with the cell cortex is the KANK N-terminal motif and ankyrin repeat domains 1 (KANK1)/kinesin family member 21A (KIF21A) complex. Genetic defects in these two proteins are associated with various cancers and developmental diseases, such as congenital fibrosis of the extraocular muscles type 1. However, the molecular mechanism governing the KANK1/KIF21A interaction and the role of the conserved ankyrin (ANK) repeats in this interaction are still unclear. In this study, we present the crystal structure of the KANK1·KIF21A complex at 2.1 Å resolution. The structure, together with biochemical studies, revealed that a five-helix-bundle-capping domain immediately preceding the ANK repeats of KANK1 forms a structural and functional supramodule with its ANK repeats in binding to an evolutionarily conserved peptide located in the middle of KIF21A. We also show that several missense mutations present in cancer patients are located at the interface of the KANK1·KIF21A complex and destabilize its formation. In conclusion, our study elucidates the molecular basis underlying the KANK1/KIF21A interaction and also provides possible mechanistic explanations for the diseases caused by mutations in KANK1 and KIF21A.
Highlights
The cross-talk between dynamic microtubules and the cell cortex plays important roles in cell division, polarity, and migration
We show that several missense mutations present in cancer patients are located at the interface of the KANK11⁄7KIF21A complex and destabilize its formation
We further demonstrated that the KANK1 ANK domain and the kinesin family member 21A (KIF21A) KANK1-binding domain (KBD) form a stable 1:1 stoichiometric complex in solution (Fig. 1D)
Summary
KANK1 recruits KIF21A to cell cortex by binding to a 38-amino acid (aa) fragment in the middle of KIF21A (aa 1142– 1180; human KIF21A: NP_001166935) through its ANK repeats [20] (Fig. 1A). Using a truncation-based approach, a 19-residue fragment of KIF21A (aa 1138 –1156; referred to as the KANK1-binding domain (KBD)) was sufficient to bind to the KANK1 ANK domain (Kd ϳ0.58 M) (Fig. 1, B and C). We further demonstrated that the KANK1 ANK domain and the KIF21A KBD form a stable 1:1 stoichiometric complex in solution (Fig. 1D). The KANK1 ANK domain contains five ANK repeats with the typical helix-turn-helix conformation (Fig. 2, A and B). Further amino acid sequence alignment analysis of the capping domain from the KANK family of proteins reveals that both the secondary structure and the residues forming the hydrophobic core of the capping domain are highly conserved
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