@misc{10481/88582,
year = {2020},
url = {https://hdl.handle.net/10481/88582},
abstract = {The cell culture-based retrotransposition reporter assay has been (and is) an
essential tool for the study of vertebrate Long INterspersed Elements
(LINEs). Developed more than 20 years ago, this assay has been instrumental
in characterizing the role of LINE-encoded proteins in retrotransposition,
understanding how ribonucleoprotein particles are formed, how host factors
regulate LINE mobilization, etc. Moreover, variations of the conventional
assay have been developed to investigate the biology of other currently
active human retrotransposons, such as Alu and SVA. Here,we describe a protocol
that allows combination of the conventional cell culture-based LINE-1
retrotransposition reporter assay with short interfering RNAs (siRNAs) and
microRNA (miRNAs) mimics or inhibitors, which has allowed us to uncover
specific miRNAs and host factors that regulate retrotransposition. The protocol
described here is highly reproducible, quantitative, robust and flexible, and
allows the study of several small RNA classes and various retrotransposons.
To illustrate its utility, here we show that siRNAs to Fanconi anaemia proteins
(FANC-A and FANC-C) and an inhibitor of miRNA-20 upregulate and
downregulate human L1 retrotransposition, respectively.},
organization = {J.L.G.-P. acknowledges funding from the European Research
Council (ERC-Consolidator ERC-STG-2012-309433), MINECO-FEDER
(SAF2017-89745-R), the Howard Hughes Medical Institute (IECS-
55007420), the Wellcome Trust-University of Edinburgh Institutional
Strategic Support Fund (ISFF2) and a private donation to his laboratory
from Ms Francisca Serrano (Trading y Bolsa para Torpes,
Granada, Spain). S.M. was supported by the Government of Andalucia
(FEDER-P08-CTS-03678). S.R.H. and P.T.-R. are funded by the Ramon
y Cajal programme (RYC-2016-21395) and by MINECO-FEDER
(SAF2015-71589-P). This paper is part of the thesis project of P.T.-R.,
within the Biomedicine Program of the University of Granada, Spain.},
publisher = {Royal Society},
title = {sRNA/L1 retrotransposition: using siRNAs and miRNAs to expand the applications of the cell culture-based LINE-1 retrotransposition assay},
doi = {10.1098/rstb.2019.0346.},
author = {Tristán Ramos, Pablo and Morell, Santiago and Toledo, Belén and García Pérez, José Luis and Rodríguez Heras, Sara},
}