Long non coding RNAs lncRNAs which are longer than nt
Long non-coding RNAs (lncRNAs), which are longer than 200 nt without protein coding capacity, are reported to be regularly abnormal expression in multiple physiological and pathological processes . Based on the previous studies, the function of lncRNAs are various in cells including epigenetic control, nuclear import, Dalbavancin regulation, cellular differentiation, alternative splice, RNA decay and modulating RNA-binding proteins [5–7]. LncRNAs have been reported to take a significant role in the processes of gastric cancer [8–10]. To date, the
role of TONSL-AS1 in gastric cancer remains unclear.
The number of lncRNA ranges from about 20,000 to 100,000 in homo sapiens [11,12]. However, the biological function of the most lncRNAs remain elusive. Many studies reported that many lncRNAs could activate enhancers or promoters, but do not reveal sequence-specific functions. This phenomenon is further confirmed by the feature that most lncRNAs are localized in the nucleus with little sequence conservation and low expression levels [13,14]. Recent studies re-inforce the notion that lncRNA could regulate nearby gene by cis [15–17]. Thus, it is possible that TONSL-AS1 affect its locus gene TONSL's level through a cis manner.
Tonsoku like DNA repair protein (TONSL), also known as NFKBIL2, is thought to be a negative regulator of NF-kappa-B mediated tran-scription . TONSL may bind NF-κB complexes and trap them in the cytoplasm, preventing them from entering the nucleus and interacting with the DNA . TONSL was not only implicated in several physio-logical processes such as homologous recombination, but also involved in cancer pathogenesis [20–22]. For instance, overexpressed TONSL has
Abbreviations: Long non-coding RNA, lncRNA; Dulbecco's modified Eagle's medium, DMEM; fetal bovine serum, FBS
∗ Corresponding author. Department of General Surgery, The Affiliated Jiangsu Shengze Hospital of Nanjing Medical University. No.1399, Shichang Road, Shengze Town, Suzhou City, Jiangsu, China. E-mail address: [email protected] (Y. Ding).
1 These authors contributed equally to this work.
been observed in lung, esophageal and cervical cancer [20,22]. Fur-thermore, it was reported that knockdown of TONSL could significantly inhibit lung cancer cell growth . However, the role of TONSL in gastric cancer remain unclear. In the present study, we revealed that TONSL-AS1 inhibited cell proliferation, anchorage dependent cell growth and tumorigenesis. TONSL, a gene which is located around TONSL-AS1 on chromosome 8, was upregulated in TONSL-AS1 inducing cells. Luciferase activity assay indicated that TONSL-AS1 activated the TONSL promoter activity. In general, this study was aimed to reveal the potential roles of TONSL-AS1 in the gastric cancer, provided a new sight on how lncRNA in-volved in pathogenesis.
2. Materials and methods
2.1. Clinical samples
Fresh frozen gastric cancer tissues and their adjacent normal tissues were obtained from the First Affiliated Hospital of Nanjing Medical University. The samples were taken within 10 min after tumor excision, and immediately stored at −80 °C until application in the experiments. Written informed consents were acquired from all patients and this study was approved by the Ethics Committee of the hospital.
All cells were purchased from The Cell Bank of Type Culture Collection of Chinese Academy of Sciences, Shanghai, China. All cell lines were authenticated using short tandem repeat DNA profiling in January 2018. Human gastric cancer cell lines (SGC-7901, MGC-803) were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS), and 1% penicillin/ streptomycin (pen/strep). All cell lines were cultured in an atmosphere of 5% CO2 at 37 °C.
2.3. RNA extraction and quantitative real-time PCR (qRT-PCR)