Fig. 1

Borealin binds microtubules through its N-terminal region. a Western blots of input, supernatant (S) and pellet (P) fraction of microtubule co-sedimentation assay with 100 nM ISB^WT, ISB^MTBM and ISB^Δ20 and indicated concentration of microtubules; probed with anti-Borealin antibody. b Graph from three independent microtubule co-sedimentation assays (mean ± s.d.), small graph is log₁₀ scale and large graph is linear scale. ISB^WT is in red, ISB^MTBM is in green and ISB^Δ20 is in blue. c Representative kymographs (from two independent experiments with 150 tracks) of single molecules of ISB-GFP diffusing on taxol-stabilized microtubules, ISF-GFP concentration 100 pM. Black scale bar 5 μm, red scale bar 1 s. d Schematic showing multiple protein interaction regions on Borealin (microtubule-binding region is characterized in this paper). Multiple sequence alignment of Borealin, basic residues are shown in green, residues important for microtubule binding are indicated with blue asterisk. e Crystal structure of INCENP-survivin-Borealin (PDB: 2QFA); microtubule binding residues R17, R19, and K20 are highlighted in dark blue. f Images from microtubule bundling assay ISB^WT, ISB^MTBM and ISB^Δ20 were incubated with 1 μM taxol-stabilized microtubules and probed with anti-tubulin and anti-6His antibody. 6His-ISB is shown in green and microtubules are shown in red (representative images from one of two independent experiments). Scale bar is 5 μm