title: A unique tubulin antiserum attenuates the rate of poleward chromosome movement in anaphase.
abstract: An antiserum against tubulin, NS20, was previously shown to specifically attenuate both fast axonal transport in vivo (Johnston, K. M. et al., Brain Res. 385, 38-45 (1986)) and in vitro (Johnston, K. M. et al., Cell Motil. Cytoskel. 7, 110-115 (1987)) and flagellar motility (Goldsmith, M. et al., Cell Motil. Cytoskel. 20, 249-262 (1991)). We hypothesized that NS20 blocked motility by binding to a multifunctional motor binding domain on the microtubules (MTs), or axonemes. Here we have examined the effect of microinjecting NS20, at metaphase, into dividing PtK2 cells. Plotting chromosome separation (CS) as a function of time, we report here that CS rates for anaphase A (chromosome-to-pole movement) were reduced by approximately 50% relative to uninjected controls. CS rates for anaphase B (spindle pole elongation) were unaffected by the NS20 antiserum. The inhibition of CS rate during anaphase A by NS20 was significantly greater than the inhibition caused by a control antitubulin serum (PC5). Two possible mechanisms underlying NS20's inhibition of CS during anaphase A were considered. NS20 could block the binding of a kinetochore-associated motor to kinetochore MTs (kMTs) or, alternatively, NS20 could stabilize kMTs against depolymerization. Our results favor the first alternative. In a cold-induced depolymerization assay, NS20 had no selective stabilizing effect on MTs. Moreover, we show that NS20 can selectively block the binding of a well characterized MT-associated motor (kinesin) to MTs, in vitro. These results suggest that NS20 may be defining a unique tubulin binding domain common to the motors underlying vesicle transport, flagellar motility, and chromosome movements during anaphase A.
authors.affiliation: Department of Anatomy, Faculty of Medicine, University of Toronto, Ontario, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Goldsmith
authors.forename: S
authors.initials: S
authors.lastname: Leyland
authors.forename: J A
authors.initials: JA
authors.lastname: Connolly
authors.forename: D
authors.initials: D
authors.lastname: van der Kooy
mesh_terms: D000705:Anaphase / Q000187:drug effects*; D000818:Animals; D000906:Antibodies / Q000378:metabolism*; D001666:Binding Sites, Antibody; D002460:Cell Line; D002875:Chromosomes / Q000187:drug effects; D016547:Kinesins / Q000037:antagonists & inhibitors; D007614:Macropodidae; D008845:Microinjections; D008870:Microtubules / Q000378:metabolism*; D014404:Tubulin / Q000276:immunology* / Q000378:metabolism
chemical_list: D000906:Antibodies; D014404:Tubulin; D016547:Kinesins