Non-autonomous VNTR (Variable Number of Tandem Repeats) composite retrotransposons – SVA (SINE-R-VNTR-Alu) and LAVA (L1-Alu-VNTR-Alu) – are specific to hominoid primates. SVA expanded in great apes, LAVA in gibbon. Both SVA and LAVA have been shown to be mobilized by the autonomous LINE-1 (L1)-encoded protein machinery in a cell-based assay in trans. The efficiency of human SVA retrotransposition in vitro has, however, been considerably lower than would be expected based on recent pedigree-based in vivo estimates. The VNTR composite elements across hominoids – gibbon LAVA, orangutan SVA_A descendants and hominine SVA_D descendants – display characteristic structures of the 5′ Alu-like domain and the VNTR. Different partner L1 subfamilies are currently active in each of the lineages. The possibility that the lineage-specific types of VNTR composites evolved in response to evolutionary changes in their autonomous partners, particularly in the nucleic acid binding L1 ORF1-encoded protein, has not been addressed. Here I report the identification and functional characterization of a highly active human SVA element using an improved mneo retrotransposition reporter cassette. The modified cassette (mneoM) minimizes splicing between the VNTR of human SVAs and the neomycin phosphotransferase stop codon. SVA deletion analysis provides evidence that key elements determining its mobilization efficiency reside in the VNTR and 5′ hexameric repeats. Simultaneous removal of the 5′ hexameric repeats and part of the VNTR has an additive negative effect on mobilization rates. Taking advantage of the modified reporter cassette that facilitates robust cross-species comparison of SVA/LAVA retrotransposition, I show that the ORF1-encoded proteins of the L1 subfamilies currently active in gibbon, orangutan and human do not display substrate preference for gibbon LAVA versus orangutan SVA versus human SVA. Finally, I demonstrate that an orangutan-derived ORF1p supports only limited retrotransposition of SVA/LAVA in trans, despite being fully functional in L1 mobilization in cis. Overall, the analysis confirms SVA as a highly active human retrotransposon and preferred substrate of the L1-encoded protein machinery. Based on the results obtained in human cells coevolution of L1 ORF1p and VNTR composites does not appear very likely. The changes in orangutan L1 ORF1p that markedly reduce its mobilization capacity in trans might explain the different SVA insertion rates in the orangutan and hominine lineages, respectively.

中文翻译：

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LINE-1 ORF1p 不能确定人类/猩猩 SVA 和长臂猿熔岩的底物偏好。

非自主 VNTR（可变数量串联重复）复合反转录转座子 - SVA (SINE-R-VNTR-Alu) 和 LAVA (L1-Alu-VNTR-Alu) - 是类人灵长类动物特有的。SVA 在类人猿中扩展，LAVA 在长臂猿中扩展。在基于细胞的反式分析中，SVA 和 LAVA 都被证明是由自主的 LINE-1 (L1) 编码的蛋白质机制动员的。然而，体外人类 SVA 逆转录转座的效率远低于基于最近基于谱系的体内估计的预期。跨类人猿的 VNTR 复合元素——长臂猿 LAVA、猩猩 SVA_A 后代和人猿 SVA_D 后代——显示出 5' Alu 样域和 VNTR 的特征结构。不同的伙伴 L1 亚家族目前在每个谱系中都很活跃。谱系特异性类型的 VNTR 复合物随着其自主伙伴的进化变化而进化的可能性，特别是在核酸结合 L1 ORF1 编码蛋白中的可能性尚未得到解决。在这里，我报告了使用改进的 mneo 逆转录转座报告盒对高活性人类 SVA 元件的鉴定和功能表征。修改后的盒式磁带 (mneoM) 最大限度地减少了人类 SVA 的 VNTR 和新霉素磷酸转移酶终止密码子之间的剪接。SVA 缺失分析提供了决定其动员效率的关键因素存在于 VNTR 和 5' 六聚体重复序列中的证据。同时去除 5' 六聚体重复和部分 VNTR 对动员率具有附加的负面影响。利用有助于 SVA/LAVA 逆转录转座的稳健跨物种比较的改良报告盒，我表明目前在长臂猿、猩猩和人类中活跃的 L1 亚科的 ORF1 编码蛋白不显示对长臂猿熔岩与猩猩的底物偏好SVA 与人类 SVA。最后，我证明了猩猩衍生的 ORF1p 仅支持有限的反式 SVA/LAVA 逆转录转座，尽管在顺式 L1 动员中具有完全功能。总体而言，该分析证实 SVA 是一种高活性的人类反转录转座子，也是 L1 编码蛋白质机器的首选底物。基于在人类细胞中获得的结果，L1 ORF1p 和 VNTR 复合材料的共同进化似乎不太可能。