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种群水平在适应热应激（高温；39度以上）过程中的动态变化。首先，我们构建了保持特定生长速率的连续转移培养系统，然后将大肠杆菌培养物转移至高温（比过去温度高2度）。我们可以进化大肠杆菌接受 45 度的条件，这是大肠杆菌可以在最低限度的条件培养基中生存的最高温度。 39度适应过程中，温度刚转变后，生长速率迅速下降，2~7天后大肠杆菌群体细胞绿色荧光蛋白（GFP）荧光标准差增强，随后10~20天生长速率恢复，GFP的SD下降并收敛于39度特异值。这些结果表明，温度变化诱导了细胞内条件的多样性，这可能会带来大肠杆菌种群可塑性的恢复，然后大肠杆菌多样性促进了适应。在45度适应的早期阶段，大肠杆菌可以以细胞密度依赖性方式生长。这种依赖性源自大肠杆菌的分泌因子，因此，温度变化可能会诱导群体感应。我们一直在从大肠杆菌培养上清液中寻找相应的分泌因子。此外，大肠杆菌细胞中的平均基因组DNA含量随着对较高温度的适应而成比例增加，有趣的是，鸟嘌呤和胞嘧啶的含量随着对较高温度的适应而减少。我们的结果表明，大肠杆菌的耐热适应和进化可能是由大肠杆菌种群之间的多样化和/或相互作用带来的，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.