Bridging telomeres, centromeres and CENP-B in Chinese Hamster

The centromere is a specialized nucleoprotein structure of the eukaryotic chromosomes which ensures proper chromosome segregation of sister chromatids during mitosis and meiosis. Despite the centromeric function is well conserved among taxa, centromeric DNA sequences are highly divergent. Indeed, centromeres are epigenetically defined by the histone H3-variant Centromere Protein A (CENP-A), whereas satellite DNA sequences have been proposed to contribute to centromere stability and organization. In spite of their divergence, satellite DNA sequences share a highly conserved 17-nucleotide motif, named CENP-B box, which is recognized by CENP-B, the unique centromeric protein to possess a DNA sequence-binding specificity, suggesting that CENP-B binding is an important feature of mammalian centromeres. On the other hand, the protein is dispensable for centromere function and the exact role of CENP-B is still controversial. Previous work from our laboratory demonstrated that, in the rodent species Chinese Hamster (Cricetulus griseus) large clusters of telomeric-like TTAGGG repeats were present at centromeric positions as consequence of chromosomal fusions and fission events during karyotype evolution. Although telomeric-like repeats represented the major centromeric satellite DNA sequences, they do not contain any CENP-B binding sites, suggesting a peculiar organization of centromeres in this species. In this thesis work, we investigated the distribution of CENP-B and its binding sites and the overall organization of Chinese hamster centromeres using a combination of cytogenetic and molecular approaches. In particular, we demonstrated that not all centromeres are bound by CENP-B, suggesting an uncoupling between CENP-B and the centromeric function. In addition, we demonstrated by ChIP-seq that the CENP-B binding site is contained into a 79-bp satellite DNA family which is not strictly centromeric but rather pericentromeric. Finally, we characterized by ChIP-seq Chinese hamster centromeric domains demonstrating at the molecular level that telomeric-like repeats are associated with the centromeric function. Surprisingly, we identified centromeric domains composed by satellite-less sequences. In our laboratory, we identified satellite-less centromeres in equid species and this finding reveals that, also in Chinese hamster, satellite DNA is uncoupled from centromeric function. Moreover, we characterized at the molecular level an active centromere which originated at the fusion point between two ancestral chromosomes. In this centromere, telomeric-like sequences are maintained in the middle of a single-copy sequence. To our knowledge, this is the first molecular report of a fusion event involving telomeres that resulted in a centromerization event at the fusion point. Taken together, these results strongly indicate that Chinese Hamster may represent a valuable model system for centromere biology.