Epistasis, aneuploidy, and functional mutations underlie evolution of resistance to induced microtubule depolymerization

Abstract

Cells with blocked microtubule polymerization are delayed in mitosis, but eventually manage to proliferate despite substantial chromosome missegregation. While several studies have analyzed the first cell division after microtubule depolymerization, we have asked how cells cope long-term with microtubule impairment. We allowed 24 clonal populations of yeast cells with beta-tubulin mutations preventing proper microtubule polymerization, to evolve for ~150 generations. At the end of the laboratory evolution experiment, cells had regained the ability to form microtubules and were less sensitive to microtubule-depolymerizing drugs. Whole-genome sequencing identified recurrently mutated genes, in particular for tubulins and kinesins, as well as pervasive duplication of chromosome VIII. Recreating these mutations and chromosome VIII disomy prior to evolution confirmed that they allow cells to compensate for the original mutation in beta-tubulin. Most of the identified mutations did not abolish function, but rather restored microtubule functionality. Analysis of the temporal order of resistance development in independent populations repeatedly revealed the same series of events: disomy of chromosome VIII followed by a single additional adaptive mutation in either tubulins or kinesins. Since tubulins are highly conserved among eukaryotes, our results have implications for understanding resistance to microtubule-targeting drugs widely used in cancer therapy.

Multiple routes of adaptation to high levels of CIN and aneuploidy in budding yeast

“…This timeline of adaptation supports the theory that aneuploidy often provides a rapid but temporary form of adaptation, as has previously been observed in yeast adapted to heat stress or a tubulin mutation (Yona et al, 2012; Pavani et al, 2021). In both of those studies, a single chromosome was gained early in the adaptation process and sometimes lost again at later time points.…”

A fitness trade-off explains the early fate of yeast aneuploids with chromosome gains

“…In yeast, the development of aneuploidy resulting from an accidental chromosome missegregation has been characterized with massive experimental data [3–6, 6–10, 13, 14, 14–16]. As a consequence of this major effort, we are in need of unifying principles to rationalize this wealth of data, and embed the underlying evolutionary dynamics into simple quantitative models.…”

“…The model introduced here can be extended to describe more complex scenarios. First, it can be applied to investigate the evolutionary consequences of a sudden increase of the mis-segregation rate [6], which can result from the usage of anti-mitotic drugs. Second, it can be used as an ingredient to build models of chromosomal instability, with a clear interest for cancer development [53].…”

“…The evolutionary dynamics leading to the emergence of aneuploidy is typically investigated with yeast models, because they can be manipulated with advanced genetics and cell- and molecular-biology methods, and hence be used to create isogenic backgrounds that only differ from each other by chromosome ploidy number, offering a direct point of comparison between euploid and aneuploid strains. Moreover, yeast models can be easily investigated with laboratory-evolution experiments, thanks to their short replication time [6, 9, 13, 14].…”

“…In the case of yeast, the literature offers extensive phenotypic data, for example growth curves of aneuploids in several environmental conditions as well as in laboratory-evolution experiments [6,9,10,[13][14][15], offering the opportunity to test for unifying trends. The available modelling studies presented so far have focused on the effect of aneuploidy on cell growth and physiology [7,[22][23][24][25].…”