Di Stefano Luciano H, Saba Leila J, Oghbaie Mehrnoosh, Jiang Hua, McKerrow Wilson, Benitez-Guijarro Maria, Taylor Martin S, LaCava John
European Research Institute for the Biology of Ageing, University Medical Center Groningen, Groningen, Netherlands.
Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY, USA.
Methods Mol Biol. 2023;2607:215-256. doi: 10.1007/978-1-0716-2883-6_12.
During their proliferation and the host's concomitant attempts to suppress it, LINE-1 (L1) retrotransposons give rise to a collection of heterogeneous ribonucleoproteins (RNPs); their protein and RNA compositions remain poorly defined. The constituents of L1-associated macromolecules can differ depending on numerous factors, including, for example, position within the L1 life cycle, whether the macromolecule is productive or under suppression, and the cell type within which the proliferation is occurring. This chapter describes techniques that aid the capture and characterization of protein and RNA components of L1 macromolecules from tissues that natively express them. The protocols described have been applied to embryonal carcinoma cell lines that are popular model systems for L1 molecular biology (e.g., N2102Ep, NTERA-2, and PA-1 cells), as well as colorectal cancer tissues. N2102Ep cells are given as the use case for this chapter; the protocols should be applicable to essentially any tissue exhibiting endogenous L1 expression with minor modifications.
在LINE-1（L1）逆转录转座子增殖以及宿主随之而来的抑制其增殖的过程中，会产生一系列异质性核糖核蛋白（RNP）；其蛋白质和RNA组成仍不清楚。与L1相关的大分子的成分可能因多种因素而有所不同，例如，在L1生命周期中的位置、该大分子是处于活跃状态还是受到抑制，以及发生增殖的细胞类型。本章介绍了一些技术，这些技术有助于从天然表达L1大分子的组织中捕获和鉴定其蛋白质和RNA成分。所描述的方案已应用于胚胎癌细胞系，这些细胞系是L1分子生物学常用的模型系统（如N2102Ep、NTERA-2和PA-1细胞），以及结直肠癌组织。本章以N2102Ep细胞为例；这些方案经过微小修改后基本上适用于任何表现出内源性L1表达的组织。
