Experimental evolution of insertion sequence expansion in intracellular bacteria

描述(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，一个新的趋势是，在细菌从自由生活方式转变为细胞内生活方式后，转座插入序列(IS)元件通常会增殖。IS元素通常是有害的。然而，它们为寄主的基因组增加了固有的流动性，特别是当它们丰富的时候。由于胞内细菌本质上是无性的，丰富的IS元素可能是这些种群遗传变异的重要来源。这突显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。这个项目的目的是检验以下两个假设，解释为什么元素经常在细胞内共生体中扩展：(1)细胞内细菌的种群规模相对较小，因此自然选择在清除有害遗传变化方面的效率较低，因为随机遗传漂移在这些种群中发挥着越来越重要的作用；(2)细胞内共生体通常沐浴在营养丰富的环境中，不必合成它们自己所需的许多代谢产物，因此许多生物合成基因可能基本上是选择性的中性区域，用于IS元素的插入。这些假说将用一种实验性的进化方法在大肠杆菌中进行验证。简而言之，复制的大肠杆菌种群(有44个常驻ISS)将在5000代中经历四种实验处理：(1)大种群规模/丰富介质，(2)小种群规模/丰富介质，(3)大种群规模/最小介质，和(4)小种群规模/最小介质。如果IS元素在小种群中的分布明显多于大种群，那么这将支持这样的假设，即由于小种群中明显的遗传漂移，当自然选择放松时，ISS会扩大。或者，如果IS元素在营养丰富的中等种群中的分布明显多于最小中等种群，那么这将支持这样的假设，即对多余的生物合成基因的宽松选择允许ISS增殖。这两个假设并不是相互排斥的，这两个假设在塑造细胞内共生体的IS扩张方面都可能是重要的。如果是这样，则IS扩展将与每种媒体类型的人口大小负相关，最小媒体值低于富媒体值。 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，ParkHill等人。2003年；Song et al.2004年；Yang et al.2005)。因此，它们在细胞内病原体中的频繁增殖可能是产生细菌遗传变异性的根本原因，这些细菌很少与其他细菌相遇并交换DNA。这突显了阐明细胞内病原体内IS成分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.