Targeted knock-down of miR21 primary transcripts using snoMEN vectors induces apoptosis in human cancer cell lines

Motoharu Ono, Kayo Ono, Fabio Avolio, Vackar Afzal, Dalila Bensaddek, Angus I. Lamond (Lead / Corresponding author)

Research output: Contribution to journalArticle

7 Citations (Scopus)
278 Downloads (Pure)

Abstract

We have previously reported an antisense technology, ‘snoMEN vectors’, for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the specific knock-down of miR21 in HeLa cells using plasmid vectors expressing miR21-targeted snoMEN RNAs and show this induces apoptosis. Knock-down is dependent on the presence of complementary sequences in the snoMEN vector and the induction of apoptosis can be suppressed by over-expression of miR21. Furthermore, we have also developed lentiviral vectors for delivery of snoMEN RNAs and show this increases the efficiency of vector transduction in many human cell lines that are difficult to transfect with plasmid vectors. Transduction of lentiviral vectors expressing snoMEN targeted to pri-miR21 induces apoptosis in human lung adenocarcinoma cells, which express high levels of miR21, but not in human primary cells. We show that snoMEN-mediated suppression of miRNA expression is prevented by siRNA knock-down of Ago2, but not by knock-down of Ago1 or Upf1. snoMEN RNAs colocalise with Ago2 in cell nuclei and nucleoli and can be co-immunoprecipitated from nuclear extracts by antibodies specific for Ago2.
Original languageEnglish
Article numbere0138668
Number of pages22
JournalPLoS ONE
Volume10
Issue number9
DOIs
Publication statusPublished - 25 Sep 2015

Fingerprint

plasmid vectors
apoptosis
Cells
cell lines
RNA
Apoptosis
microRNA
Cell Line
MicroRNAs
Neoplasms
Plasmids
human cell lines
Cell Nucleolus
Small Nucleolar RNA
cell nucleolus
cells
cell nucleus
small interfering RNA
adenocarcinoma
Messenger RNA

Cite this

@article{c06678fd32a142a09ec0d13678f67363,
title = "Targeted knock-down of miR21 primary transcripts using snoMEN vectors induces apoptosis in human cancer cell lines",
abstract = "We have previously reported an antisense technology, ‘snoMEN vectors’, for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the specific knock-down of miR21 in HeLa cells using plasmid vectors expressing miR21-targeted snoMEN RNAs and show this induces apoptosis. Knock-down is dependent on the presence of complementary sequences in the snoMEN vector and the induction of apoptosis can be suppressed by over-expression of miR21. Furthermore, we have also developed lentiviral vectors for delivery of snoMEN RNAs and show this increases the efficiency of vector transduction in many human cell lines that are difficult to transfect with plasmid vectors. Transduction of lentiviral vectors expressing snoMEN targeted to pri-miR21 induces apoptosis in human lung adenocarcinoma cells, which express high levels of miR21, but not in human primary cells. We show that snoMEN-mediated suppression of miRNA expression is prevented by siRNA knock-down of Ago2, but not by knock-down of Ago1 or Upf1. snoMEN RNAs colocalise with Ago2 in cell nuclei and nucleoli and can be co-immunoprecipitated from nuclear extracts by antibodies specific for Ago2.",
author = "Motoharu Ono and Kayo Ono and Fabio Avolio and Vackar Afzal and Dalila Bensaddek and Lamond, {Angus I.}",
year = "2015",
month = "9",
day = "25",
doi = "10.1371/journal.pone.0138668",
language = "English",
volume = "10",
journal = "PLoS ONE",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "9",

}

Targeted knock-down of miR21 primary transcripts using snoMEN vectors induces apoptosis in human cancer cell lines. / Ono, Motoharu; Ono, Kayo; Avolio, Fabio; Afzal, Vackar; Bensaddek, Dalila; Lamond, Angus I. (Lead / Corresponding author).

In: PLoS ONE, Vol. 10, No. 9, e0138668, 25.09.2015.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Targeted knock-down of miR21 primary transcripts using snoMEN vectors induces apoptosis in human cancer cell lines

AU - Ono, Motoharu

AU - Ono, Kayo

AU - Avolio, Fabio

AU - Afzal, Vackar

AU - Bensaddek, Dalila

AU - Lamond, Angus I.

PY - 2015/9/25

Y1 - 2015/9/25

N2 - We have previously reported an antisense technology, ‘snoMEN vectors’, for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the specific knock-down of miR21 in HeLa cells using plasmid vectors expressing miR21-targeted snoMEN RNAs and show this induces apoptosis. Knock-down is dependent on the presence of complementary sequences in the snoMEN vector and the induction of apoptosis can be suppressed by over-expression of miR21. Furthermore, we have also developed lentiviral vectors for delivery of snoMEN RNAs and show this increases the efficiency of vector transduction in many human cell lines that are difficult to transfect with plasmid vectors. Transduction of lentiviral vectors expressing snoMEN targeted to pri-miR21 induces apoptosis in human lung adenocarcinoma cells, which express high levels of miR21, but not in human primary cells. We show that snoMEN-mediated suppression of miRNA expression is prevented by siRNA knock-down of Ago2, but not by knock-down of Ago1 or Upf1. snoMEN RNAs colocalise with Ago2 in cell nuclei and nucleoli and can be co-immunoprecipitated from nuclear extracts by antibodies specific for Ago2.

AB - We have previously reported an antisense technology, ‘snoMEN vectors’, for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the specific knock-down of miR21 in HeLa cells using plasmid vectors expressing miR21-targeted snoMEN RNAs and show this induces apoptosis. Knock-down is dependent on the presence of complementary sequences in the snoMEN vector and the induction of apoptosis can be suppressed by over-expression of miR21. Furthermore, we have also developed lentiviral vectors for delivery of snoMEN RNAs and show this increases the efficiency of vector transduction in many human cell lines that are difficult to transfect with plasmid vectors. Transduction of lentiviral vectors expressing snoMEN targeted to pri-miR21 induces apoptosis in human lung adenocarcinoma cells, which express high levels of miR21, but not in human primary cells. We show that snoMEN-mediated suppression of miRNA expression is prevented by siRNA knock-down of Ago2, but not by knock-down of Ago1 or Upf1. snoMEN RNAs colocalise with Ago2 in cell nuclei and nucleoli and can be co-immunoprecipitated from nuclear extracts by antibodies specific for Ago2.

U2 - 10.1371/journal.pone.0138668

DO - 10.1371/journal.pone.0138668

M3 - Article

VL - 10

JO - PLoS ONE

JF - PLoS ONE

SN - 1932-6203

IS - 9

M1 - e0138668

ER -