The centromere, on which kinetochore proteins assemble, ensures precise chromosome segregation. Centromeres are largely specified by the histone H3 variant CENP-A (also known as Cse4 in yeasts). Structurally, centromere DNA sequences are highly diverse in nature. However, the evolutionary consequence of these structural diversities on de novo CENP-A chromatin formation remains elusive. Here, we report the identification of centromeres, as the binding sites of four evolutionarily conserved kinetochore proteins, in the human pathogenic budding yeast Candida tropicalis. Each of the seven centromeres comprises a 2 to 5 kb non-repetitive mid core flanked by 2 to 5 kb inverted repeats. The repeat-associated centromeres of C. tropicalis all share a high degree of sequence conservation with each other and are strikingly diverged from the unique and mostly non-repetitive centromeres of related Candida species—Candida albicans, Candida dubliniensis, and Candida lusitaniae. Using a plasmid-based assay, we further demonstrate that pericentric inverted repeats and the underlying DNA sequence provide a structural determinant in CENP-A recruitment in C. tropicalis, as opposed to epigenetically regulated CENP-A loading at centromeres in C. albicans. Thus, the centromere structure and its influence on de novo CENP-A recruitment has been significantly rewired in closely related Candida species. Strikingly, the centromere structural properties along with role of pericentric repeats in de novo CENP-A loading in C. tropicalis are more reminiscent to those of the distantly related fission yeast Schizosaccharomyces pombe. Taken together, we demonstrate, for the first time, fission yeast-like repeat-associated centromeres in an ascomycetous budding yeast.
Centromeres aid in high fidelity chromosome segregation. Paradoxically, centromere DNA sequences are rapidly evolving in fungi, plants, and animals. Centromere DNA sequences in fungi can be unique in each chromosome or share conserved features such as motifs for sequence specific protein binding, pericentric repeats, or transposon-rich elements. Ascomycetous fungi, in particular, show a wide range of diversity in centromere sequence elements. However, no ascomycetous budding yeast species is known to possess repeat-associated centromeres in all of its chromosomes. Here, we identified and mapped all seven centromeres in an ascomycete, a rapidly emerging human pathogenic yeast, Candida tropicalis. The repeat-associated centromeres of highly homogeneous DNA sequences in C. tropicalis are significantly diverged from the mostly non-repetitive unique centromeric DNA sequences of its closely related sequenced species, Candida albicans, Candida dubliniensis and Candida lusitaniae. Structurally, the centromeres of C. tropicalis more closely resemble those of the distantly related fission yeast Schizosaccharomyces pombe. Thus, we discover rapidly diverging repeat-associated centromeres in an ascomycetous budding yeast and provide evidence of emergence of repeat-associated centromeres via two independent evolutionary events in ascomycetous fungi.