不同温度下酿酒酵母细胞分裂周期蛋白Cdc5在有丝分裂中的分子动力学研究

Kinetics of cell division cyclin protein Cdc5 in budding yeast during mitosis at different temperatures

摘要:

[目的] 探究不同温度下酿酒酵母细胞分裂周期蛋白Cdc5蛋白在有丝分裂中的分子动力学变化。[方法] 本研究以酿酒酵母（Saccharomyces cerevisiae）为材料，采用活细胞成像的方法，探究Cdc5蛋白在不同温度下在酿酒酵母有丝分裂过程中的精细分子动力学变化；通过测量OD₅₉₅绘制生长曲线图，看其宏观的分裂情况是否与微观下Cdc5蛋白的分子动力学变化一致；利用流式细胞术检测细胞的细胞周期变化的情况。[结果] 在胞质分裂时，Cdc5蛋白从母细胞进入子细胞，并在芽颈处发生聚集。25℃条件下细胞中Cdc5蛋白在芽颈处的聚集时间长，37℃条件下Cdc5蛋白在芽颈处聚集时间短，两者间存在显著差异；但两个温度下，细胞中Cdc5蛋白的表达量没有显著性差异。同时，温度也会影响Cdc5蛋白在降解过程中的动力学行为，包括Cdc5蛋白在母细胞与子细胞中荧光强度峰值出现的次数和时间。生长曲线结果显示，酿酒酵母单一细胞分裂周期的变化影响了其宏观的细胞生长，且酵母分裂速度越快，子细胞长宽比越小；细胞周期结果表明，37℃下Cdc5蛋白的动力学变化与酿酒酵母细胞周期变化一致，酿酒酵母细胞周期从G₀/G₁期进入S期，亦加速了酿酒酵母的分裂。[结论] 本研究首次探究了不同温度下酿酒酵母有丝分裂中Cdc5蛋白的精细分子动力学及对应的酵母的宏观生长情况，结果表明温度会对Cdc5蛋白的动力学产生影响，且其精细分子动力学与酿酒酵母的分裂速度成正相关，该结果为进一步研究其在细胞有丝分裂中的功能提供了前期研究基础。

关键词:酿酒酵母;有丝分裂;Cdc5蛋白;分子动力学;细胞周期

Abstract:

[Objective] To explore the molecular dynamics changes of cell division cyclin protein Cdc5 in budding yeast during mitosis at different temperatures.[Methods] In this study, Saccharomyces cerevisiae was used as the experimental material objective to explore the molecular dynamic changes of Cdc5 protein in the mitosis process of budding yeast at different temperatures by using living cell imaging method; draw the growth curve by measuring OD₅₉₅ to see whether the macro division is consistent with the micro dynamic changes of Cdc5 protein; use flow cytometry to detect the cell cycle changes.[Results] During cytokinesis, Cdc5 protein entered into daughter cells from mother cells and aggregated at bud neck. The aggregation time of Cdc5 protein at the bud neck was long at 25℃, and the aggregation time of Cdc5 protein at the bud neck was short at 37℃. There was a significant difference between them. However, there was no significant difference in the expression of Cdc5 protein between the two temperatures. At the same time, the temperature also affected the dynamics of Cdc5 protein during degradation process, including the occurence of frequency and time of the peak of fluorescence intensity of Cdc5 in mother cells and daughter cells. The growth curve results showed that the single cell division cycle of budding yeast affected its macroscopic cell growth, and the faster the division rate of budding yeast, the smaller the ratio of length to width of the daughter cell. The cell cycle results showed that the dynamic changes of Cdc5 protein at 37℃ were consistent with the cell cycle changes of budding yeast. The cell cycle results showed that the cell cycle of budding yeast changed from G₀/G₁ phase to S phase at 37℃, which also accelerated the division of budding yeast.[Conclusion]] This study was to explore the molecular dynamics of Cdc5 protein in the mitosis of budding yeast at different temperatures and the corresponding macro growth of budding yeast for the first time. The results showed that the temperature had an effect on the dynamics of Cdc5 protein, and its molecular dynamics was positively correlated with the division speed of budding yeast, which provided a basis for further study of its function in mitosis.

Key words:budding yeast;mitosis;cell division cycle protein 5;molecular dynamics;cell cycle