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在培养的哺乳动物细胞中高频逆转换

J V Moran¹, S E Holmes, T P Naas

¹Department of Genetics, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.

|November 29, 1996

概括

人类L1元素L1.2和LRE2在培养的哺乳动物细胞中积极逆转移,产生与疾病相关的插入. 在L1.2蛋白中的突变显著降低了这种逆转换活性.

科学领域:

分子生物学分子生物学
遗传学是一个遗传学.
细胞生物学细胞生物学

背景情况:

人类LINE-1 (L1) 元素是涉及遗传疾病的逆转移子.
之前的研究发现L1.2和LRE2是潜在的致病性L1元素.

研究的目的:

在培养的哺乳动物细胞中研究人类L1元素L1.2和LRE2的逆转移活性.
描述L1逆转换产物的性质,并确定涉及的关键蛋白质域.

主要方法:

稳定表达的L1.2和LRE2元素从一个Episome在HeLa细胞.
对逆转移插入部位的分析,包括5'截断,多元A通道,以及目标部位的重复/删除.
在L1.2编码蛋白质的保存域中引入点突变,以评估它们对逆转换的影响.

主要成果:

无论是L1.2和LRE2元素都显示出高频逆转换到各种染色体位置.
逆转移L1产品表现出类似于内源L1插入的特征.
在关键的L1.2蛋白域中的点突变大大减少了逆转换 (100至1000倍).
在小鼠细胞系中,L1.2也有效地逆转移.

结论:

人类L1元素L1.2和LRE2能够在哺乳动物细胞中进行活性逆转换.
在L1.2内的特定蛋白质域对于其逆转换功能至关重要.
L1.2在物种之间进行逆转移的能力表明,L1基载体在随机插入性突变发生过程中的潜在应用.

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