Galetto Román, Moumen Abdeladim, Giacomoni Véronique, Véron Michel, Charneau Pierre, Negroni Matteo
Unité de Régulation Enzymatique des Activités Cellulaires CNRS-URA 2185 and Groupe de Virologie Moléculaire et Vectorologie, Institut Pasteur, 25-28 rue du Dr. Roux, 75724 Paris, France.
J Biol Chem. 2004 Aug 27;279(35):36625-32. doi: 10.1074/jbc.M405476200. Epub 2004 Jun 24.
By frequently rearranging large regions of the genome, genetic recombination is a major determinant in the plasticity of the human immunodeficiency virus type I (HIV-1) population. In retroviruses, recombination mostly occurs by template switching during reverse transcription. The generation of retroviral vectors provides a means to study this process after a single cycle of infection of cells in culture. Using HIV-1-derived vectors, we present here the first characterization and estimate of the strength of a recombination hot spot in HIV-1 in vivo. In the hot spot region, located within the C2 portion of the gp120 envelope gene, the rate of recombination is up to ten times higher than in the surrounding regions. The hot region corresponds to a previously identified RNA hairpin structure. Although recombination breakpoints in vivo cluster in the top portion of the hairpin, the bias for template switching in this same region appears less marked in a cell-free system. By modulating the stability of this hairpin we were able to affect the local recombination rate both in vitro and in infected cells, indicating that the local folding of the genomic RNA is a major parameter in the recombination process. This characterization of reverse transcription products generated after a single cycle of infection provides insights in the understanding of the mechanism of recombination in vivo and suggests that specific regions of the genome might be prompted to yield different rates of evolution due to the presence of circumscribed recombination hot spots.
通过频繁重排基因组的大片段，基因重组是人类免疫缺陷病毒I型（HIV-1）群体可塑性的主要决定因素。在逆转录病毒中，重组大多在逆转录过程中通过模板转换发生。逆转录病毒载体的产生提供了一种在培养细胞单次感染周期后研究这一过程的方法。利用源自HIV-1的载体，我们在此首次对HIV-1体内重组热点的强度进行了表征和估计。在位于gp120包膜基因C2部分的热点区域，重组率比周围区域高出多达10倍。该热点区域对应于先前鉴定出的一种RNA发夹结构。虽然体内重组断点聚集在发夹的顶部，但在无细胞系统中，同一区域模板转换的偏向似乎不那么明显。通过调节这种发夹的稳定性，我们能够在体外和感染细胞中影响局部重组率，这表明基因组RNA的局部折叠是重组过程中的一个主要参数。对单次感染周期后产生的逆转录产物的这一表征为理解体内重组机制提供了见解，并表明由于存在限定的重组热点，基因组的特定区域可能会以不同的速率进化。
