Experimental evolution of insertion sequence expansion in intracellular bacteria

描述（由申请人提供）：随着细胞内细菌共生体和病原体的完整基因组序列的积累，一个新趋势是，转座插入序列（is）元件通常在细菌从自由生活方式转变为细胞内生活方式后增殖。IS元素通常是有害的。然而，它们为宿主的基因组增加了固有的流动性，尤其是当它们数量丰富时。由于细胞内细菌本质上是无性的，大量的IS元素可能是这些群体遗传变异的重要来源。这强调了阐明细胞内共生体和病原体中IS元素的进化动力学的重要性。这个项目的目的是测试以下两个假设，为什么is元素经常在细胞内共生体中扩展：(1)细胞内细菌的群体规模相对较小，因此自然选择在清除有害遗传变化方面的效果较差，因为随机遗传漂变在这些群体中起着越来越重要的作用；(2)细胞内共生体经常处于营养丰富的环境中，不需要合成许多自身所需的代谢物，因此许多生物合成基因可能本质上是is元件插入的选择性中性区域。这些假设将使用大肠杆菌的实验进化方法进行验证。简而言之，重复大肠杆菌群体（有44个常驻ISs）将接受4种实验处理，共5000代：(1)大群体/富培养基，(2)小群体/富培养基，(3)大群体/最小培养基，(4)小群体/最小培养基。如果IS元素在小群体中比在大群体中传播得更多，那么这将支持这样的假设，即当自然选择因小群体中明显的遗传漂变而放松时，ISs会扩大。或者，如果IS元素在营养丰富的培养基群体中比在最低限度的培养基群体中传播得更多，那么这将支持对多余生物合成基因的宽松选择允许ISs增殖的假设。这两种假设并不是相互排斥的，它们都可能在细胞内共生体中形成IS的扩展中发挥重要作用。如果是这样，那么IS的扩展将与每种媒体类型的人口规模负相关，最小媒体值低于富媒体值。

DESCRIPTION (provided by applicant): As complete genome sequences of intracellular bacterial symbionts and pathogens accumulate, an emerging trend is that transposable insertion sequence (IS) elements often proliferate after bacteria transition from a free-living to an intracellular lifestyle. IS elements are generally detrimental. However, they add inherent fluidity to their hosts' genomes, particularly when they are abundant. Because intracellular bacteria are essentially asexual, profuse IS elements may be an important source of genetic variation for these populations. This underscores the importance of elucidating the evolutionary dynamics of IS elements within intracellular symbionts and pathogens. The objective of this project is to test the following two hypotheses for why IS elements often expand in intracellular symbionts: (1) intracellular bacteria have relatively small population sizes, so natural selection is less effective at purging deleterious genetic changes because stochastic genetic drift plays an increasingly important role in these populations, and (2) intracellular symbionts are often bathed in a nutrient-rich environment and do not have to synthesize many of their own required metabolites, so many biosynthetic genes may be essentially selectively neutral territory for IS element insertion. These hypotheses will be tested using an experimental evolution approach with E. coli. In short, replicate E. coli populations (which have 44 resident ISs) will be subjected to four experimental treatments for 5000 generations: (1) large population size/rich medium, (2) small population size/rich medium, (3) large population size/minimal medium, and (4) small population size/minimal medium. If IS elements spread significantly more in small populations than large populations, then this will support the hypothesis that ISs expand when natural selection is relaxed due to pronounced genetic drift in small populations. Alternatively, if IS elements spread significantly more in nutritionally rich medium populations than minimal medium populations, then this will support the hypothesis that relaxed selection on superfluous biosynthetic genes allows ISs to proliferate. These two hypotheses are not mutually exclusive and both could be important in shaping IS expansion in intracellular symbionts. If so, then IS expansion will negatively correlate to population size for each media type, with the minimal media values lower than the rich media values. Project Narrative: IS elements add inherent fluidity to their hosts' genomes (e.g., Parkhill et al. 2003; Song et al. 2004; Yang et al. 2005). Therefore, their frequent proliferation in intracellular pathogens may be fundamentally important in generating genetic variability in bacteria that rarely encounter and exchange DNA with other bacteria. This underscores the importance of elucidating the evolutionary dynamics of IS elements within intracellular pathogens.