title: PRC1-labeled microtubule bundles and kinetochore pairs show one-to-one association in metaphase.
abstract: In the mitotic spindle, kinetochore microtubules form k-fibers, whereas overlap or interpolar microtubules form antiparallel arrays containing the cross-linker protein regulator of cytokinesis 1 (PRC1). We have recently shown that an overlap bundle, termed bridging fiber, links outermost sister k-fibers. However, the relationship between overlap bundles and k-fibers throughout the spindle remained unknown. Here, we show that in a metaphase spindle more than 90% of overlap bundles act as a bridge between sister k-fibers. We found that the number of PRC1-GFP-labeled bundles per spindle is nearly the same as the number of kinetochore pairs. Live-cell imaging revealed that kinetochore movement in the equatorial plane of the spindle is highly correlated with the movement of the coupled PRC1-GFP-labeled fiber, whereas the correlation with other fibers decreases with increasing distance. Analysis of endogenous PRC1 localization confirmed the results obtained with PRC1-GFP PRC1 knockdown reduced the bridging fiber thickness and interkinetochore distance throughout the spindle, suggesting a function of PRC1 in bridging microtubule organization and force balance in the metaphase spindle.
authors.affiliation: Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Bruno
authors.initials: B
authors.lastname: Polak
authors.affiliation: Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Patrik
authors.initials: P
authors.lastname: Risteski
authors.affiliation: Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia.
authors.forename: Sonja
authors.initials: S
authors.lastname: Lesjak
authors.affiliation: Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia tolic@irb.hr.
authors.forename: Iva M
authors.identifier: 0000-0003-1305-7922
authors.initials: IM
authors.lastname: Tolić
mesh_terms: D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002877:Chromosomes, Human; D015870:Gene Expression; D020868:Gene Silencing; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism*; D008677:Metaphase; D008870:Microtubules / Q000378:metabolism*; D057054:Molecular Imaging / Q000379:methods; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D008941:Spindle Apparatus
chemical_list: D018797:Cell Cycle Proteins; C116662:PRC1 protein, human; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins
keywords: bridging fiber; interpolar microtubules; k‐fiber; mitosis; overlap microtubules