A unique kinesin-8 surface loop provides specificity for chromosome alignment.
Bottom Line: To address this question, we engineered chimeric kinesins that contain the Kif4A, Kif18B (kinesin-8), or Kif5B (kinesin-1) motor domain fused to the C-terminal tail of Kif18A.Mutational studies of Kif18A indicate that this control depends on both its C-terminus and a unique, positively charged surface loop, called loop2, within the motor domain.These data support a model in which microtubule-attenuating kinesins are molecularly "tuned" to control the dynamics of specific subsets of spindle microtubules.
Affiliation: Department of Molecular Physiology and Biophysics, University of Vermont, Burlington, VT 05405.Show MeSH
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Mentions: To determine whether loop2 is required for Kif18A's regulation of K-fiber lengths during mitosis, we assayed the ability of our chimeric kinesins and loop2 mutants to facilitate chromosome alignment and regulate spindle length in cells depleted of endogenous Kif18A. Cells depleted of Kif18A were transfected with GFP-tagged kinesins and then treated ∼18 h later with MG132 (20 μM), which prevents anaphase entry (Figure 6A). Cells were then fixed and immunofluorescently stained with antibodies against centromeric (ACA) and centrosomal (γ-tubulin) proteins. Under these conditions, the majority of cells transfected with scrambled control siRNAs and a GFP control plasmid had well-aligned kinetochores. In contrast, cells transfected with Kif18A siRNAs and a GFP control plasmid displayed unaligned kinetochores (Figure 6A).
Affiliation: Department of Molecular Physiology and Biophysics, University of Vermont, Burlington, VT 05405.