TY - JOUR
T1 - Whole-genome-scale identification of novel non-protein-coding RNAs controlling cell proliferation and survival through a functional forward genetics strategy
AU - Tonge, D. P.
AU - Darling, D.
AU - Farzaneh, F.
AU - Williams, G. T.
N1 - Funding Information:
Work in the Molecular Medicine Group at King’s is supported by the Wellcome Trust, Medical Research Council (MRC), LifeArc and Biochemical Biomedical Research Council (BBSRC), as well as the Experimental Cancer Medicine Centre at King’s College London, the Cancer Research UK Centre at King’s Health Partners and by the National Institute for Health Research (NIHR) Biomedical Research Centre based at Guy’s and St Thomas’ NHS Foundation Trust and King’s College London.
Publisher Copyright:
© 2022, The Author(s).
PY - 2022/12
Y1 - 2022/12
N2 - Identification of cell fate-controlling lncRNAs is essential to our understanding of molecular cell biology. Here we present a human genome-scale forward-genetics approach for the identification of lncRNAs based on gene function. This approach can identify genes that play a causal role, and immediately distinguish them from those that are differentially expressed but do not affect cell function. Our genome-scale library plus next-generation-sequencing and bioinformatic approach, radically upscales the breadth and rate of functional ncRNA discovery. Human gDNA was digested to produce a lentiviral expression library containing inserts in both sense and anti-sense orientation. The library was used to transduce human Jurkat T-leukaemic cells. Cell populations were selected using continuous culture ± anti-FAS IgM, and sequencing used to identify sequences controlling cell proliferation. This strategy resulted in the identification of thousands of new sequences based solely on their function including many ncRNAs previously identified as being able to modulate cell survival or to act as key cancer regulators such as AC084816.1*, AC097103.2, AC087473.1, CASC15*, DLEU1*, ENTPD1-AS1*, HULC*, MIRLET7BHG*, PCAT-1, SChLAP1, and TP53TG1. Independent validation confirmed 4 out of 5 sequences that were identified by this strategy, conferred a striking resistance to anti-FAS IgM-induced apoptosis.
AB - Identification of cell fate-controlling lncRNAs is essential to our understanding of molecular cell biology. Here we present a human genome-scale forward-genetics approach for the identification of lncRNAs based on gene function. This approach can identify genes that play a causal role, and immediately distinguish them from those that are differentially expressed but do not affect cell function. Our genome-scale library plus next-generation-sequencing and bioinformatic approach, radically upscales the breadth and rate of functional ncRNA discovery. Human gDNA was digested to produce a lentiviral expression library containing inserts in both sense and anti-sense orientation. The library was used to transduce human Jurkat T-leukaemic cells. Cell populations were selected using continuous culture ± anti-FAS IgM, and sequencing used to identify sequences controlling cell proliferation. This strategy resulted in the identification of thousands of new sequences based solely on their function including many ncRNAs previously identified as being able to modulate cell survival or to act as key cancer regulators such as AC084816.1*, AC097103.2, AC087473.1, CASC15*, DLEU1*, ENTPD1-AS1*, HULC*, MIRLET7BHG*, PCAT-1, SChLAP1, and TP53TG1. Independent validation confirmed 4 out of 5 sequences that were identified by this strategy, conferred a striking resistance to anti-FAS IgM-induced apoptosis.
UR - http://www.scopus.com/inward/record.url?scp=85122430097&partnerID=8YFLogxK
U2 - 10.1038/s41598-021-03983-5
DO - 10.1038/s41598-021-03983-5
M3 - Article
AN - SCOPUS:85122430097
SN - 2045-2322
VL - 12
JO - Scientific Reports
JF - Scientific Reports
IS - 1
M1 - 182
ER -
