Dissecting the evolution of resistance to microtubule hyper-stabilization
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Date
2024-01-31Embargo Date
2024-07-29
Author
Macaluso, Francesca
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Abstract
When microtubules are hyper-stabilized, they do not provide the proper
tension at the kinetochore of the metaphase chromosomes. Low tension leads
to the activation of the spindle assembly checkpoint which stops cells in
mitosis. Eventually cells can bypass the checkpoint and resume proliferation
with defective microtubules; some cells die in this process, but others
missegregate chromosomes and eventually become resistant to the
microtubule hyper-stabilizing stress. For example, cancer cells develop
resistance to chemotherapeutic drugs that hyper-stabilize microtubules such
as taxanes. Here, we aimed to investigate the evolution of resistance to
microtubule hyper-stabilizing conditions. We characterized in yeast the tub2-
150 mutation which hyper-stabilizes microtubules similarly to taxanes and
shows a growth defect compared to WT cells. We evolved in the laboratory
haploid yeast cells carrying the tub2-150 mutation for over ~190 generations.
We then investigated by Next Generation Sequencing the genetic alterations
in cells that had resumed cell proliferation. We found that tub2-150 mutants
were able to evolve and recover the initial growth defect by two temporally
distinct solutions. The first evolutionary solution was the selection of
recurrent aneuploidies, in particular the disomy of chromosome XII. We
found that overexpression of Stu2, a regulator of microtubule-kinetochore
attachments, was responsible for the disomy. We confirmed that STU2
duplication was able to partially rescue the tub2-150 growth defect by
lowering the mis-segregation rate of the mutant. In the long-term,
aneuploidies were replaced by point mutations of the TUB genes. Some
representative chosen TUB mutations, when re-introduced in the mutant
ancestor before evolution, largely rescued the tub2-150 growth defect by
destabilizing microtubules to different extents. Interestingly, several
compensatory TUB mutations that emerged during the evolution experiment
had been previously found in mammalian cell lines resistant to taxane-like
treatments or in breast cancer patients. These results provide new insights on
the evolutionary dynamics underlying resistance to microtubules hyper stabilization and potentially lead to the development of new strategies in order
to stop or delay the emergence of resistance to drugs which stabilize
microtubules.