Inhibition of anaphase spindle elongation in vitro by a peptide antibody that recognizes kinesin motor domain

• Published in: Proceedings of the National Academy of Sciences - PNAS 1993-07, Vol.90 (14), p.6611-6615
• Main Authors: Hogan, C.J, Wein, H, Wordeman, L, Scholey, J.M, Sawin, K.E, Cande, W.Z
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - pharmacology; ALGAE; Amino Acid Sequence; Anaphase; Anaphase - physiology; Antibodies; Antibodies - pharmacology; Antibody Specificity; ANTICORPS; ANTICUERPOS; Antigens; BACILLARIOPHYCEAE; Binding, Competitive; Biological and medical sciences; Cell biology; Cell cycle, cell proliferation; Cell division; Cell physiology; Cells; Cylindrotheca fusiformis; Diatoms; Fundamental and applied biological sciences. Psychology; Gels; Immunity (Disease); Kinesin - immunology; Kinesin - physiology; Marine; Microtubules; MITOSE; MITOSIS; Mitotic spindle apparatus; Molecular and cellular biology; Molecular Sequence Data; Nervous system; PEPTIDE; Peptide Fragments - immunology; PEPTIDOS; PROTEINAS; PROTEINE; Spindle Apparatus - drug effects; Spindle Apparatus - physiology; Spindle Apparatus - ultrastructure
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.90.14.6611

Isolated central spindles or spindles in detergent-permeabilized cells from the diatom Cylindrotheca fusiformis can undergo ATP-dependent reactivation of spindle elongation in vitro. We have used a peptide antibody raised against a 10-amino acid portion common to the kinesin superfamily motor domain to look for kinesin-like motor activity during anaphase B of mitosis. The peptide antibody localizes to central spindles. Upon ATP reactivation of spindle elongation, antigens recognized by the antibody are associated exclusively with the central spindle midzone where antiparallel microtubules of each half-spindle overlap. The antibody recognizes several polypeptides by immunoblot using isolated spindle extracts. One of these polypeptides behaves like kinesin with respect to nucleotide-specific binding to and release from taxol-stabilized microtubules. Preincubation of the spindle model with the peptide antibody inhibits subsequent ATP reactivation of spindle elongation. Coincubation of the peptide antibody with peptide antigen rescues spindle function. These results support a role for kinesin-related protein(s) in spindle elongation (anaphase B) of mitosis and suggest that one or several polypeptides that we have identified in spindle extracts may fulfill this function.