Analyses of intercellular interaction and genome structure change during heat tolerant evolution by using experimental evolution system with Escherichia coli

我们研究了细胞和大肠杆菌在适应热应激（高温；39度以上）过程中的种群水平动态。首先，我们构建了保持特定生长速率的连续转移培养体系，然后将大肠杆菌培养物转移到比以往温度高2度的高温环境中。我们可以进化出大肠杆菌，使其能够接受45度的最高温度。大肠杆菌可以在最低条件的培养基中存活。在39度适应期，刚经过温度转变，大肠杆菌群体的生长速度迅速下降，荧光标准差在2~7d内增强，随后生长速度恢复，GFP SD在10 ~ 20d内下降，并收敛到39度特异性值。这些结果表明，温度变化引起的胞内条件多样性可能使大肠杆菌种群恢复可塑性，并通过多样性促进适应。在45度适应初期，大肠杆菌能以细胞密度依赖的方式生长。这种依赖性来源于大肠杆菌分泌的因子，因此，温度变化可能诱导群体感应。我们一直在大肠杆菌培养上清中寻找相应的分泌因子。此外，大肠杆菌细胞的平均基因组DNA含量随温度的升高而增加，鸟嘌呤和胞嘧啶含量随温度的升高而降低。结果表明，大肠杆菌的耐热性适应和进化可能是由大肠杆菌群体间的多样化和互作引起的，并且通过进化表型的固定过程观察到基因组DNA含量的增加和G/C含量的减少。

We have researched about the dynamics both of cellular and E. coli population level during adaptation to heat stress (high temperature; above 39 degree). First, we constructed the serially transfer culture system maintaining specific growth rate, then transferred E. coli culture to high temperature (2 degree above past temperature). We could evolve E. coli to accept 45 degree condition that was highest temperature E. coli could survive in minimum conditioned medium. During 39 degree adaptation, just after temperature transition, growth rate decreased rapidly and standard deviation of fluorescence of cellular green fluorescence protein (GFP) of E. coli population enhanced in 2~7days, then growth rate recovered and SD of GFP was decreased for 10-20 days and converged to 39 degree specific values. These results indicated that temperature shift induced diversity of intracellular conditions that might bring the recovery of plasticity of E. coli population, and then adaptation was facilitated by E. coli diversity. At the early stage of 45 degree adaptation, E. coli could grow cell density dependent-manner. This dependency was derived from secreted factors from E. coli, therefore, temperature shift might induce quorum sensing. We have been searching corresponding secreted factors from E. coli culture supernatant. Moreover, average genome DNA content in E. coli cell increased in proportion to adaptation to higher temperature, and interestingly content both of guanine and cytosine decreased according to adaptation to higher temperature. Our results suggested that heat tolerant adaptation and evolution of E. coli might be brought by diversification and/or interaction among E. roll population, and genome DNA content increment and G/C content decrement were observed through fixation process of evolved phenotype.