De Paepe Marianne, Hutinet Geoffrey, Son Olivier, Amarir-Bouhram Jihane, Schbath Sophie, Petit Marie-Agnès
INRA, UMR1319, Micalis, domaine de Vilvert, Jouy en Josas, France; AgroParisTech, UMR1319, Micalis, domaine de Vilvert, Jouy en Josas, France.
INRA, UR1077, MIG, domaine de Vilvert, Jouy en Josas, France.
PLoS Genet. 2014 Mar 6;10(3):e1004181. doi: 10.1371/journal.pgen.1004181. eCollection 2014 Mar.
Bacteriophages (or phages) dominate the biosphere both numerically and in terms of genetic diversity. In particular, genomic comparisons suggest a remarkable level of horizontal gene transfer among temperate phages, favoring a high evolution rate. Molecular mechanisms of this pervasive mosaicism are mostly unknown. One hypothesis is that phage encoded recombinases are key players in these horizontal transfers, thanks to their high efficiency and low fidelity. Here, we associate two complementary in vivo assays and a bioinformatics analysis to address the role of phage encoded recombinases in genomic mosaicism. The first assay allowed determining the genetic determinants of mosaic formation between lambdoid phages and Escherichia coli prophage remnants. In the second assay, recombination was monitored between sequences on phage λ, and allowed to compare the performance of three different Rad52-like recombinases on the same substrate. We also addressed the importance of homologous recombination in phage evolution by a genomic comparison of 84 E. coli virulent and temperate phages or prophages. We demonstrate that mosaics are mainly generated by homology-driven mechanisms that tolerate high substrate divergence. We show that phage encoded Rad52-like recombinases act independently of RecA, and that they are relatively more efficient when the exchanged fragments are divergent. We also show that accessory phage genes orf and rap contribute to mosaicism. A bioinformatics analysis strengthens our experimental results by showing that homologous recombination left traces in temperate phage genomes at the borders of recently exchanged fragments. We found no evidence of exchanges between virulent and temperate phages of E. coli. Altogether, our results demonstrate that Rad52-like recombinases promote gene shuffling among temperate phages, accelerating their evolution. This mechanism may prove to be more general, as other mobile genetic elements such as ICE encode Rad52-like functions, and play an important role in bacterial evolution itself.
噬菌体在数量和遗传多样性方面都在生物圈中占据主导地位。特别是，基因组比较表明，温和噬菌体之间存在显著水平的水平基因转移，这有利于高进化速率。这种普遍存在的镶嵌现象的分子机制大多未知。一种假设是，噬菌体编码的重组酶是这些水平转移的关键参与者，这得益于它们的高效率和低保真性。在这里，我们结合了两种互补的体内试验和生物信息学分析，以研究噬菌体编码的重组酶在基因组镶嵌现象中的作用。第一个试验允许确定类λ噬菌体和大肠杆菌原噬菌体残余物之间镶嵌形成的遗传决定因素。在第二个试验中，监测噬菌体λ上序列之间的重组，并允许比较三种不同的Rad52样重组酶在同一底物上的性能。我们还通过对84种大肠杆菌烈性噬菌体和温和噬菌体或原噬菌体的基因组比较，探讨了同源重组在噬菌体进化中的重要性。我们证明，镶嵌体主要由耐受高底物差异的同源驱动机制产生。我们表明，噬菌体编码的Rad52样重组酶独立于RecA起作用，并且当交换的片段存在差异时，它们相对更有效。我们还表明，辅助噬菌体基因orf和rap有助于镶嵌现象。生物信息学分析通过表明同源重组在最近交换片段的边界处的温和噬菌体基因组中留下痕迹，加强了我们的实验结果。我们没有发现大肠杆菌烈性噬菌体和温和噬菌体之间发生交换的证据。总之，我们的结果表明，Rad52样重组酶促进了温和噬菌体之间的基因改组，加速了它们的进化。这种机制可能更具普遍性，因为其他移动遗传元件，如整合性接合元件（ICE）也编码Rad52样功能，并在细菌进化本身中发挥重要作用。
