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，一个新兴的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，它们增加了宿主基因组固有的流动性，特别是当它们大量存在时。由于细胞内细菌基本上是无性的，丰富的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的IS元件的进化动力学的重要性。该项目的目的是测试以下两个假设，为什么IS元件经常在细胞内共生体中扩增：（1）细胞内细菌具有相对小的群体大小，因此自然选择在清除有害的遗传变化方面不太有效，因为随机遗传漂变在这些群体中起着越来越重要的作用，和（2）细胞内共生体通常沐浴在营养丰富的环境中，并且不必合成许多它们自身所需的代谢物，因此许多生物合成基因可能对于IS元件插入而言基本上是选择性中性区域。这些假设将使用实验进化方法与E。杆菌简而言之，复制E。大肠杆菌群体（其具有44个常驻IS）将经历四种实验处理5000代：（1）大群体大小/丰富培养基，（2）小群体大小/丰富培养基，（3）大群体大小/最小培养基，和（4）小群体大小/最小培养基。如果IS元素在小群体中的传播比在大群体中的传播更显著，那么这将支持这样的假设，即当自然选择由于小群体中明显的遗传漂变而放松时，IS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小培养基群体中传播得更明显，那么这将支持对多余的生物合成基因进行放松选择允许IS增殖的假设。这两个假设是不相互排斥的，都可能是重要的，在塑造IS扩张细胞内共生体。如果是这样，则IS扩展将与每种介质类型的群体大小负相关，最小介质值低于丰富介质值。 项目叙述：IS元素为其宿主的基因组增加了固有的流动性（例如，帕克希尔等人，2003年;宋等人，2004年;杨等人，2005年）。因此，它们在细胞内病原体中的频繁增殖可能在很少遇到其他细菌并与其他细菌交换DNA的细菌中产生遗传变异性方面具有根本重要性。这强调了阐明细胞内病原体的IS元件的进化动力学的重要性。

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.