Abstract

The regional kinetochores of vertebrate chromosomes are propagated stably over thousands of generations. Kinetochores assemble on preferred DNA sequences, but there is no absolute sequence requirement. This, and studies of highly unusual human stable dicentric chromosomes have led to the hypothesis that kinetochore assembly has an epigenetic component. This was supported by the discovery that CENP -A, a critical determinant that marks the site of kinetochore assembly, is a specialized centromeric H3 family member. Thus kinetochores are built on a platform of specialized chromatin. I will discuss work in which we use a synthetic artificial chromosome that was designed to allow targeting of chimeric molecules into the kinetochore chromatin for “epigenetic engineering” of that chromatin. This approach enables us to modify one single kinetochore within a human cell, whilst leaving all other kinetochores untouched and fully functional. Our aim is to identify the suite of chromatin posttranslational modifications that is conducive to kinetochore assembly and maintenance, and to determine the functional consequences when these modifications are removed or altered. Our data have thus far uncovered a remarkable plasticity of functional centromeres in vivo. Kinetochores can tolerate profound changes in their chromatin environment, but they appear to be critically sensitive to the level of centromeric transcription.