Institut Pasteurgrid.428999.7, Université de Paris, Trypanosome Molecular Biology, Department of Parasites and Insect Vectors, Paris, France.
Université de Paris, Sorbonne Paris Cité, Paris, France.
mBio. 2022 Apr 26;13(2):e0384721. doi: 10.1128/mbio.03847-21. Epub 2022 Mar 1.
In the mammalian host, Trypanosoma brucei is coated in a single-variant surface glycoprotein (VSG) species. Stochastic switching of the expressed allows the parasite to escape detection by the host immune system. DNA double-strand breaks (DSB) trigger VSG switching, and repair via gene conversion results in an antigenically distinct being expressed from the single active bloodstream-form expression site (BES). The single active BES is marked by exclusion 2 (VEX2) protein. Here, we have disrupted monoallelic expression by stably expressing a second telomeric from a ribosomal locus. We found that cells expressing two contained one VEX2 focus that was significantly larger in size than the wild-type cells; this therefore suggests the ectopic is expressed from the same nuclear position as the active BES. Unexpectedly, we report that in the double -expressing cells, the DNA sequence of the ectopic copy is lost following a DSB in the active BES, despite it being spatially separated in the genome. The loss of the ectopic is dependent on active transcription and does not disrupt the number or variety of templates used to repair a BES DSB and elicit a VSG switch. We propose that there are stringent mechanisms within the cell to reinforce monoallelic expression during antigenic variation. The single-cell parasite Trypanosoma brucei causes the fatal disease human African trypanosomiasis and is able to colonize the blood, fat, skin, and central nervous system. Trypanosomes survive in the mammalian host owing to a dense protective protein coat that consists of a single-variant surface glycoprotein species. Stochastic switching of one VSG for an immunologically distinct one enables the parasite to escape recognition by the host immune system. We have disrupted monoallelic antigen expression by expressing a second and report that following DSB-triggered VSG switching, the DNA sequence of the ectopic is lost in a transcription-dependent manner. We propose that there are strict requirements to ensure that only one variant antigen is expressed following a VSG switch, which has important implications for understanding how the parasite survives in the mammalian host.
在哺乳动物宿主中，布氏锥虫被一层单一变异的表面糖蛋白（VSG）所覆盖。表达的随机切换允许寄生虫逃避宿主免疫系统的检测。DNA 双链断裂（DSB）触发 VSG 切换，通过基因转换修复导致从单个活性血流形式表达位点（BES）表达具有不同抗原性的 。单个活性 BES 由 排除 2（VEX2）蛋白标记。在这里，我们通过稳定表达第二个端粒 从核糖体基因座来破坏单等位基因 表达。我们发现表达两个 的细胞包含一个 VEX2 焦点，其大小明显大于野生型细胞；因此，这表明异位 是从与活性 BES 相同的核位置表达的。出乎意料的是，我们报告说，在双表达细胞中，尽管在基因组中空间分离，但在活性 BES 中的 DSB 后，异位 序列丢失。尽管异位 的丢失不依赖于活性转录，但它不破坏用于修复 BES DSB 并引发 VSG 切换的模板的数量或种类。我们提出，在细胞内存在严格的机制来加强抗原变异期间的单等位基因表达。单细胞寄生虫布氏锥虫引起致命的人类非洲锥虫病，并且能够定殖血液、脂肪、皮肤和中枢神经系统。锥虫在哺乳动物宿主中存活是由于其密集的保护性蛋白质外壳，该外壳由单一变异的表面糖蛋白组成。一种 VSG 免疫上独特的随机切换使寄生虫能够逃避宿主免疫系统的识别。我们通过表达第二个 来破坏单等位基因抗原表达，并报告说，在 DSB 触发的 VSG 切换后，异位 的 DNA 序列以转录依赖性方式丢失。我们提出，有严格的要求来确保在 VSG 切换后仅表达一种变体抗原，这对于理解寄生虫如何在哺乳动物宿主中存活具有重要意义。
