LncRNA-LINC01089 inhibits lung adenocarcinoma cell proliferation and promotes apoptosis via sponging miR-543

Highlights

• LINC01089 indicated improved overall survival of LUAD patients.

• LINC01089 was low-expressed in LUAD.

• Upregulating LINC01089 inhibited LUAD cell proliferation and promoted apoptosis.

• MiR-543 was targeted by LINC01089, and high-expressed in LUAD tissues in LUAD.

• The effects of miR-543 and LINC01089 were mutually counteracted in LUAD.

Abstract

LINC01089, a newly discovered long non-coding RNA (lncRNA), has been reported to inhibit the progression of various types of cancers. This study aimed to characterize LINC01089 in the pathogenesis of lung adenocarcinoma (LUAD). LINC01089 expression in LUAD tissues or/and cells and its association with the overall survival of LUAD patients was analyzed in The Cancer Genome Atlas (TCGA)-LUAD database, by qRT-PCR or by Kaplan-Meier’s curve. Databases of StarBase, LncBase, and DEmiRNA were used to predict and confirm the interaction between LINC01089 and potential LINC01089-targeted microRNAs (miRNAs). The expressions of these miRNAs in LUAD tissues or/and cells were determined by qRT-PCR, and dual-luciferase reporter assay was performed to validate lncRNA-miRNA interaction. The expressions of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax) and Cleaved caspase-3 in LUAD cells were analyzed by Western blot. LINC01089 improved overall survival of LUAD patients and was low-expressed in LUAD. Upregulating LINC01089 expression reduced LUAD cell viability, inhibited colony formation, enhanced apoptosis, accompanied by downregulated Bcl-2 and miR-543 and upregulated Bax and Cleaved caspase-3. MiR-543 was determined as a target gene of LINC01089, and was high-expressed in LUAD tissues. Upregulating miR-543 expression induced the opposite effects to LINC01089 upregulation on these cellular biological behaviors and the expressions of Bcl-2, Bax and Cleaved caspase-3. Moreover, the effects of miR-543 upregulation and LINC01089 upregulation were mutually counteracted by each other. LINC01089 inhibited lung adenocarcinoma cell proliferation and promoted apoptosis via sponging miR-543.

Introduction

Lung cancer is one of the most prevalent malignancies and a leading cause of death in the US, with an estimated incidence around 13 % and death rates of 23 % in 2020 (Siegel et al., 2020). Moreover, in the US, cancer-related death caused by lung cancer was notably more than that caused by other cancers in 2017 (Siegel et al., 2020). Lung cancer can be histologically divided into small cell lung carcinoma (SCLC), and non-small-cell lung carcinoma (NSCLC), which accounts for 85 % of all the types (Alghamdi et al., 2018; Sher et al., 2008). Lung adenocarcinoma (LUAD) is a subtype of NSCLC, and the most commonly diagnosed type of lung cancer, with an incidence of 40 % (Denisenko et al., 2018). LUAD develops from type II alveolar cells, which consists of small airway epithelium, and can secrete mucus and restore epithelium after lung injury (Mason, 2006; Noguchi et al., 1995). Through high-resolution computed tomography (CT), LUAD can be preliminarily detected with the typical presentation of pure or ground-glass nodules (GGNs) (Higashi et al., 1998; Huang et al., 2012; Wu et al., 2015). However, GGNs also show alveolar inflammation and hemorrhage (benign lesions) (Scafoglio et al., 2018). Thus, biomarkers to enhance the precision of LUAD diagnosis as well as to reduce mortality are urgently desired.

Long non-coding RNAs (lncRNAs), a class of RNAs endogenously existing in cells with more than 200 nucleotides, participate in various biological processes, including cancer occurrence and progression (Bhan et al., 2017). LncRNAs were subjected to common post-transcriptional modifications, including 5’-capping, 3’-polyadenylation, and splicing (Quinn and Chang, 2016). LncRNAs may serve as signals to promote transcription, or as decoys to repress transcription, or as epigenetic regulators, or as scaffolds to interact with various protein partners to form ribonucleoprotein complexes (Mercer et al., 2009; W. X. Peng et al., 2017). The prominent classes of lncRNAs share a common functionality in their ability to regulate gene expression through sponging miRNAs, and such a phenomenon refers to the hypothesis of competing endogenous RNA (ceRNA) (Salmena et al., 2011). cBioPortal analysis showed that lncRNA alteration frequencies are high in LUAD, with 625 lncRNAs presenting alteration rates over 1% and 36 over 10 %, moreover, these alterations of lncRNAs are associated with overall survival of LUAD patients (B. Liu et al., 2017). Reversely regulating LUAD-altered lncRNA expression could inhibit LUAD progression through modulating LUAD cell biological behaviors, such as apoptosis (Dong et al., 2018), proliferation and viability (Liu et al., 2019b), and migration and invasion (Z. Peng et al., 2018). LINC01089, a lncRNA located at chromosome 12, also refers to as LncRNA Inhibiting Metastasis (LIMT), is previously found to be low-expressed after epidermal growth factor (EGF) stimulation in MCF-10A mammary cells. Study showed that low-expressed LIMT promotes migration and invasion, and predicts poor prognosis in breast cancer (BC) patients (Sas-Chen et al., 2016). Overexpressed LINC01089 in BC inhibits motility in vitro and retards metastasis in animal models (Sas-Chen et al., 2016). The tumor suppressing effect of LINC01089 is also reported in colorectal cancer (CRC) (M. Li and Guo, 2020), gastric cancer (GC) (F. Wang and Yang, 2020) and cervical cancer (CC) (S. Li et al., 2020). Moreover, the interaction between lncRNAs and microRNAs (miRNAs), that is, the competitive binding of lncRNAs to its targeted miRNA is implicated in the tumor-suppressing activities (Tang et al., 2019). However, the expression of LINC01089 and its effect on LUAD remained unclear.

The objective of this study was to assess the expression of LINC01089 in LUAD, and to investigate the modulation of LINC01089 on LUAD cell biological behaviors in LUAD progression. Moreover, we also determined the miRNA through which LINC01089 exerted the anti-LUAD effects.