Faithful chromosome segregation is crucial for fertility, development, and tissue homeostasis. Despite such highly conserved roles, genomic analyses suggest that the chromosome segregation machinery evolves extremely rapidly. This is a fascinating conundrum because chromosomes must segregate accurately to ensure organismal viability and reproduction. Solving this puzzle requires experimental systems that test hypotheses emerging from genomic analyses. In the Dudka Lab, we are building such systems by combining molecular evolution, gene editing, experimental evolution, confocal microscopy, and mouse models.
Functional Constraints of Centromeric DNA Evolution
Highly repetitive centromeric DNA satellites occupy 6.2% of the human genome and can differ 37-fold between individuals. Despite mounting evidence that this variation is deleterious (linked to Down syndrome and poor cancer prognosis), the functional constraints that result in this divergence are very poorly understood. We discovered that two recently emerged mouse centromeric satellites have distinct DNA shapes due to sequence divergence. Using hybrid mouse oocytes, we showed that architectural proteins from the HMGA family recognize the differences in those DNA shapes and then tightly package one of those centromeric satellites but not the other. Satellite packaging failure is catastrophic and disrupts chromosome segregation. Therefore, we proposed a model that can explain why closely related species show striking differences in satellite sequence and copy number. Under our model, differences in DNA shape-recognizing architectural proteins across species contribute to the extraordinary satellite divergence (Dudka et al., Nature 2025; Dudka et al., in revision). We use mammalian tissue culture cells as a model of satellite expansion, leveraging experimental evolution assays and digital droplet PCR to understand the role of architectural proteins and DNA packaging in satellite divergence.
Functional Impacts of Adaptive Protein Evolution
Our molecular evolution analyses reveal that a whopping 30% (out of 100) of centromeric proteins show signatures of adaptive evolution in rodents and primates. These adaptations occur in proteins with diverse functions: centromere assembly, microtubule attachment modulation, and stabilization of correct attachments (Dudka et al., Journal of Cell Biology 2023). Our work on a critical centromeric protein that creates microtubule attachments at centromeres, CENP-T, suggests that adaptive evolution modulates centromere protein function and promotes robust female gametogenesis (Dudka et al., Current Biology 2025). We can discover how these adaptations maintain faithful segregation by creating "mal-adapted" alleles using gene editing to swap adaptively evolving regions between closely related rodent and primate species, creating evolutionary mismatches between the protein and the centromere. We are now systematically testing how adaptations in functionally distinct centromeric proteins regulate chromosome segregation and female fertility using cell lines and mouse models.
Health Implications of Mutations in Adaptive Sites
Chromosome mis-segregation underlies infertility, congenital diseases, and cancer. The vast majority of missense mutations in centromeric proteins are variants of uncertain significance (VUS). Given that many of these mutations occur in adaptively evolving sites, we hypothesize that molecular evolution analyses can reveal new disease-associated mutations in patients,
especially those suffering from infertility. We aim to identify new disease-associated mutations in centromeric proteins by intersecting molecular evolution analyses with known human mutations. We use gene editing in human cells and confocal microscopy to test the impact of these mutations on chromosome segregation.