Bradley David, Hogrebe Alexander, Dandage Rohan, Dubé Alexandre K, Leutert Mario, Dionne Ugo, Chang Alexis, Villén Judit, Landry Christian R
Institut de Biologie Intégrative et des Systèmes (IBIS), Université Laval, Québec, QC, Canada.
Department of Biochemistry, Microbiology and Bioinformatics, Université Laval, Québec, QC, Canada.
bioRxiv. 2023 Oct 10:2023.10.08.561337. doi: 10.1101/2023.10.08.561337.
The fidelity of signal transduction requires the binding of regulatory molecules to their cognate targets. However, the crowded cell interior risks off-target interactions between proteins that are functionally unrelated. How such off-target interactions impact fitness is not generally known, but quantifying this is required to understand the constraints faced by cell systems as they evolve. Here, we use the model organism to inducibly express tyrosine kinases. Because yeast lacks tyrosine kinases, most of the resulting tyrosine phosphorylation is spurious. This provides a suitable system to measure the impact of artificial protein interactions on fitness. We engineered 44 yeast strains each expressing a tyrosine kinase, and quantitatively analysed their phosphoproteomes. This analysis resulted in ~30,000 phosphosites mapping to ~3,500 proteins. Examination of the fitness costs in each strain revealed a strong correlation between the number of spurious pY sites and decreased growth. Moreover, the analysis of pY effects on protein structure and on protein function revealed over 1000 pY events that we predict to be deleterious. However, we also find that a large number of the spurious pY sites have a negligible effect on fitness, possibly because of their low stoichiometry. This result is consistent with our evolutionary analyses demonstrating a lack of phosphotyrosine counter-selection in species with tyrosine kinases. Taken together, our results suggest that, alongside the risk for toxicity, the cell can tolerate a large degree of non-functional crosstalk as interaction networks evolve.
信号转导的保真度需要调节分子与其同源靶标结合。然而，细胞内部拥挤，存在功能不相关蛋白质之间的非靶标相互作用风险。这种非靶标相互作用如何影响适应性通常尚不清楚，但量化这一点对于理解细胞系统在进化过程中面临的限制是必要的。在这里，我们使用模式生物来诱导表达酪氨酸激酶。由于酵母缺乏酪氨酸激酶，因此产生的大多数酪氨酸磷酸化都是假的。这提供了一个合适的系统来测量人工蛋白质相互作用对适应性的影响。我们构建了44个酵母菌株，每个菌株都表达一种酪氨酸激酶，并对它们的磷酸化蛋白质组进行了定量分析。该分析导致约30000个磷酸化位点映射到约3500种蛋白质。对每个菌株适应性成本的检查显示，假磷酸化酪氨酸（pY）位点的数量与生长下降之间存在很强的相关性。此外，对pY对蛋白质结构和蛋白质功能影响的分析揭示了1000多个我们预测有害的pY事件。然而，我们也发现大量的假pY位点对适应性的影响可以忽略不计，这可能是因为它们的化学计量较低。这一结果与我们的进化分析一致，该分析表明在具有酪氨酸激酶的物种中缺乏磷酸酪氨酸反选择。综上所述，我们的结果表明，除了毒性风险外，随着相互作用网络的进化，细胞能够容忍很大程度的无功能串扰。
