MicroRNA-18a Attenuates DNA Damage Repair through Suppressing the Expression of Ataxia Telangiectasia Mutated in Colorectal Cancer

Abstract

BackgroundmiR-18a is one of the most up-regulated miRNAs in colorectal cancers (CRC) based on miRNA profiling. In this study, we examined the functional significance of miR-18a in CRC.MethodsExpression of miR-18a was investigated in 45 CRC patients. Potential target genes of miR-18a were predicted by in silico search and confirmed by luciferase activity assay and Western blot. DNA damage was measured by comet assay. Gene function was measured by cell viability, colony formation and apoptosis assays.ResultsThe up-regulation of miR-18a was validated and confirmed in 45 primary CRC tumors compared with adjacent normal tissues (p<0.0001). Through in silico search, the 3′UTR of Ataxia telangiectasia mutated (ATM) contains a conserved miR-18a binding site. Expression of ATM was down-regulated in CRC tumors (p<0.0001) and inversely correlated with miR-18a expression (r = -0.4562, p<0.01). Over-expression of miR-18a in colon cancer cells significantly reduced the luciferase activity of the construct with wild-type ATM 3′UTR but not that with mutant ATM 3′UTR, inferring a direct interaction of miR-18a with ATM 3′UTR. This was further confirmed by the down-regulation of ATM protein by miR-18a. As ATM is a key enzyme in DNA damage repair, we evaluated the effect of miR-18a on DNA double-strand breaks. Ectopic expression of miR-18a significantly inhibited the repair of DNA damage induced by etoposide (p<0.001), leading to accumulation of DNA damage, increase in cell apoptosis and poor clonogenic survival.ConclusionmiR-18a attenuates cellular repair of DNA double-strand breaks by directly suppressing ATM, a key enzyme in DNA damage repair.

Role for DNA Damage Signaling in Pulmonary Arterial Hypertension

“…In brief, cells were treated with 1 μmol/L (micro M) etoposide for 1 hour and after withdrawal of etoposide, cells were incubated in fresh medium for 2 hours to allow DNA repair, as previously described. 25 The remaining DNA damage was then assessed (γ-H2AX). For ABT-888 treatments, cells were incubated with 10 μmol/L (micro M) ABT-888 for 48 hours before etoposide treatment, and ABT-888 was kept in medium for the recovery period.…”

MicroRNAs in Colorectal Cancer: from Diagnosis to Targeted Therapy

“…miR-143 was then found to be down-regulated in colon but not in rectum related cancer DOI:http://dx.doi.org/10.7314/APJCP.2014.15.17.6989 MicroRNAs in Colorectal Cancer: from Diagnosis to Targeted Therapies (Akao et al, 2006a, Xi et al, 2007, Vogelstein et al, 1988 NF2, CCND, miR-1 Down MET (Reid et al, 2012) miR-7 Down, up XRCC2, YY1,PAX6, (Motoyama et al, 2009, Xu et al, 2014, Zhang et al, 2013c, Li et al, 2014b) miR-9 Down α-Catenin (Cekaite et al, 2012, Zhu et al, 2012) miR-10b Down (Nishida et al, 2012, Burk et al, 2008, Zhu et al, 2012) miR-16 Down Cdx2 (Ma et al, 2013, Volinia et al, 2006, Tagawa et al, 2012 Up THBS1, MYC, CDKN1A,TMBIM1, E2F1 (Lanza et al, 2007, Arndt et al, 2009, Volinia et al, 2006, Motoyama et al, 2009, Diosdado et al, 2009, Cummins et al, 2006 Up ATM (Cummins et al, 2006, Sarver et al, 2009, Arndt et al, 2009, Wang et al, 2010, Wu et al, 2013a Up TF (Diosdado et al, 2009, Yu et al, 2013a, Bandres et al, 2006, Cummins et al, 2006 Up SFPQ and MYBL2 (Cummins et al, 2006, Arndt et al, 2009, Kurokawa et al, 2012 Up PTEN, TMP1 (Tan et al, 2013, Bovell et al, 2013, Earle et al, 2010, Yantiss et al, 2009…”

Radiosensitizing effects of miR‐18a‐5p on lung cancer stem‐like cells via downregulating both ATM and HIF‐1α

“…For example, miR‐18a‐5p enhanced radiosensitivity via downregulating ATM in cervical cancer,52 colorectal cancer,53 and breast cancer 54. It was addressed that miR‐18 targeted heat shock transcription factor 2 (HSF2),55 while HSF2 upregulated HIF‐1α 56.…”