The dynamic protein architecture of human kinetochores

Accurate segregation of genomic DNA during mitosis is one of the cell’s most vital functions. Errors in this process result in defects that are implicated in many human diseases and developmental disorders. Key to this process is the attachment of chromosomes to microtubules of the mitotic spindle, via complex protein machines at the centromere known as kinetochores. In addition to providing the spindle microtubule attachment site, kinetochores also generate and propagate a “warning system” known as the spindle assembly checkpoint (SAC) that prevents anaphase onset until all chromosomes are properly aligned at the spindle equator. Lack of resolution capability in the light microscope and lack of contrast in the electron microscope have frustrated attempts to generate a map of protein architecture within kinetochores. Here we demonstrate a new light microscopy method, which we term K-SHREC (Kinetochore Speckle High-Resolution Colocalizaton) that measures distances between fluorescent labels at an accuracy of