Parallel reorganization of protein function in the spindle checkpoint pathway through evolutionary paths in the fitness landscape that appear neutral in laboratory experiments

Abstract

Regulatory networks often increase in complexity during evolution through gene duplication and divergence of component proteins. Two models that explain this increase in complexity are: 1) adaptive changes after gene duplication, such as resolution of adaptive conflicts, and 2) non-adaptive processes such as duplication, degeneration and complementation. Both of these models predict complementary changes in the retained duplicates, but they can be distinguished by direct fitness measurements in organisms with short generation times. Previously, it has been observed that repeated duplication of an essential protein in the spindle checkpoint pathway has occurred multiple times over the eukaryotic tree of life, leading to convergent protein domain organization in its duplicates. Here, we replace the paralog pair in S. cerevisiae with a single-copy protein from a species that did not undergo gene duplication. Surprisingly, using quantitative fitness measurements in laboratory conditions stressful for the spindle-checkpoint pathway, we find no evidence that reorganization of protein function after gene duplication is beneficial. We then reconstruct several evolutionary intermediates from the inferred ancestral network to the extant one, and find that, at the resolution of our assay, there exist stepwise mutational paths from the single protein to the divergent pair of extant proteins with no apparent fitness defects. Parallel evolution has been taken as strong evidence for natural selection, but our results suggest that even in these cases, reorganization of protein function after gene duplication may be explained by neutral processes.

Author summary

Parallel evolution of protein domain organization following gene duplication has been demonstrated in the spindle checkpoint pathway leading to the hypothesis that this organization is likely to be adaptive. We test this hypothesis by reconstructing budding yeast strains with a spindle checkpoint pathway containing a protein with ancestral domain organization, and systematically perform stepwise duplication, degeneration and complementation of the duplicated protein. We show that, under laboratory conditions where the spindle checkpoint pathway is necessary for growth, degeneration of the ancestral pathway organization to the extant sub-functionalized proteins is consistent with a neutral model of duplication-degeneration-complementation.