The mechanisms involved in mitotic movements are central to our knowledge of cell biology yet remain poorly understood. Understanding the regulation of these mechanisms is crucial to development of an integrated model of cell division and cellular motility in general. Recently a model system has been developed where anaphase B (spindle elongation) can be studied in vitro using isolated, functional spindles from a diatom. Using this system it was found that spindle elongation in vitro requires a highly phosphorylated environment. Several phosphorylated proteins associated with the microtubule overlap zone were also identified. Protein phosphorylation thus may play a key role in the regulation of mitosis. The identification and characterization of the protein kinase(s) involved is proposed. An in vitro quantitative assay will be developed whereby phosphorylation levels of overlap zone proteins can be evaluated. Phosphorylated overlap zone proteins may be integral to spindle motor function. Also proposed is an immuno-analysis of these proteins. A monoclonal antibody library will be raised against phosphorylated overlap zone proteins. Any antibodies found to inhibit anaphase B will be used to purify the antigen for in vitro characterization. Antibodies also will be tried on higher plant cells to see if these proteins are common to the higher plant mitotic machinery as well. Mitosis is central to genetic inheritance in eukaryotic cells. Knowledge of the molecular mechanisms involved with the ordered movement of chomosomes is essential to any understanding of the mechanics of cell division as well as the transmission of genetic information. The results of this study should greatly enhance the minimal understanding we now have.
