Forschen Sci Open HUB for Scientific Research International Journal of Cancer Research and Molecular Mechanisms Open Access Copyright: © 2015 Ting T, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Volume: 1.2 Research Article RNA Interference-Mediated Knockdown on DKC1 Gene in Chemosensitized Colorectal Cancer Cell Lines Tammy Ting 1 , Rahman Jamal 1 , Roslan Harun 1,2 , Wan Zurinah Wan Ngah 1,3 and Norflza Mohd Mokhtar 1,4 * 1 UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Jalan Ya’acob Latiff, 56000 Cheras, Kuala Lumpur, Malaysia 2 Department of Medicine, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Malaysia 3 Department of Biochemistry, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Malaysia 4 Department of Physiology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Malaysia Received date: 09 June 2015; Accepted date: 24 June 2015; Published date: 27 June 2015. Citation: Ting T, Jamal R, Harun R, Wan Ngah WZ, Mokhtar NM (2015) RNA Interference-Mediated Knockdown on DKC1 Gene in Chemosensitized CRC Cell Lines. Int J Cancer Res Mol Mech 1(2): doi http:// dx.doi.org/10.16966/2381-3318.106 Copyright: © 2015 Ting T, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. * Corresponding author: Norflza Mohd Mokhtar, UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Jalan Ya’acob Latiff, 56000 Cheras, KL, Malaysia, Tel: 006 03 91718459; Fax:006 003 91717185; E-mail: norflza@ppukm.ukm.edu.my Abstract Background Dyskerin (DKC1) gene, a tolemerase ribonucleoprotein complex, has been reported to be up-regulated in various sporadic cancers, including colorectal cancer (CRC). This study was performed to investigate its potential as a therapeutic target for CRC. Materials and Methods The RNA interference (RNAi) technique was employed to down-regulate DKC1 expression in two human CRC cell lines, HCT116 and HT- 29, and followed by 5-fuorouracil (5-FU) treatment. Functional assays were carried out. The effcacy of RNAi was assessed by quantitative polymerase chain reaction (qPCR) and Western blotting. Results RNAi targeting DKC1 reduced mRNA and protein levels signifcantly after 48 and 72 hours post-transfection respectively with concomitant decrease in cell viability (P<0.05) in both cell lines. HCT116 cells were arrested in G1 phase of the cell cycle. 5-FU treatment following silencing further reduced cell viability and arrested HCT116 cells in G2 phase. RNAi treatment also reduced HCT116 cell migration signifcantly. Conclusion Silencing of DKC1 in combination with 5-FU may represent a good strategy to inhibit the CRC growth. Keywords: Colorectal cancer; RNA Interference; 5-Fluorouracil; DKC1 protein; Cell cycle Introduction Dyskerin is a highly conserved protein encoded by the DKC1 gene in eukaryotes [1]. It is present in small nucleolar ribonucleoprotein particles that have been shown to have pleiotropic functions for all basic cellular events such as protein expression, cell growth and cell proliferation [2]. Dyskerin is an integral component of the telomerase ribonucleoprotein complex and is required for the stabilization of the telomerase RNA component, normal telomerase activity and telomere maintenance [3]. It is also essential in rRNA processing and normal ribosome biogenesis by converting the specifc uridine residues of ribosomal RNA to pseudouridine [2]. Recently, its role in internal ribosome entry site (IRES)-mediated translation has also been reported [4].  Dyskerin expression is strongly correlated with active cell proliferation [5]. Its expression is up-regulated under experimental conditions that promotes cell growth and proliferation, and through oncogenic stimulation in breast [6] and colon cancers [7]. Recent studies have also identifed up- regulation of the DKC1 gene in association with hepatocellular carcinoma [8], oral squamous cell carcinoma [5] and prostate cancer [9]. Since up regulation of the DKC1 gene is associated with cell proliferation, the DKC1 gene can be a potential target for cancer therapy. Te fuoropyrimidine drug, 5-fuorouracil (5-FU), is widely used in CRC treatment since 1957 [10]. Its mechanism of action includes inhibition of thymidylate synthase, incorporation of its metabolites into RNA and DNA, and induction of cell cycle arrest and apoptosis [11]. However, the overall response rate for 5-FU in colorectal cancer (CRC) patients is low and depends on the DNA mismatch repair status [12]. Terefore, new treatment strategies to improve the efcacy of this drug as an anti-cancer agent are urgently needed. RNA interference (RNAi) is a process of sequence-specifc post- transcriptional gene silencing in a wide range of organisms and is initiated by double-stranded RNA that is homologous in sequence to the targeting gene [13]. To explore the potential of DKC1 as a novel therapeutic target, we applied siRNA targeting DKC1 to reduce its expression, followed by 5-FU treatment in CRC cell lines. Te aim of this study is to determine the efects of siRNA and the combination of siRNA with 5-FU treatment on chemosensitivity of tumour cells. ISSN 2381-3318