Centromeres both mediate and
monitor interaction of chromosomes with the mitotic spindle. Budding
yeast centromeres are specified by a tripartite CEN DNA motif.
Five proteins that bind CEN DNA in vitro have been identified. We
have shown that two of these, Ndc10p and Cbf1p, are indeed centromere
proteins using in vivo cross-linking followed by immunoprecipitation.
The essential Cse4 and Mif2 proteins also affect centromere function
based on their mutant phenotypes and genetic interactions with
centromere components. Importantly, Cse4p and Mif2p share homology with
the mammalian centromere proteins, CENP-A and CENP-C. Cse4p and Mif2p
have eluded biochemical detection as CEN DNA binding proteins.
However, by cross-linking, both proteins interact specifically with
CEN DNA in vivo, providing evidence for structural conservation
between yeast and mammalian centromeres. By fine mapping, none of the
four interactions extends beyond the vicinity of the CEN DNA.
This is especially interesting for Cse4p, which like CENP-A, possesses a
C-terminal histone fold similar to that of histone H3. Cse4p genetically
interacts with histone H4, and mutations in both Cse4p and histone H4
affect centromeric chromatin structure. Thus, functional centromeres in
yeast may be built in the context of a single specialized Cse4p-containing nucleosome. We are defining the structural requirements for
the Cse4p- CEN DNA interaction and using Cse4p as a cytological
marker for centromere assembly and position.
Return to YGM 1998 Abstract Index