Fig. 7: Interaction of bridgin with centromeric chromatin is established by the basic domain enriched with positively charged residues.

a Schematic of the bridgin constructs wherein its basic domain was replaced with the basic DNA-binding domain from human Ki67. b Representative micrographs of Bgi1^FL expressing SHR879, Bgi1^BDΔ expressing SHR880, and the domain-swap chimera Bgi1^BDΔ + Ki67^BD expressing SHR926 (SHR832::3×-FLAG-GFP-Bgi1^BDΔ+Ki67^BD). Scale bar, 3 μm. c Signal intensities of bridgin constructs mentioned in b were measured at metaphase and normalized to the mean intensity of Bgi1^FL. The data represent the results of three independent experiments with 100 cells each. The red dot represents the mean of one experiment; mean ± S.D. is shown. Kruskal–Wallis one-way analysis followed by Dunn’s multiple comparison test was used to calculate the statistical significance of differences (the p values show the difference compared to Bgi1^FL). d The extent of complementation of bridgin constructs mentioned in c, wild-type control CNVY121, and the bridgin-null mutant SHR832 was measured. Indicated cell populations were measured 9 h post incubation at 37°C. All values were normalized to wild-type control levels. Defects in nuclear segregation were measured as mentioned in Fig. 4b. The data represent the mean ± S.D. of three independent experiments. One-way ANOVA test with Tukey’s multiple comparison test was used to calculate the statistical significance of differences between the net missegregation and large-bud arrest population across strains. e Cells of varying numbers, 2 × 10⁴, 2 × 10³, 200, 100, and 50, of Bgi1^BDΔ expressing SHR880 and the domain-swap chimera Bgi1^BDΔ+Ki67^BD expressing SHR926 strains, were spotted on mentioned media plates. f Experimental design to determine Bgi1-BD interaction with chromatin in vivo by MNase IP. g FLAG immunoprecipitation of kinetochore proteins. 3×FLAG-tagged proteins of Spc25 (SHR861,SPC25::SPC25-3×FLAG), CENP-C^Mif2 (SHR896), FLAG-GFP (FG) control (SHR918), Bgi1^FL (SHR879), Bgi1^BDΔ (SHR880), Bgi1^BD (SHR917), the domain-swap chimera Bgi1^BDΔ+Ki67^BD (SHR926), and untagged wild-type control (H99) were pulled down and levels of interacting histone H4 were determined. Bait proteins are marked by blue arrows. h Relative enrichment of FLAG-tagged proteins described in g at centromeres following the MNase IP. qPCR was performed using CEN2 and CEN14 and nonentromeric primer sets described in Fig. 2a followed by normalization to noncentromeric controls. The data represent the mean ± S.D. of two independent experiments. Source data are available as a Source Data file.