Functional Screening Identifies miRNAs Influencing Apoptosis and Proliferation in Colorectal Cancer
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Functional Screening Identifies miRNAs Influencing Apoptosis and Proliferation in Colorectal Cancer. / Christensen, Lise Lotte; Holm, Anja; Rantala, Juha; Kallioniemi, Olli; Rasmussen, Mads H; Ostenfeld, Marie S; Dagnaes-Hansen, Frederik; Oster, Bodil; Schepeler, Troels; Tobiasen, Heidi; Thorsen, Kasper; Sieber, Oliver M; Gibbs, Peter; Lamy, Philippe; Hansen, Torben F; Jakobsen, Anders; Riising, Eva M; Helin, Kristian; Lubinski, Jan; Hagemann-Madsen, Rikke; Laurberg, Søren; Orntoft, Torben F; Andersen, Claus L.
In: PLOS ONE, Vol. 9, No. 6, 2014, p. e96767.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Functional Screening Identifies miRNAs Influencing Apoptosis and Proliferation in Colorectal Cancer
AU - Christensen, Lise Lotte
AU - Holm, Anja
AU - Rantala, Juha
AU - Kallioniemi, Olli
AU - Rasmussen, Mads H
AU - Ostenfeld, Marie S
AU - Dagnaes-Hansen, Frederik
AU - Oster, Bodil
AU - Schepeler, Troels
AU - Tobiasen, Heidi
AU - Thorsen, Kasper
AU - Sieber, Oliver M
AU - Gibbs, Peter
AU - Lamy, Philippe
AU - Hansen, Torben F
AU - Jakobsen, Anders
AU - Riising, Eva M
AU - Helin, Kristian
AU - Lubinski, Jan
AU - Hagemann-Madsen, Rikke
AU - Laurberg, Søren
AU - Orntoft, Torben F
AU - Andersen, Claus L
PY - 2014
Y1 - 2014
N2 - MicroRNAs (miRNAs) play a critical role in many biological processes and are aberrantly expressed in human cancers. Particular miRNAs function either as tumor suppressors or oncogenes and appear to have diagnostic and prognostic significance. Although numerous miRNAs are dys-regulated in colorectal cancer (CRC) only a small fraction has been characterized functionally. Using high-throughput functional screening and miRNA profiling of clinical samples the present study aims at identifying miRNAs important for the control of cellular growth and/or apoptosis in CRC. The high-throughput functional screening was carried out in six CRC cell lines transfected with a pre-miR library including 319 synthetic human pre-miRs. Phenotypic alterations were evaluated by immunostaining of cleaved cPARP (apoptosis) or MKI67 (proliferation). Additionally, TaqMan Human MicroRNA Array Set v2.0 was used to profile the expression of 667 miRNAs in 14 normal colon mucosa and 46 microsatellite stable stage II CRC patients. Among the miRNAs that induced growth arrest and apoptosis in the CRC cell lines, and at same time were dys-regulated in the clinical samples, miR-375 was selected for further analysis. Independent in vitro analysis of transient and stable transfected CRC cell lines confirmed that miR-375 reduces cell viability through the induction of apoptotic death. We identified YAP1 as a direct miR-375 target in CRC and show that HELLS and NOLC1 are down-stream targets. Knock-down of YAP1 mimicked the phenotype induced by miR-375 over-expression indicating that miR-375 most likely exerts its pro-apoptotic role through YAP1 and its anti-apoptotic down-stream targets BIRC5 and BCL2L1. Finally, in vivo analysis of mouse xenograft tumors showed that miR-375 expression significantly reduced tumor growth. We conclude that the high-throughput screening successfully identified miRNAs that induce apoptosis and/or inhibit proliferation in CRC cells. Finally, combining the functional screening with profiling of CRC tissue samples we identified clinically relevant miRNAs and miRNA targets in CRC.
AB - MicroRNAs (miRNAs) play a critical role in many biological processes and are aberrantly expressed in human cancers. Particular miRNAs function either as tumor suppressors or oncogenes and appear to have diagnostic and prognostic significance. Although numerous miRNAs are dys-regulated in colorectal cancer (CRC) only a small fraction has been characterized functionally. Using high-throughput functional screening and miRNA profiling of clinical samples the present study aims at identifying miRNAs important for the control of cellular growth and/or apoptosis in CRC. The high-throughput functional screening was carried out in six CRC cell lines transfected with a pre-miR library including 319 synthetic human pre-miRs. Phenotypic alterations were evaluated by immunostaining of cleaved cPARP (apoptosis) or MKI67 (proliferation). Additionally, TaqMan Human MicroRNA Array Set v2.0 was used to profile the expression of 667 miRNAs in 14 normal colon mucosa and 46 microsatellite stable stage II CRC patients. Among the miRNAs that induced growth arrest and apoptosis in the CRC cell lines, and at same time were dys-regulated in the clinical samples, miR-375 was selected for further analysis. Independent in vitro analysis of transient and stable transfected CRC cell lines confirmed that miR-375 reduces cell viability through the induction of apoptotic death. We identified YAP1 as a direct miR-375 target in CRC and show that HELLS and NOLC1 are down-stream targets. Knock-down of YAP1 mimicked the phenotype induced by miR-375 over-expression indicating that miR-375 most likely exerts its pro-apoptotic role through YAP1 and its anti-apoptotic down-stream targets BIRC5 and BCL2L1. Finally, in vivo analysis of mouse xenograft tumors showed that miR-375 expression significantly reduced tumor growth. We conclude that the high-throughput screening successfully identified miRNAs that induce apoptosis and/or inhibit proliferation in CRC cells. Finally, combining the functional screening with profiling of CRC tissue samples we identified clinically relevant miRNAs and miRNA targets in CRC.
U2 - 10.1371/journal.pone.0096767
DO - 10.1371/journal.pone.0096767
M3 - Journal article
C2 - 24892549
VL - 9
SP - e96767
JO - PLoS ONE
JF - PLoS ONE
SN - 1932-6203
IS - 6
ER -
ID: 113597077