Kinetochore size variation in mammalian chromosomes: an image analysis study with evolutionary implications.

The kinetochore, a proteinaceous plate that is the site for attachment of spindle microtubules to the metaphase chromosome, can be visualized using anti-kinetochore indirect immunofluorescence. We have used computer-assisted image analysis to measure the variation of kinetochore surface areas, as reflected by immunofluorescence areas, in cell lines derived from rat kangaroo, Chinese hamster and common rat, to determine if our size estimates correlate well with those obtained using measurements from electron micrographs. In addition, we used male and female human fibroblast cell lines, as well as a transformed human female cell line as well as a transformed human female cell line (HeLa), to examine kinetochore size variation among cells, between sexes, and between cell lines. We found that our system gave reproducible estimates of kinetochore size, and that these sizes correlated very well (r = 0.95) with the electron micrograph measurements. In examining variation within humans, we observed measurable differences between cell lines. Despite this difference, all the human lines had size distributions that were leptokurtotic and positively skewed. The fact that very few chromosomes exhibited areas smaller than the mode gives support to the idea that mammalian chromosomes may require a specific, minimum amount of kinetochore material in order to maintain stable attachment to the mitotic spindle. On the other hand, the positive skewness seems to indicate that larger kinetochores, possibly the result of events such as Robertsonian fusions, are fully functional. The retention of this plasticity may allow the chromosomes to maintain an evolutionary adaptability that might otherwise be lost.