Nuclear proteins that become part of the mitotic apparatus: a role in nuclear assembly?

A structure located at the poles of the mitotic spindle is described, which may function as a centre for post-mitotic nuclear assembly. Evidence in support of this function is incomplete, but comes from two different kinds of experiments, which are reviewed here. First, fluorescence microscopy studies show that mitotic chromosomes at telophase or late anaphase are drawn into juxtaposition with this polar structure and second, the structure is made up in part of a non-histone chromosomal protein that in interphase cells can be detected only in the nucleus. Studies of this nuclear-mitotic apparatus protein (NuMA protein) are reported here. Monoclonal antibodies specific for the NuMA protein have been used in immunofluorescence studies to visualize the prenucleus-like polar structure and to identify the NuMA protein by immunoblotting after electrophoretic separation. The NuMA protein is a non-histone chromosomal protein of molecular weight 250000 relative to standard protein molecular weight markers in sodium dodecyl sulphate/polyacrylamide gel electrophoresis. Experiments are described that indicate several difficulties in studying the possible affinity and association of NuMA protein with mitotic chromosomes. Metaphase chromosomes isolated by the polyamine procedure of Lewis and Laemmli have bound NuMA protein detectable by immunofluorescence or by immunoblotting, but measurements made at different stages of chromosome purification show that most of the NuMA protein is separated from the chromosomes using this purification procedure. Chromosomes purified from mixtures of human and Chinese hamster cells (the latter have none of the human form of NuMA recognized by a monoclonal antibody) have human NuMA protein bound to the hamster chromosomes. Results suggest that in cell extracts exchange reactions of NuMA protein can occur, which must be avoided in the study of its natural function.