Fig. 6: EB1; like ARK2 associates with spindle and kinetochore during male gametogony.

A Live-cell imaging of EB1-GFP (green) showing its location on spindles and spindle poles. DNA is stained with Hoechst dye (blue). More than 50 images were analysed in more than three different experiments. Scale bar = 5 μm. B ChIP-seq analysis of EB1-GFP profiles for all 14 chromosomes showing its centromeric binding. Signals are plotted on a normalised read per million (RPM) basis. Red lines at the top indicate the ends of chromosomes; circles on the bottom indicate centromere locations. NDC80-GFP was used as a positive control and IgG was used as a negative control. C Live-cell imaging showing the location of EB1-GFP (green) and kinetochore marker NDC80-mCherry (red) in a gametocyte activated for 1–2 min. DNA is stained with Hoechst dye (blue). More than 50 images were analysed in more than three different experiments. Scale bar = 5 µm. R is Pearson’s coefficient for co-localisation. D Live-cell imaging showing the location of EB1-GFP (green) and ARK2-mCherry (red) in a gametocyte activated for 1–2 min. DNA is stained with Hoechst dye (blue). More than 50 images were analysed in more than three different experiments. Scale bar = 5 µm. R is Pearson’s coefficient for co-localisation. E Live-cell imaging showing the location of EB1-mCherry (red) and basal body marker SAS4-GFP (green) in gametocytes activated for 1–2 min (upper panel) and 4 min (lower panel). DNA is stained with Hoechst dye (blue). More than 50 images were analysed in more than three different experiments. Scale bar = 5 µm. F 3D-SIM image showing location of EB1 (green) and NDC80 (purple) in a gametocyte activated for 1 min. DNA is stained with Hoechst dye (blue). More than ten images were analysed in more than three different experiments. Scale bar = 1 μm. G 3D-SIM image showing location of EB1 (green) and ARK2 (purple) in a gametocyte activated for 3–4 min. DNA is stained with Hoechst dye (blue). More than ten images were analysed in more than three different experiments. Scale bar = 1 μm. H 3D-SIM image showing location of EB1 (purple) and cytoplasmic SAS4 (green) in gametocyte activated for 1 min. DNA is stained with Hoechst dye (blue). More than ten images were analysed in more than three different experiments. Scale bar = 1 μm. I STED confocal microscopy showing co-localisation of EB1 (purple) and α-tubulin (green) at spindle but not with cytoplasmic MTs in a gametocyte activated for 1 min. DNA is stained with SiR DNA (blue). More than ten images were analysed in more than three different experiments. Scale bar = 1 μm.